Methods and compositions for transfecting cells

Abstract

Provided herein are methods and compositions for transfecting cells or an organism comprising the cells for use as therapeutic, medicinal products, or consumption products or generating thereof. The compositions may comprise a saccharide, a nucleic acid molecule, and / or a lipid molecule. The compositions and methods may promote or facilitate uptake of a nucleic acid by the cells.

Description

METHODS AND COMPOSITIONS FOR TRANSFECTING CEEESBACKGROUND

[0001] Transfection can introduce nucleic acid molecules into eukaryotic cells. The nucleic acid molecule being transfected into the cells may promote changes in the structural and / or functional properties of the transfected cells or the progenies thereof. The cells with different structural and / or functional properties may be used in a wide variety of purposes, including but not limited to food production, diagnostics production, biologic drug production, cell therapies, virus production, biosensors, tissue engineering, or in drug discovery.SUMMARY

[0002] Wide-spread or industrial-scale application of cell for variety of purposes, including but not limited to food production, diagnostics’ production, biologic drug production, cell therapies, virus production, biosensors, tissue engineering, or in drug discovery entails efficient transfection of nucleic acids into cells. The cells generated using the methods, compositions, or kits may have desirable expression level of the transfected nucleic acids. Provided herein are methods, compositions, and / or kits for transfection that are highly efficient and able to produce high and stable gene expression. The methods, compositions, or kits disclosed herein can be broadly applicable for a wide variety of purposes, cell types, and / or conditions without the need of using of animal derived materials or genetic modifications. The methods, compositions, or kits disclosed herein may also avoid causing immunogenic reaction and / or cancer. The methods, compositions, or kits disclosed herein may have a low cytotoxicity. The methods, compositions, or kits disclosed herein may not use animal-derived products but still provide a high transfection efficiency. The methods, compositions, or kits disclosed herein may use carriers that are biodegradable or biocompatible. The methods, compositions, or kits disclosed herein may allow for large payloads (e.g., the size of the transfected nucleic acids) and / or entail a low cost to practice / manufacture. The methods, compositions, or kits disclosed herein may also allow delivery of nucleic acids into the cells of an organism in vivo. Such delivery be tissue / cell-type specific. These methods, compositions, or kits may thus overcome the limitations imposed by currently available viral or non-viral transfection and facilitate wide-spread or industrial-scale applications of transfections of cells.

[0003] Provided herein, are methods. In an aspect, a method comprises: (a) providing a cell with a composition comprising: (i) a nucleic acid molecule, and (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or anycombination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); and (b) delivering the composition into the cell.

[0004] In some embodiments, the saccharide molecule comprises the sorbitan, the mannide, the trehalose, the functional variant thereof, the derivative thereof, or any combination thereof. In some embodiments, the saccharide molecule comprises a second ester group. In some embodiments, the saccharide molecule comprises the lactose comprising the ester group. In some embodiments, the composition further comprises a lipid molecule.

[0005] Provided herein, are methods. In an aspect, a method comprises: (a) providing a cell with a composition comprising: (i) a nucleic acid molecule, (ii) a saccharide molecule and / or a lipid molecule; and (iii) a polar organic solvent miscible with water; and (b) delivering the composition into the cell.

[0006] In some embodiments, the polar organic solvent is at most about 16 %, by volume, of the composition. In some embodiments, the polar organic solvent is at most about 8 %, by volume, of the composition. In some embodiments, the polar organic solvent is DMSO, DMF, ethanol, propanol, 2-propanol, ethylene glycol, a functional variant thereof, a derivative thereof, or any combination thereof. In some embodiments, the polar organic solvent is DMSO. In some embodiments, the polar organic solvent is ethanol. In some embodiments, the composition comprises an alcohol-based solvent. In some embodiments, the saccharide molecule comprises a monosaccharide. In some embodiments, the saccharide molecule comprises a disaccharide. In some embodiments, the saccharide molecule comprises an oligosaccharide. In some embodiments, the saccharide molecule comprises a polysaccharide. In some embodiments, the composition comprises the saccharide molecule and the lipid molecule. In some embodiments, the lipid molecule is cationic in an aqueous neutral solution. In some embodiments, the lipid molecule comprises DOTAP. In some embodiments, the lipid molecule comprises a lipid molecule having a structure of formula (I), (II), or (III). In some embodiments, the lipid molecule is selected from the group consisting of a compound of formula (IV)-(IX). In some embodiments, the saccharide molecule is neutral in an aqueous neutral solution. In some embodiments, the saccharide molecule comprises sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate, a derivative thereof, or any combination thereof. In some embodiments, the saccharide molecule comprises a lipid moiety. In some embodiments, the lipid moiety comprises a fatty acyl. In some embodiments, the fatty acyl is monounsaturated. In some embodiments, the fatty acyl is polyunsaturated. In someembodiments, the lipid moiety comprises a fatty hydrocarbon chain. In some embodiments, the fatty hydrocarbon chain is saturated. In some embodiments, the fatty hydrocarbon chain is monounsaturated. In some embodiments, the fatty hydrocarbon chain is polyunsaturated. In some embodiments, the nucleic acid molecule comprises a ribonucleic acid (RNA); optionally wherein the RNA comprises a messenger RNA (mRNA), a self-amplifying RNA (saRNA), a small interfering RNA (siRNA), a transfer RNA (tRNA), a small activating RNA (RNAa), an endless / circular RNA (eRNA), a complementary DNA (cDNA), a plasmid DNA (pDNA), or any combination thereof.

[0007] Provided herein, are compositions. In an aspect, a composition comprises: a lipid molecule having a structure of formula (I), formula (II), or formulawherein: R1is independently C6-C22 alkyl,C6-C22 alkenyl, or C6-C22 alkynyl; each of R2and R3is independently H, C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl; R4is C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl; R5is independentlyC1-C4 alkyl; and X is independently an anionic radical selected from the group consisting of: hydroxide, halide, sulfate, methylsulfate, ethylsulfate, and phosphate ions.

[0008] In some embodiments, the lipid molecule is selected from the group consisting of:embodiments, the composition further comprises a nucleic acid molecule. In some embodiments, the composition further comprises a saccharide molecule. In some embodiments, the saccharide molecule comprises a monosaccharide. In some embodiments, the saccharide molecule comprises a disaccharide. In some embodiments, the saccharide molecule comprises an oligosaccharide. In some embodiments, the saccharide molecule comprises a polysaccharide. In some embodiments, the saccharide molecule is neutral in an aqueous neutral solution. In some embodiments, the saccharide molecule comprises a sorbitan, a mannide, a trehalose, a lactose, a functional variant thereof, a derivative thereof, or any combination thereof; optionally wherein the saccharide molecule comprises sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate, a derivative thereof, or any combination thereof. In some embodiments, the saccharide molecule comprises a lipid moiety. In some embodiments, the lipid moiety comprises a fatty acyl. In some embodiments, the fatty acyl is monounsaturated. In some embodiments, the fatty acyl is polyunsaturated. In some embodiments, the lipid moiety comprises a fatty hydrocarbon chain. In some embodiments, the fatty hydrocarbon chain is saturated. In some embodiments, the fatty hydrocarbon chain is monounsaturated. In some embodiments, the fatty hydrocarbon chain is polyunsaturated. In some embodiments, the nucleic acid molecule comprises a ribonucleic acid (RNA); or wherein the nucleic acid molecule comprises a messenger RNA (mRNA), a selfamplifying RNA (saRNA), a small interfering RNA (siRNA), a transfer RNA (tRNA), a small activating RNA (RNA), an endl ess / circular RNA (eRNA), a complementary DNA (cDNA), a plasmid DNA (pDNA), or any combination thereof.

[0009] Provided herein, are methods. In an aspect, a method comprises: (a) providing a cell with the composition as described herein; and (b) delivering the composition into the cell.

[0010] In some embodiments, the providing in (a) comprises contacting the composition with an organism comprising the cell. In some embodiments, the organism comprises an animal. In some embodiments, the animal comprises a human. In some embodiments, the animal comprises a non-human animal. In some embodiments, the non-human animal comprises a rodent. In some embodiments, the providing in (a) comprises contacting the cell that is cultured in a culture medium. In some embodiments, further comprising converting the cell or a progeny thereof into a meat product. In some embodiments, further comprising generating a therapeutic product using the cell or a progeny thereof.

[0011] In some embodiments, the composition further comprises a second lipid molecule different from the lipid molecule. In some embodiments, the second lipid molecule is a structural lipid molecule. In some embodiments, the second lipid molecule comprises a l,2-di-(9Z-octadecenoyl)- sn-glycero-3 -phosphoethanolamine (DOPE); optionally wherein a ratio of the nucleic acid molecule and the second lipid molecule is at least about 0.1, at least about 0.5, at least about 1, at least about 2, at least about 4, at least about 6, at least about 8, at least about 10, or at least about 20. In some embodiments, further comprising, generating a nanoparticle molecule comprising the nucleic acid molecule, the saccharide molecule, the lipid molecule, and the second lipid molecule. In some embodiments, a ratio of the nucleic acid molecule and the saccharide molecule, by weight, is at least about 0.1, at least about 0.5, at least about 1, at least about 2, at least about 4, at least about 6, at least about 8, at least about 10, at least about 12, at least about 14, at least about 16, at least about 20, at least about 30, at least about 40, at least about 50, or at least about 100. In some embodiments, a ratio of the nucleic acid molecule and the lipid molecule is, by weight, is at least about 0.1 , at least about 0.5, at least about 1, at least about 2, at least about 4, at least about 6, at least about 8, at least about 10, at least about 12, at least about 14, at least about 16, at least about 20, at least about 30, at least about 40, at least about 50, or at least about 100.

[0012] Provided herein, are compositions for use in a method of treating a disease. In an aspect, a composition for use in a method of treating a disease comprises: (i) a nucleic acid molecule, wherein the nucleic acid molecule is a therapeutic nucleic acid molecule, and (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2).

[0013] In some embodiments, further comprising (iii) a cationic lipid molecule. In some embodiments, the cationic lipid molecule is selected from the group consisting of DOTAP and any compound of Formula (I)-(IX). In some embodiments, further comprising a structural lipid. In some embodiments, the structural lipid comprises DOPE.

[0014] Additional aspects and advantages of the present disclosure will become readily apparent to those skilled in this art from the following detailed description, wherein only illustrativeembodiments of the present disclosure are shown and described. As will be realized, the present disclosure is capable of other and different embodiments, and its several details are capable of modifications in various obvious respects, all without departing from the disclosure. Accordingly, the drawings and description are to be regarded as illustrative in nature, and not as restrictive.INCORPORATION BY REFERENCE

[0015] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. To the extent publications and patents or patent applications incorporated by reference contradict the disclosure contained in the specification, the specification is intended to supersede and / or take precedence over any such contradictory material.BRIEF DESCRIPTION OF THE DRAWINGS

[0016] The novel features of the disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the disclosure are utilized, and the accompanying drawings (also “Figure” and “FIG.” herein), of which:

[0017] FIG. 1A depicts an image showing green fluorescent protein (GFP) expression after selfamplifying RNA (saRNA) transfection with N-(l-(2,3-dioleoyloxy) propyl)-N,N,N- trimethylanunonium chloride (DOTAP) (saRNA / DOTAP mass ratio 1 / 4). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0018] FIG. IB depicts an image showing green fluorescent protein (GFP) expression after saRNA transfection with DOTAP / Sorbitan trioleate (STO) (saRNA / DOTAP / STO mass ratio 1 / 4 / 10). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0019] FIG. 1C depicts an image showing GFP expression after saRNA transfection with DOTAP / Sorbitan trioleate (SMO) (saRNA / DOTAP / SMO mass ratio 1 / 4 / 10). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0020] FIG. ID depicts DIC image (left) and image showing GFP expression (right) of 3- dimensional (3D) cultured cells after saRNA transfection with DOTAP / Sorbitan monooleate (SMO)(saRNA / DOTAP / SMO mass ratio 1 / 2 / 40). The black and white image is obtained by converting green fluorescence into white and grey shades in the right image.

[0021] FIG. IE shows that alcohol-based solvent (ethanol) can increase the transfection efficiency of the transfection complex in 3D culture. The bar graph shows the transfection efficiency of transfection complexes comprising DOTAP / SMO (saRNA / DOTAP / SMO mass ratio 1 / 2 / 40) in DMSO or ethanol as solvent quantified as percent of GFP-positive cells by flow cytometry.

[0022] FIG. 2A depicts images showing a comparison of transfection efficiency of a first exemplary transfection complex (with the same saccharide / cationic lipid) with or without a helper lipid. The black and white image is obtained by converting green fluorescence into white and grey shades in the right image.

[0023] FIG. 2B depicts images showing a comparison of transfection efficiency of a second exemplary transfection complex (with the same saccharide / cationic lipid) with or without a helper lipid. The black and white image is obtained by converting green fluorescence into white and grey shades in the right image.

[0024] FIG. 2C shows that alcohol-based solvent (ethanol) can increase the transfection efficiency of the transfection complex in 3D culture. The bar graph shows the transfection efficiency of transfection complexes comprising DOTAP / SMO / DOPE (saRNA / DOTAP / SMO / DOPE mass ratio 1 / 2 / 40 / 2) in different solvents (DMSO, ethanol, or JEtMessenger) quantified as percent of GFP- positive cells by flow cytometry.

[0025] FIG. 3A depicts an image showing that myosin heavy chain (MyHC) expression was observed in 3D cultured cells after myoblast determination protein 1 (MyoD)-saRNA transfection using DOTAP / DOPE / SMO (saRNA / DOTAP / SMO / DOPE mass ratio 1 / 2 / 40 / 2), detected by fluorescence microscopy of cells stained with MyHC antibody. The black and white image is obtained by converting fluorescence into white and grey shades.

[0026] FIG. 3B depicts images comparing GFP expression after transfection of plasmid deoxynucleic acid (DNA) (pDNA) in 2D culture using DOTAP / DOPE / SMO (saRNA / DOTAP / SMO / DOPE mass ratio 1 / 2 / 40 / 2) and lipofectamine. Black and white image of fluorescence microscopy with green fluorescence converted into white and grey shades.

[0027] FIG. 4 depicts an image showing GFP expression after saRNA transfection in 3D using the lipid of formular (IV) / DOPE / SMO (saRNA / cationic lipid / SMO / DOPE mass ratio 1 / 2 / 40 / 2). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0028] FIG. 5A depicts an image showing GFP expression after saRNA transfection in 3D using the lipid of formular (IV) / DOPE / SMO (saRNA / lipid / SMO / DOPE mass ratio 1 / 4 / 32 / 8). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0029] FIG. 5B depicts an image showing GFP expression after saRNA transfection in 3D using the lipid of formular (V)DOPE / SMO (saRNA / lipid / DOPE / SMO mass ratio 1 / 4 / 6 / 24). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0030] FIG. 5C depicts an image showing GFP expression after saRNA transfection in 3D using the lipid of formular (VI) / DOPE / SMO (saRNA / lipid / DOPE / SMO mass ratio 1 / 5.3 / 6.7 / 16). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0031] FIG. 5D depicts an image showing GFP expression after saRNA transfection in 3D using the lipid of formular (VII) / DOPE / SMO (saRNA / lipid / DOPE / SMO mass ratio 1 / 5.3 / 6.7 / 16). The black and white image is obtained by converting green fluorescence into white and grey shades.

[0032] FIG. 6A shows the 1H NMR spectra of a synthesized fourth exemplary lipid.

[0033] FIG. 6B shows the 1H NMR spectra of a synthesized fifth exemplary lipid.

[0034] FIG. 7 depicts a schematic workflow for transfecting cells with compositions comprising saccharide and / or lipid molecules as described and nucleic acid molecules.

[0035] FIG. 8 depicts a schematic workflow for transfecting cells with compositions comprising saccharide and / or lipid molecules as described and nucleic acid molecules in vivo.

[0036] FIG. 9A shows in vivo image analysis (IVIS) of transection of cells using the methods described herein. The left panel shows the radiance measured from animals at timepoints 6 hours and 24 hours post transfection. The right panel shows the same values in logarithmic scale and statistical differences are calculated. In each group n=4, statistical analysis: 2way ANOVA, Tukey’s multiple comparison test, ** p < 0.01, *** p < 0.0001.

[0037] FIG. 9B shows in vivo image analysis (IVIS) of transection of cells using the methods described herein measured from all ex vivo tissues. The left panel shows the radiance measured from the ex vivo tissues. The right panel shows the same values in logarithmic scale and statistical differences are calculated. In each group n=4, statistical analysis: 2way ANOVA, Tukey’s multiple comparison test, *p< 0.05, ** p < 0.01, *** p < 0.0001.

[0038] FIG. 10 shows knockdown of GAPDH mRNA levels in HEK-293 cells following delivery of saRNA[GFP] (transfection control) and / or 50 pM siRNA [GAPDH] using Lipofectamine2000 (transfection control) or the delivery composition, comprising a cationic lipid and a sorbitan polysaccharide. Analysis by qPCR.

[0039] FIG 11. shows knockdown of GAPDH mRNA levels in three human cell lines (A549, MRC- 5, and HEK-293) following delivery of 50 pM siRNA [GAPDH] using Lipofectamine2000 (transfection control) or the delivery composition, comprising a cationic lipid and a sorbitan polysaccharide. Control cells were not transfected. Analysis by qPCR.

[0040] While various embodiments of the disclosure have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions may occur to those skilled in the art without departing from the disclosure. It should be understood that various alternatives to the embodiments of the disclosure described herein may be employed.DETAILED DESCRIPTIONCompositions and Methods for TransfectionOverview

[0041] Transfection is a process to introduce exogenous / foreign nucleic acid molecules into eukaryotic cells. The nucleic acid molecule being transfected into the cells may promote changes in the structural and / or functional properties of the transfected cells or the progenies thereof. The cells with different structural and / or functional properties may be used in a wide variety of purposes, including but not limited to food production, diagnostics’ production, biologic drug production, cell therapies, virus production, biosensors, tissue engineering, or in drug discovery.

[0042] Transfection of nucleic acids can comprise stable and transient transfections. Stable transfection can comprise those that allows for long-term expression of a transgene by integrating foreign nucleic acids (such as deoxyribonucleic acid (DNA)) into the host nuclear genome or maintaining an episomal vector in the host nucleus as an extra-chromosomal element. The transgene may then be constitutively expressed even with the replication of cells. Transient transfection may not require integrating nucleic acids into the host cell genome. Nucleic acids may be transfected in the form of a plasmid or as oligonucleotides.

[0043] Nucleic acids may be packaged into a transfection complex (for example, the lipoplex as described herein) with various components for facilitating the transfection. The transfection complex may facilitate the transfection via various physical or chemical properties of the components. Such components can comprise nucleic acid vector or plasmids; chemical group or entities such as lipid molecules, saccharide molecules, or a combination thereof; or both.

[0044] Currently available transfection methods can be limited due to a lack of effectiveness, scalability, or reliability. Currently available transfection methods may not be applicable at industrial or manufacturing scales. Currently available transfection methods can be classified as viral or non- viral methods. Non-viral transfection methods using non-viral vectors (e.g., chemical, polymer, or lipid based) can result in high cytotoxicity and a high percentage of cell loss. Non-viral transfection methods can entail a high cost to practice. Non-viral vectors may suffer from low transfection efficiency and undesirable gene expression of the nucleic acids being introduced into the cells. Additionally, non-viral transfection methods may entail a narrow range of physiochemical conditions for transfection complex formation that limit the transfection efficiency, as opposed to animal-derived components. Non-viral transfection may also use carriers that are not biodegradable or biocompatible, which can limit the application of the transfected cells, for example, for consumption purposes. Viral methods using viral vectors may generate immunogenic responses against the viral vectors. The viral vectors may be carcinogenic. The viral vectors may also only allow for small payloads (e.g., the size of the transfected nucleic acids) and / or entail a high cost to manufacture. In some cases, even certain existing methods may allow delivery of the nucleic acids into the cells of an organism in vivo, such delivery may not be effective enough for certain purposes (such as achieving a therapeutic effect using the nucleic acid). In some cases, such delivery may not have sufficient tissue / cell-type specificity. Hence, current transfection methods, viral or non-viral based, are limited for their wide-spread or industrial-scale applications. The methods, compositions, or kits described herein have the beneficial advantages of circumventing the issues described herein.

[0045] Provided herein, are methods and compositions for transfecting nucleic acids into cells. The methods may comprise contacting a cell with a composition as described herein. In some cases, the composition may comprise a saccharide molecule, a lipid molecule, or a combination thereof; as described herein. The composition may further comprise a solvent. The composition may further comprise a nucleic acid molecule. The composition comprising the saccharide molecule, the lipid molecule, the solvent, the nucleic acid molecule, or the combination thereof may form a transfection complex for facilitating the transfection of the nucleic acid into a cell. Various components of the transfection complex may be maintained as a particular ratio, as described herein, to obtain a desirable transfection efficiency. The solvent may comprise an organic solvent. The solvent may comprise a polar organic solvent miscible with water. A cell taking up the nucleic acid may undergo alternations in protein expression that result in differentiation, growth, maintenance of viability, or a combination thereof.

[0046] The efficiency of a cell taking up a nucleic acid molecule using traditional methods may be low. The low efficiency of the cell taking up the nucleic acid may limit the applicability of the methods and compositions for differentiating the cell. The lack of biodegradability of non-viral vectors used in traditional methods may limit the applicability of the methods and compositions for altering and / or changing the protein expression and may limit cell differentiation. The instant disclosure provides methods and compositions for increasing the transfection efficiency, thereby reducing the amount of nucleic acid molecules, cells, or other reagents to transfect the cells with the nucleic acid molecules.

[0047] A transfected cell may alter protein or gene expression. A transfected cell may undergo differentiation. A differentiated cell may have a different structural and / or functional characteristic relative to a cell that is not differentiated. In some cases, a cell with a different structural and / or functional characteristic may be used in a wide variety of purposes. In some cases, a cell with a different structural and / or functional characteristic may be edible. In some cases, a cell with a different structural and / or functional characteristic may be used to produce an edible meat product.

[0048] A saccharide or lipid of a composition described herein may be biodegradable. Biodegradable saccharides or lipids can be metabolized by various enzymes (e.g., esterase, peptidases) to minimize any toxicity, immunogenicity, and / or carcinogenicity.

[0049] Additionally, currently available transfection complex may comprise using solvent injection in which lipid-based transfection complex is prepared by rapidly injecting a solution of solid lipids in water-miscible solvents or a water-miscible solvent mixture into water. A component of the complex can comprise a cationic substance or ionizable substance generating positive charge at pH below physiological pH. This cationic substance can be a cationic polymer or lipid. DOTAP chloride (N-[l-(2,3-Dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride) is traditionally used as the cationic substance. However, DOTAP is expensive to obtain and thus limits the scale and application of the products generated from transfections. Provided herein are cationic substances that are not cost-prohibitive to obtain and thus facilitates the scale and application of the products generated from transfections. Furthermore, provided herein are additional components for generating the transfection complex that can increase the transfection efficiency of the nucleic acids. One such component can comprise an alcohol-based solvent (such as ethanol). Additionally, provided herein are dosages for individual components of the transfection complex that can provide beneficial transfection efficiency of the nucleic acids, thus also limiting the cost of generating the reagents andfacilitating the application of the transfection. Also provided herein are methods for generating the components of the complex and the complex.Saccharide

[0050] In some cases, the composition may comprise a saccharide molecule. A saccharide molecule may facilitate the uptake of the nucleic acid by binding to the glucose transporter (e.g., GLUT1 or GLUT4) or other glucose receptors. Once the saccharide molecule of the transfection complex binds to the transporter or receptor on a cell, the transfection complex can be collectively translocated across the cell membrane of the cell using the transporter or receptor. The saccharide may also facilitate the transfection by interacting with other components of the cell membrane. Additionally, saccharide may facilitate the transfection by serving as the structural component of the transfection complex or facilitate the formation of the complex.

[0051] In some instances, a saccharide may comprise a carbohydrate. In some instances, a saccharide may comprise a carbohydrate moiety. A saccharide may be a sugar. A saccharide may comprise a sugar moiety. A saccharide may comprise a carbohydrate monomer or a monosaccharide. In some cases, a saccharide may comprise a polymeric form of monosaccharides. The monosaccharides of a polymeric form of a saccharide may be linked by glycosidic bonds. In some cases, a polymeric form of saccharide may comprise a disaccharide, an oligosaccharide, a polysaccharide, or a combination thereof. In some cases, a saccharide may comprise at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000 or more carbohydrate moieties. In some cases, a saccharide may comprise at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000, or 100000 carbohydrate moieties. In some cases, a saccharide may comprise at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000 or more monosaccharides. In some cases, a saccharide may comprise at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 50, 100, 150, 200, 250, 300, 350, 400, 450, 500, 1000, 2000, 3000, 4000, 5000, 6000, 7000, 8000, 9000, 10000, or 100000 monosaccharides.

[0052] In some instances, a saccharide may be a monosaccharide, disaccharide, oligosaccharide, or polysaccharide. In some instances, a saccharide may be a monosaccharide. In some instances, a saccharide may be a disaccharide. In some instances, a saccharide may be an oligosaccharide. In some instances, a saccharide may be a polysaccharide.

[0053] In some instances, a saccharide may be functionalized or derivatized with various moieties. The term “derivative” or a grammatically equivalent term, when referring to a saccharide, can mean a modified saccharide modified via functionalization or derivatization of the saccharide with various chemical moieties.

[0054] A saccharide or a derivative thereof may have cationic moieties. A saccharide or a derivative thereof may have a cationic charge in an aqueous solution. A saccharide or a derivative thereof may have a cationic charge in a neutral solution. A saccharide or a derivative thereof may have a neutral charge in an aqueous solution. A saccharide or a derivative thereof may have a neutral charge in a neutral solution. A saccharide or a derivative thereof may have an anionic charge in an aqueous solution. A saccharide or a derivative thereof may have an anionic charge in a neutral solution. A saccharide or a derivative thereof may have a cationic charge in an aqueous solution. A saccharide or a derivative thereof may have a cationic charge in a neutral solution. A saccharide or a derivative thereof may have a cationic charge in an aqueous neutral solution. A saccharide or a derivative have a neutral charge in an aqueous neutral solution. A saccharide or a derivative thereof may have an anionic charge in an aqueous neutral solution.

[0055] A corresponding cationic charge density of cationic moieties may be measured in mequivalent of basic group per gram of the saccharide molecule (mequiv / g). In some cases, the cationic charge density of cationic moieties may be determined by the polyelectrolyte titration method, conductometric titration, and / or acid / base titration. In some cases, the corresponding cationic charge density of a saccharide or a derivative thereof may be at least about 0.5 mequiv / g, at least about 1 mequiv / g, at least about 1.5 mequiv / g, at least about 2 mequiv / g, at least about 2.5 mequiv / g, at least about 3 mequiv / g, at least about 3.5 mequiv / g, at least about 4 mequiv / g, at least about 4.5 mequiv / g, at least about 5 mequiv / g, at least about 5.5 mequiv / g, at least about 6 mequiv / g, at least about 6.5 mequiv / g, at least about 7 mequiv / g, at least about 7.5 mequiv / g, at least about 8 mequiv / g, at least about 8.5 mequiv / g, at least about 9 mequiv / g, at least about 9.5 mequiv / g, at least about 10 mequiv / g, at least about 10.5 mequiv / g, at least about 11 mequiv / g, at least about 11.5 mequiv / g, at least about 12 mequiv / g, at least about 12.5 mequiv / g, at least about 13 mequiv / g, at least about 13.5 mequiv / g, at least about 14 mequiv / g, at least about 14.5 mequiv / g, at least about 15 mequiv / g, at least about 15.5 mequiv / g, at least about 16 mequiv / g, at least about 16.5 mequiv / g, at least about 17 mequiv / g, at least about 17.5 mequiv / g, at least about 18 mequiv / g, at least about 18.5 mequiv / g, at least about 19 mequiv / g, at least about 19.5 mequiv / g, or at least about 20 mequiv / g. In some cases, the corresponding cationic charge density of a saccharide or a derivative thereof may beat most about 0.5 mequiv / g, at most about 1 mequiv / g, at most about 1.5 mequiv / g, at most about 2 mequiv / g, at most about 2.5 mequiv / g, at most about 3 mequiv / g, at most about 3.5 mequiv / g, at most about 4 mequiv / g, at most about 4.5 mequiv / g, at most about 5 mequiv / g, at most about 5.5 mequiv / g, at most about 6 mequiv / g, at most about 6.5 mequiv / g, at most about 7 mequiv / g, at most about 7.5 mequiv / g, at most about 8 mequiv / g, at most about 8.5 mequiv / g, at most about 9 mequiv / g, at most about 9.5 mequiv / g, at most about 10 mequiv / g, at most about 10.5 mequiv / g, at most about 11 mequiv / g, at most about 11.5 mequiv / g, at most about 12 mequiv / g, at most about12.5 mequiv / g, at most about 13 mequiv / g, at most about 13.5 mequiv / g, at most about 14 mequiv / g, at most about 14.5 mequiv / g, at most about 15 mequiv / g, at most about 15.5 mequiv / g, at most about 16 mequiv / g, at most about 16.5 mequiv / g, at most about 17 mequiv / g, at most about 17.5 mequiv / g, at most about 18 mequiv / g, at most about 18.5 mequiv / g, at most about 19 mequiv / g, at most about19.5 mequiv / g, or at most about 20 mequiv / g. In some cases, the corresponding cationic charge density of a saccharide or a derivative thereof may be from 0.005 to 2000 mequiv / g. In some cases, the corresponding cationic charge density of cationic moieties of a saccharide or a derivative thereof may be from 0.05 to 200 mequiv / g. In some cases, the corresponding cationic charge density of cationic moieties of a saccharide or a derivative thereof may be from 0.5 to 20 mequiv / g.

[0056] In some instances, a saccharide or a derivative thereof may comprise an average molecular mass of at least about 0.05 kilodaltons (kDa), at least about 0.1 kDa, at least about 0.2 kDa, at least about 0.3 kDa, at least about 0.4 kDa, at least about 0.5 kDa, at least about 0.6 kDa, at least about 0.7 kDa, at least about 0.8 kDa, at least about 0.9 kDa, at least about 1 kDa, at least about 2 kDa, at least about 3 kDa, at least about 4 kDa, at least about 5 kDa, at least about 6 kDa, at least about 7 kDa, at least about 8 kDa, at least about 9 kDa, at least about 10 kDa, at least about 20 kDa, at least about 30 kDa, at least about 40 kDa, at least about 50 kDa, at least about 60 kDa, at least about 80 kDa, at least about 100 kDa, at least about 120 kDa, at least about 140 kDa, at least about 160 kDa, at least about 180 kDa, at least about 200 kDa, at least about 220 kDa, at least about 240 kDa, at least about 260 kDa, at least about 280 kDa, at least about 300 kDa, at least about 350 kDa, at least about 400 kDa, at least about 450 kDa, at least about 500 kDa, at least about 550 kDa, at least about 600 kDa, at least about 650 kDa, at least about 700 kDa, at least about 750 kDa, at least about 800 kDa, at least about 850 kDa, at least about 900 kDa, at least about 950 kDa, at least about 1000 kDa, at least about 1100 kDa, at least about 1200 kDa, at least about 1300 kDa, at least about 1400 kDa, at least about 1500 kDa, at least about 1600 kDa, at least about 1700 kDa, at least about 1800 kDa, at least about 1900 kDa, at least about 2000 kDa or more. In some instances, a saccharide or a derivativethereof may comprise an average molecular mass of at most about 0.05 kilodaltons (kDa), at most about 0.1 kDa, at most about 0.2 kDa, at most about 0.3 kDa, at most about 0.4 kDa, at most about 0.5 kDa, at most about 0.6 kDa, at most about 0.7 kDa, at most about 0.8 kDa, at most about 0.9 kDa, at most about 1 kDa, at most about 2 kDa, at most about 3 kDa, at most about 4 kDa, at most about 5 kDa, at most about 6 kDa, at most about 7 kDa, at most about 8 kDa, at most about 9 kDa, at most about 10 kDa, at most about 20 kDa, at most about 30 kDa, at most about 40 kDa, at most about 50 kDa, at most about 60 kDa, at most about 80 kDa, at most about 100 kDa, at most about 120 kDa, at most about 140 kDa, at most about 160 kDa, at most about 180 kDa, at most about 200 kDa, at most about 220 kDa, at most about 240 kDa, at most about 260 kDa, at most about 280 kDa, at most about 300 kDa, at most about 350 kDa, at most about 400 kDa, at most about 450 kDa, at most about 500 kDa, at most about 550 kDa, at most about 600 kDa, at most about 650 kDa, at most about 700 kDa, at most about 750 kDa, at most about 800 kDa, at most about 850 kDa, at most about 900 kDa, at most about 950 kDa, at most about 1000 kDa, at most about 1100 kDa, at most about 1200 kDa, at most about 1300 kDa, at most about 1400 kDa, at most about 1500 kDa, at most about 1600 kDa, at most about 1700 kDa, at most about 1800 kDa, at most about 1900 kDa, or at most about 2000 kDa. In some instances, a saccharide or a derivative thereof may comprise a monosaccharide, a disaccharide, a trisaccharide, an oligosaccharide, or a polysaccharide.

[0057] A saccharide may comprise a sorbitan, a mannide, a trehalose, a lactose, a functional variant thereof, or a derivative thereof, or any combination thereof. A saccharide may comprise a sorbitan, a functional variant thereof, or a derivative thereof. A saccharide may comprise a trehalose, a functional variant thereof, or a derivative thereof. A saccharide may comprise a mannide a functional variant thereof, or a derivative thereof. A saccharide may comprise a lactose, a functional variant thereof, or a derivative thereof.

[0058] In some cases, the saccharide may comprise at least a derivation group. In some cases, the saccharide described herein may comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more derivation groups. In some cases, the saccharide described herein may comprise at most 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 derivation groups. The derivation group may comprise an ester group, an ether group, a carbamate, a thioester group, or a combination thereof. A saccharide may comprise sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate, a derivative thereof, or any combination thereof.

[0059] A saccharide may comprise sorbitan monooleate (SMO, a derivative thereof, or any combination thereof. A saccharide may comprise sorbitan dioleate, a derivative thereof, or any combination thereof. A saccharide may comprise sorbitan trioleate (STO), a derivative thereof, or any combination thereof. A saccharide may comprise sorbitan monostearate, a derivative thereof, or any combination thereof. A saccharide may comprise mannide monooleate (MMO, a derivative thereof, or any combination thereof. A saccharide may comprise lactose oleate, a derivative thereof, or any combination thereof. A saccharide may comprise trehalose monooleate, a derivative thereof, or any combination thereof.

[0060] In some cases, the transfection complex described herein may comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more saccharide molecules. In some cases, the transfection complex described herein may comprise at most 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 more saccharide molecules. The multiple saccharide molecules can be the same. The multiple saccharide molecules can be different.

[0061] In some cases, the saccharide described herein may comprise at least a lipid moiety. For example, a saccharide described herein may comprise a glycolipid. The lipid moiety may determine the physical properties of the lipid bilayer (e.g., flexibility and the rate of lipid exchange). In some cases, the saccharide described herein may comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more lipid moieties. In some cases, the saccharide described herein may comprise at most 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 lipid moieties.

[0062] In some instances, a lipid moiety or a derivative thereof may comprise an average molecular mass of at least about 0.05 kilodaltons (kDa), at least about 0.1 kDa, at least about 0.2 kDa, at least about 0.3 kDa, at least about 0.4 kDa, at least about 0.5 kDa, at least about 0.6 kDa, at least about 0.7 kDa, at least about 0.8 kDa, at least about 0.9 kDa, at least about 1 kDa, at least about 2 kDa, at least about 3 kDa, at least about 4 kDa, at least about 5 kDa, at least about 6 kDa, at least about 7 kDa, at least about 8 kDa, at least about 9 kDa, at least about 10 kDa, at least about 20 kDa, at least about 30 kDa, at least about 40 kDa, at least about 50 kDa, at least about 60 kDa, at least about 80 kDa, at least about 100 kDa, at least about 120 kDa, at least about 140 kDa, at least about 160 kDa, at least about 180 kDa, at least about 200 kDa, at least about 220 kDa, at least about 240 kDa, at least about 260 kDa, at least about 280 kDa, at least about 300 kDa, at least about 350 kDa, at least about 400 kDa, at least about 450 kDa, at least about 500 kDa, at least about 550 kDa, at least about 600 kDa, at least about 650 kDa, at least about 700 kDa, at least about 750 kDa, at least about 800 kDa, at least about 850 kDa, at least about 900 kDa, at least about 950 kDa, at least about 1000 kDa, at least about 1100 kDa, at least about 1200 kDa, at least about 1300 kDa, at least about 1400 kDa, at leastabout 1500 kDa, at least about 1600 kDa, at least about 1700 kDa, at least about 1800 kDa, at least about 1900 kDa, at least about 2000 kDa or more. In some instances, a lipid moiety or a derivative thereof may comprise an average molecular mass of at most about 0.05 kilodaltons (kDa), at most about 0.1 kDa, at most about 0.2 kDa, at most about 0.3 kDa, at most about 0.4 kDa, at most about 0.5 kDa, at most about 0.6 kDa, at most about 0.7 kDa, at most about 0.8 kDa, at most about 0.9 kDa, at most about 1 kDa, at most about 2 kDa, at most about 3 kDa, at most about 4 kDa, at most about 5 kDa, at most about 6 kDa, at most about 7 kDa, at most about 8 kDa, at most about 9 kDa, at most about 10 kDa, at most about 20 kDa, at most about 30 kDa, at most about 40 kDa, at most about 50 kDa, at most about 60 kDa, at most about 80 kDa, at most about 100 kDa, at most about 120 kDa, at most about 140 kDa, at most about 160 kDa, at most about 180 kDa, at most about 200 kDa, at most about 220 kDa, at most about 240 kDa, at most about 260 kDa, at most about 280 kDa, at most about 300 kDa, at most about 350 kDa, at most about 400 kDa, at most about 450 kDa, at most about 500 kDa, at most about 550 kDa, at most about 600 kDa, at most about 650 kDa, at most about 700 kDa, at most about 750 kDa, at most about 800 kDa, at most about 850 kDa, at most about 900 kDa, at most about 950 kDa, at most about 1000 kDa, at most about 1100 kDa, at most about 1200 kDa, at most about 1300 kDa, at most about 1400 kDa, at most about 1500 kDa, at most about 1600 kDa, at most about 1700 kDa, at most about 1800 kDa, at most about 1900 kDa, or at most about 2000 kDa.

[0063] The lipid moiety may comprise at least a fatty acid, at least a cholesterol group, or a combination thereof. In some cases, the fatty acid may comprise one or more oleate, laurate, stearate, a derivative thereof, or a combination thereof. In some cases, the fatty acid may comprise one or more oleate or a derivative thereof. In some cases, the fatty acid may comprise one or more laurate or a derivative thereof. In some cases, the fatty acid may comprise one or more stearate or a derivative thereof. The fatty acid may be saturated. The fatty acid may be monounsaturated. The fatty acid may be polyunsaturated.

[0064] In some cases, the oleate may comprise monooleate, dioleate, trioleate, or polyoleate. In some cases, the laurate may comprise monolaurate, dilaurate, trilaurate, or polylaurate. In some cases, the stearate may comprise monostearate, distearate, tristearate, or polystearate. The fatty acid may be derived from oleic or myristic acid).

[0065] The lipid moiety may comprise at least a fatty acyl, at least a cholesterol group, or a combination thereof. In some cases, the fatty acyl may comprise one or more oleate, laurate, stearate, a derivative thereof, or a combination thereof. In some cases, the fatty acyl may comprise one ormore oleate or a derivative thereof. In some cases, the fatty acyl may comprise one or more laurate or a derivative thereof. In some cases, the fatty acyl may comprise one or more stearate or a derivative thereof. The fatty acyl may be monounsaturated. The fatty acyl may be polyunsaturated.

[0066] The lipid moiety may comprise at least a fatty hydrocarbon chain, at least a cholesterol group, or a combination thereof. In some cases, the fatty hydrocarbon chain may comprise one or more oleate, laurate, stearate, a derivative thereof, or a combination thereof. In some cases, the fatty hydrocarbon chain may comprise one or more oleate or a derivative thereof. In some cases, the fatty hydrocarbon chain may comprise one or more laurate or a derivative thereof. In some cases, the fatty hydrocarbon chain may comprise one or more stearate or a derivative thereof. The fatty hydrocarbon chain may be monounsaturated. The fatty hydrocarbon chain may be polyunsaturated.

[0067] In some cases, a cationic lipid may be a hydrophilic lipid anchor, a linker group, a positively charged headgroup, or a combination thereof. In some cases, a cationic lipid may be a hydrophilic lipid anchor, a linker group, a positively charged headgroup, or a combination thereof. The linker can also provide sites for the introduction of novel side chains to enhance targeting, uptake, and trafficking. The positively charged head group on the cationic lipid may interact with the negatively charged nucleic acid (or a negatively charged moiety of a nucleic acid). The headgroups may be singly- or multiplicatively-charged as primary, secondary, tertiary, and / or quaternary amines. Multivalent headgroups, such as spermine, in a “T-shape” configuration may be efficient for promoting the uptake of nucleic acid molecules by a cell. In some case, an increase in the linker length may increase the uptake of nucleic acid molecules by a cell.

[0068] In some cases, the saccharide, its lipid moiety, or its derivation group may be biodegradable. For example, subsequent to being administrated into a subject, a derivative thereof, or any combination thereof may be degradable within the subject. Biodegradable lipids can be metabolized by various enzymes (e.g., esterase, peptidases) to minimize any toxicity.Lipids

[0069] In some cases, the compositions disclosed herein may comprise a lipid molecule. Lipids may facilitate update of a nucleic acid molecule by a cell. The cationic charges of the lipid may reduce or neutralize the anionic charge of the nucleic acid molecule and / or the plasma membrane of the cell, reducing the electrostatic repulsion of the nucleic acid molecule and the plasma membrane of the cell. Thus, a lipid molecule may be a cationic lipid. In some cases, a lipid may be a structural component of a transfection complex. In some cases, through lipid-lipid interaction with the plasmamembrane of a cell, lipids may also facilitate the endocytosis of the nucleic acid molecule by the cell.

[0070] A lipid or a derivative thereof may have cationic moieties. A lipid or a derivative thereof may have a cationic charge in an aqueous solution. A lipid or a derivative thereof may have a cationic charge in a neutral solution. A lipid or a derivative thereof may have a neutral charge in an aqueous solution. A lipid or a derivative thereof may have a neutral charge in a neutral solution. A lipid or a derivative thereof may have an anionic charge in an aqueous solution. A lipid or a derivative thereof may have an anionic charge in a neutral solution. A lipid or a derivative thereof may have a cationic charge in an aqueous solution. A lipid or a derivative thereof may have a cationic charge in a neutral solution. A lipid or a derivative thereof may have a cationic charge in an aqueous neutral solution. A lipid or a derivative have a neutral charge in an aqueous neutral solution. A lipid or a derivative thereof may have an anionic charge in an aqueous neutral solution.

[0071] A corresponding cationic charge density of cationic moieties may be measured in mequivalent of basic group per gram of the lipid molecule (mequiv / g). In some cases, the cationic charge density of cationic moieties may be determined by the poly electrolyte titration method, conductometric titration, and / or acid / base titration. In some cases, the corresponding cationic charge density of a lipid or a derivative thereof may be at least about 0.5 mequiv / g, at least about 1 mequiv / g, at least about 1.5 mequiv / g, at least about 2 mequiv / g, at least about 2.5 mequiv / g, at least about 3 mequiv / g, at least about 3.5 mequiv / g, at least about 4 mequiv / g, at least about 4.5 mequiv / g, at least about 5 mequiv / g, at least about 5.5 mequiv / g, at least about 6 mequiv / g, at least about 6.5 mequiv / g, at least about 7 mequiv / g, at least about 7.5 mequiv / g, at least about 8 mequiv / g, at least about 8.5 mequiv / g, at least about 9 mequiv / g, at least about 9.5 mequiv / g, at least about 10 mequiv / g, at least about 10.5 mequiv / g, at least about 11 mequiv / g, at least about 11.5 mequiv / g, at least about 12 mequiv / g, at least about 12.5 mequiv / g, at least about 13 mequiv / g, at least about 13.5 mequiv / g, at least about 14 mequiv / g, at least about 14.5 mequiv / g, at least about 15 mequiv / g, at least about 15.5 mequiv / g, at least about 16 mequiv / g, at least about 16.5 mequiv / g, at least about 17 mequiv / g, at least about 17.5 mequiv / g, at least about 18 mequiv / g, at least about 18.5 mequiv / g, at least about 19 mequiv / g, at least about 19.5 mequiv / g, or at least about 20 mequiv / g. In some cases, the corresponding cationic charge density of a lipid or a derivative thereof may be at most about 0.5 mequiv / g, at most about 1 mequiv / g, at most about 1.5 mequiv / g, at most about 2 mequiv / g, at most about 2.5 mequiv / g, at most about 3 mequiv / g, at most about 3.5 mequiv / g, at most about 4 mequiv / g, at most about 4.5 mequiv / g, at most about 5 mequiv / g, at most about 5.5 mequiv / g, atmost about 6 mequiv / g, at most about 6.5 mequiv / g, at most about 7 mequiv / g, at most about 7.5 mequiv / g, at most about 8 mequiv / g, at most about 8.5 mequiv / g, at most about 9 mequiv / g, at most about 9.5 mequiv / g, at most about 10 mequiv / g, at most about 10.5 mequiv / g, at most about 11 mequiv / g, at most about 11.5 mequiv / g, at most about 12 mequiv / g, at most about 12.5 mequiv / g, at most about 13 mequiv / g, at most about 13.5 mequiv / g, at most about 14 mequiv / g, at most about 14.5 mequiv / g, at most about 15 mequiv / g, at most about 15.5 mequiv / g, at most about 16 mequiv / g, at most about 16.5 mequiv / g, at most about 17 mequiv / g, at most about 17.5 mequiv / g, at most about 18 mequiv / g, at most about 18.5 mequiv / g, at most about 19 mequiv / g, at most about 19.5 mequiv / g, or at most about 20 mequiv / g. In some cases, the corresponding cationic charge density of a lipid or a derivative thereof may be from 0.005 to 2000 mequiv / g. In some cases, the corresponding cationic charge density of cationic moieties of a lipid or a derivative thereof may be from 0.05 to 200 mequiv / g. In some cases, the corresponding cationic charge density of cationic moieties of a lipid or a derivative thereof may be from 0.5 to 20 mequiv / g.

[0072] In some cases, the transfection complex described herein may comprise at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more lipid molecules. In some cases, the transfection complex described herein may comprise at most 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 more lipid molecules. The multiple lipid molecules can be the same. The multiple lipid molecules can be different. In some case, the transfection complex can comprise at least 2 different lipid molecules.

[0073] A composition or transfection complex described herein can comprise a lipid molecule. The lipid molecule may comprise a cationic lipid. The cationic lipid molecule can comprise ethanolamine, carnitine, a derivative thereof, a functional variant thereof, of a combination thereof. The cationic lipid molecule can comprise ethanolamine. The cationic lipid molecule can comprise ethanolamine, a derivative thereof, or a functional variant thereof. The cationic lipid molecule can comprise carnitine. The cationic lipid molecule can comprise carnitine, a derivative thereof, or a functional variant thereof. The cationic lipid molecule can comprise a structure of formula (I), formula (II), or formula (II):

[0074] The cationic lipid molecule of a composition or transfection complex described herein can comprise a lipid molecule having a structure of formula (I). The cationic lipid molecule of acomposition or transfection complex described herein can comprise a lipid molecule having a structure of formula (II). The cationic lipid molecule of a composition or transfection complex described herein can comprise a lipid molecule having a structure of formula (III).

[0075] In some cases, R1is C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl. In some cases, R1is Ce- C22 alkyl. In some cases, R1is C6-C22 alkenyl. In some cases, R1is C6-C22 alkynyl. In some cases, each of R2and R3is H, C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl. In some cases, R2is H. In some cases, R2is C6-C22 alkyl. In some cases, R2is C6-C22 alkenyl. In some cases, R2is C6-C22 alkynyl. In some cases, R3is H. In some cases, R3is C6-C22 alkyl. In some cases, R3is C6-C22 alkenyl. In some cases, R3is C6-C22 alkynyl. In some cases, R4is C6-C22 alkyl, C6-C22 alkenyl, or Ce- C22 alkynyl. In some cases, R4is C6-C22 alkyl. In some cases, R4is C6-C22 alkenyl. In some cases, R4is C6-C22 alkynyl. In some cases, R5is C1-C4 alkyl. In some cases, X is independently an anionic radical selected from the group consisting of: hydroxide, halide, sulfate, methylsulfate, ethylsulfate, and phosphate ions. In some cases, X is an anionic radical. In some cases, X is hydroxide. In some cases, X is halide. In some cases, X is sulfate. In some cases, X is methylsulfate. In some cases, X is ethylsulfate. In some cases, X is phosphate ions. In some cases, R1is independently C6-C22 alkyl, Ce- C22 alkenyl, or C6-C22 alkynyl; each of R2and R3is independently H, C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl; R4is C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl; R5is independently C1-C4 alkyl; and X is independently an anionic radical selected from the group consisting of: hydroxide, halide, sulfate, methylsulfate, ethylsulfate, and phosphate ions.

[0076] In some cases, the cationic lipid molecule comprises:.

[0077] In some cases, the cationic lipid molecule comprises formula IV. In some cases, the cationic lipid molecule comprises formula V. In some cases, the cationic lipid molecule comprises formulaVI. In some cases, the cationic lipid molecule comprises formula VII. In some cases, the cationic lipid molecule comprises formula VIII. In some cases, the cationic lipid molecule comprises formula IX. A composition or transfection complex as described herein may comprise a cationic lipid molecule, wherein the cationic lipid molecule is selected from the group consisting of DOTAP, DDA, DLin-KC2-DMA (KC2), DLin-MC3-DMA (MC3), cKK-E12, C12-200, and any one of Formula (I)-(IX). The cationic lipid molecule may be selected from the group consisting of DOTAP and any one of Formula (IV)-(IX).

[0078] In some cases, the composition or transfection complex described herein may comprise a structural lipid molecule. In some cases, the composition or transfection complex described herein may comprise a second lipid molecule different from a cationic lipid molecule as described herein. The structural lipid molecule may be selected from the group consisting of DOPE (1,2-Distearoyl- sn-glycero-3-phosphocholine), DSPC (l,2-distearoyl-sn-glycero-3-phosphorylcholine), SQDG (glycolipid), MGDG (monogalactosyldiacylglycerol)), DGDG (digalactosyldiacylglycerol), CycloPC ( 1 -palmitoy 1 -2-cis-9, 10-methylenehexadecanoy l-sn-glycero-3 -phosphocholine) and / or DGTS(Diacylglyceryltrimethylhomo-Ser). A structural lipid molecule may comprise l,2-di-(9Z- octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE).

[0079] In some instances, a lipid or a derivative thereof may comprise an average molecular mass of at least about 0.05 kilodaltons (kDa), at least about 0.1 kDa, at least about 0.2 kDa, at least about 0.3 kDa, at least about 0.4 kDa, at least about 0.5 kDa, at least about 0.6 kDa, at least about 0.7 kDa, at least about 0.8 kDa, at least about 0.9 kDa, at least about 1 kDa, at least about 2 kDa, at least about 3 kDa, at least about 4 kDa, at least about 5 kDa, at least about 6 kDa, at least about 7 kDa, at least about 8 kDa, at least about 9 kDa, at least about 10 kDa, at least about 20 kDa, at least about 30 kDa, at least about 40 kDa, at least about 50 kDa, at least about 60 kDa, at least about 80 kDa, at leastabout 100 kDa, at least about 120 kDa, at least about 140 kDa, at least about 160 kDa, at least about 180 kDa, at least about 200 kDa, at least about 220 kDa, at least about 240 kDa, at least about 260 kDa, at least about 280 kDa, at least about 300 kDa, at least about 350 kDa, at least about 400 kDa, at least about 450 kDa, at least about 500 kDa, at least about 550 kDa, at least about 600 kDa, at least about 650 kDa, at least about 700 kDa, at least about 750 kDa, at least about 800 kDa, at least about 850 kDa, at least about 900 kDa, at least about 950 kDa, at least about 1000 kDa, at least about 1100 kDa, at least about 1200 kDa, at least about 1300 kDa, at least about 1400 kDa, at least about 1500 kDa, at least about 1600 kDa, at least about 1700 kDa, at least about 1800 kDa, at least about 1900 kDa, at least about 2000 kDa or more. In some instances, a lipid or a derivative thereof may comprise an average molecular mass of at most about 0.05 kilodaltons (kDa), at most about 0.1 kDa, at most about 0.2 kDa, at most about 0.3 kDa, at most about 0.4 kDa, at most about 0.5 kDa, at most about 0.6 kDa, at most about 0.7 kDa, at most about 0.8 kDa, at most about 0.9 kDa, at most about 1 kDa, at most about 2 kDa, at most about 3 kDa, at most about 4 kDa, at most about 5 kDa, at most about 6 kDa, at most about 7 kDa, at most about 8 kDa, at most about 9 kDa, at most about 10 kDa, at most about 20 kDa, at most about 30 kDa, at most about 40 kDa, at most about 50 kDa, at most about 60 kDa, at most about 80 kDa, at most about 100 kDa, at most about 120 kDa, at most about 140 kDa, at most about 160 kDa, at most about 180 kDa, at most about 200 kDa, at most about 220 kDa, at most about 240 kDa, at most about 260 kDa, at most about 280 kDa, at most about 300 kDa, at most about 350 kDa, at most about 400 kDa, at most about 450 kDa, at most about 500 kDa, at most about 550 kDa, at most about 600 kDa, at most about 650 kDa, at most about 700 kDa, at most about 750 kDa, at most about 800 kDa, at most about 850 kDa, at most about 900 kDa, at most about 950 kDa, at most about 1000 kDa, at most about 1100 kDa, at most about 1200 kDa, at most about 1300 kDa, at most about 1400 kDa, at most about 1500 kDa, at most about 1600 kDa, at most about 1700 kDa, at most about 1800 kDa, at most about 1900 kDa, or at most about 2000 kDa.

[0080] In some cases, a lipid may comprise a liposome. In some cases, a lipid may comprise a nanoparticle. In some cases, a lipid may comprise a liposome, a polymer nanoparticle, and a lipid- nanoparticle. In some cases, a liposome is complexed with an associated nucleic acid for subsequent delivery of the nucleic acid to a cell or tissue. In some cases, a liposome may comprise cholesterol. In some cases, a lipid nanoparticle may condense and deliver various nucleic acid molecules to a cell. In some instances, a nanoparticle may comprise a solid lipid (i.e., lipid that remains solid at room temperature and body temperature) or a liquid lipid (i.e., oil, which remains liquid at room temperature and body temperature, for example, vegetable oil or a lipid extracted from humanadipose tissue). In some instances, a nanoparticle may comprise a lipid layer (e.g., lipid monolayer) enclosing the nanoparticle core where a nucleic acid-cationic lipid is entrapped. In some cases, the lipid layer can be about 2 to 10 nm. In some cases, the lipid layer can be about 5 nm. In some instances, a lipid layer can be a lipid monolayer. In some cases, a lipid layer can comprise two or more lipid layers. In some instances, a lipid layer can be selected from: phospholipids such as lecithin, (L-a-phosphatidylcholine) or phosphatidylcholines with saturated and unsaturated fatty acids. In some cases, a lipid layer may also comprise other phospholipids, such as phosphatidic acids, phosphatidylethanolamines, phosphatidylglycerols, phosphatidylserines, phosphatidylinositols, cardiolipins, or a combination thereof. In some cases, a lipid layer can comprise lipid-PEG (lipid-poly ethyleneglycol) conjugates with various molecular weights of PEG.

[0081] In some cases, a nanoparticle can be a solid lipid nanoparticle. In some cases, a solid lipid nanoparticle can comprise a lipid layer enclosing the nanoparticle core, where a solid lipid can be disposed in the nanoparticle core along with the nucleic acid-cationic lipid conjugate. In some instances, a solid lipid can be a lipid that remains a solid at the body temperature of an animal or cell, so that a solid lipid can be selected based on the animal or the cell. In some instances, a solid lipid can be selected from the following group: monoglycerides (e.g. glycerol monostearate), diglycerides (e.g., glycerol behenate), triglycerides (e.g., tristearin, trimyristin, trilaurin), waxes (e.g., acetyl palmitate), fatty acids (e.g., stearic acid, palmitic acid), steroids (e.g., cholesterol), and a combination thereof.

[0082] In some cases, a lipid particle further may comprise a PEG-lipid. The PEG-lipid can prevent the aggregation of particles. PEG may comprise a linear, water-soluble polymer of ethylene PEG repeating units with two terminal hydroxyl groups and are classified by their molecular weights. In some embodiments, the PEG moiety of the PEG-lipid may comprise an average molecular weight from about 600 to about 5,000 Da. In some cases, the average molecular weight is 2,000 Da. In some cases, the average molecular weight is 750 Da. In some instances, the PEG-lipid is selected from one or more of the following: PEG-Maleimide, PEG-PDP, PEG-Biotin, PEG- Amine, PEG-DBCO, PEG- Azide, PEG-Cyanur, PEG-Succinyl, PEG-Folate and / or PEG-Carboxylic acid. In some instances, the PEG-lipid is 1,2- DMG PEG 2000 (1,2 dimyristoyl-sn-glycero-3-phosphoethanolamine-N- [methoxy- (polyethylene glycol)-2000. In some instances, the PEG-lipid is a combination of 1,2- DMGPEG2000 and l,3-DMGPEG2000 (l,3-DMGPEG2000 (1,3 dimynstoyl-sn-glycero-3- phosphoethanolamine-N-[methoxy- (polyethylene glycol)-2000), optionally in a 97:3 ratio. In some instances, the PEG-lipid is 1,2- DMG PEG 1000 (1,2 dimyristoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy- (polyethylene glycol)- 1000], In some instances, the PEG-lipid is 1,2- DMGPEG 3000 (1,2 dimyristoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy-(poly ethylene glycol)-3000]. In some instances, the PEG-lipid may comprise from about 1 % to about 10 % of the total lipid present in the particle. In some cases, the sterol may comprise from about 1 % to about 5 %, 1 % to about 3 %, about 1 % to about 2 %, about 1 % to 1.5 % of the total lipid present in the particle. In some cases, the PEG-lipid may comprise 2 % of the total lipid present. In some cases, the PEG-lipid may comprise 1.5 % of the total lipid present. In other cases, the PEG-lipid may comprise from about 1 % to about 5 % of the total lipid present in the particle. In some cases, the PEG-lipid is 1,2- DMGPEG 2000 (1,2 dimyristoyl-sn-glycero-3- phosphoethanolamine-N-[methoxy- (polyethylene glycol)-2000 and may comprise from about 1 % to about 2 % of the total lipid present. In some instances, the PEG-lipid is a combination of 1 ,2- DMGPEG2000 and l,3-DMGPEG2000 (l,3-DMGPEG2000 (1,3 dimynstoyl-sn-glycero-3- phosphoethanolamine-N-[methoxy- (polyethylene glycol)-2000), and may comprise from about 1 % to about 2 % of the total lipid present. In some instances, the PEG-lipid is 1,2- DMGPEG 1000 (1,2 dimyristoyl-sn-glycero-3-phosphoethanolamine-N- [methoxy- (polyethylene glycol)-1000], and may comprise from about 1 mol % to about 2 mol % of the total lipid present. In some instances, the PEG-lipid is 1,2- DMGPEG 3000 (1,2 dimyristoyl-sn-glycero-3-phosphoethanolamine-N- [methoxy- (polyethylene glycol)-3000], and may comprise from about 1 % to about 2 % of the total lipid present. In some instances, the lipid particle further may comprise a sterol.

[0083] In some instances, the sterol may be a cholesterol or a derivative thereof. Examples of cholesterol derivatives may comprise cholestanol, cholestanone, cholestenone, coprostanol, cholesteryl-2'-hydroxyethyl ether, cholesteryl-4'-hy-droxybutyl ether, and mixtures thereof. In some cases, the sterol may comprise from about 10 % to about 50 mol % of the total lipid present in the particle. In some cases, the sterol may comprise from about 10 % to about 20 %, about 10 % to about 15 %, about 30 % to about 40 % of the total lipid present in the particle. In some cases, the sterol may comprise about 38 % of the total lipid present. In other cases, the sterol may comprise from about 30 mol % to about 40 mol % of the total lipid present in the particle. In some cases, the sterol may be cholesterol or a derivative thereof and may comprise from about 30 % to about 40 % of the total lipid present. In some cases, the lipid particle further may comprise one or more stabilizing agents. Stabilizing agents ensure integrity of the lipid mixture. In some cases, the one or more stabilizing agents are polyethylene glycol-lipids. Suitable polyethylene glycol-lipids include PEG-modified phosphatidylethanolamine, PEG-modified phosphatidic acid, PEG-modifiedceramides (e.g., PEG-CerC14 or PEG-CerC20), PEG-modified dialkylamines, PEG-modified diacylglycerols, PEG-modified alkylglycerols. Representative polyethylene glycol-lipids include PEG-c-DOMG, PEG-c-DMA, and PEG-s-DMG. In one embodiment, the polyethylene glycol-lipid may be N-[(methoxy poly(ethylene glycol)2000)carbamyl]-l,2-dimyristyloxlpropyl-3 -amine (PEG- c-DMA).

[0084] In some cases, a cationic lipid is selected from one or more of the following: 1 ,2- dilinoleyloxy-N,N-dimethylaminopropane (DLinDMA), 1 ,2-dilinolenyloxy-N,N- dimethylaminopropane (DLenDMA), dilinoleyl-4-(2-dimethylaminoethyl)-[l,3]-dioxolane (DLin-K- C2-DMA; "XTC2"), 2,2-dilinoleyl-4-(3-dimethylaminopropyl)-[l,3]-dioxolane (DLin-K-C3-DMA),2.2-dilinoley l-4-( 4-dimethylamino buty l)-[ 1,3 ]-dioxo lane (DLin-K-C4-DMA), 2,2-dilinoleyl-5- dimethylaminomethyl- [1,3 ] -dioxane (DLin-K6-DMA), 2,2-dilinoleyl-4-Nmethylpepiazino- [1, 3 ] - dioxolane (DLin-K-MPZ), 2,2-dili-noleyl-4-dimethylaminomethyl-[l,3]-dioxolane (DLin-KDMA),1.2-dilinoleylcarbamoyloxy-3 -dimethylaminopropane (DLin-C-AP), 1 , 2-dilinoley oxy-3 - (dimethylamino acetoxypropane (DLin-DAC), 1, 2-dilinoley oxy-3-morpholinopropane (DLin-MA),1.2-dilinoleoyl-3 -dimethylaminopropane (DLinDAP), 1, 2-dilinoley lthio-3-dimethylaminopropane (D Lin-S-D MA), 1 -linoleoyl-2-linoleyloxy-3-dimethylaminopropane (DLin-2-D MAP), 1,2- dilinoley loxy-3-trimethy laminopropane chloride salt (DLin-TMA.Cl), l,2-dilinoleoyl-3-trimethy- laminopropane chloride salt (DLin-TAP.Cl), 1 ,2-dilinoleyloxy-3-(N-methylpiperazino) propane (DLin-MPZ), 3-(N,Ndilinoleylamino)-l,2-propanediol (DLinAP), 3-(N,Ndioleylamino)-l,2- propanedio (DOAP), l,2-dilinoleyloxo-3-(2-N,N-dimethylamino) ethoxypropane (DLin-EG-DMA), N,N-dioleyl-N,N-dimethylammonium chloride (DO DAC), l,2-dioleyloxy-N,N- dimethylaminopropane (DODMA), 1,2-disteary loxy-N,N-dimethy laminopropane (DSD MA), N- (l-(2,3-dioley loxy) propy l)-N,N,N-trimethy ammonium chloride (DOTMA), N,N-distearyl-N,N- dimethylammonium bromide (DDAB), N-(l-(2,3-dioleoyloxy) propyl)-N,N,N-trimethylanunonium chloride (DOTAP), 3-(N N',N'dimethylaminoethane)-carbamoyl) cholesterol (DC-Chol), N-(l,2- dimyristyloxyprop-3-yl)- N,N-dimethyl-N-hydroxyethyl ammonium bromide (DMRIE), 2,3- dioleyloxy-N-[2(spermine- carboxamido) ethy l]-N,N-dimethy 1-1-propanaminiumtrifiuoroacetate (DOSPA). Dioctadecylamidoglycyl spermine (DOGS), 3-dimethylamino-2-(cholest-5-en-3-beta- oxybutan-4-oxy)-l-(cis,cis-9, 12-octadecadienoxy)propane (CLinDMA), 2-[5'-(cholest-5-en-3-beta- oxy)-3'-oxapentoxy)-3-dimethyl-l-(cis,cis-9'l-2'-octadecadienoxy) propane (CpLinDMA), N,N- dimethyl-3,4-dioleyloxybenzylamine (DMOBA), 1 ,2-N,N'dioleylcarbamyl-3-dimethylaminopropane (DOcarbDAP), 1 ,2-N,N'-dilinoleylcarbamyl-3-dimethy laminopropane (DLincarbDAP), or mixturesthereof. In some cases, a cationic lipid is selected from one or more of DOTAP (DOTAP methosulfate, N-(2,3-Dioleoyloxy-l-propyl) trimethylammonium methyl sulfate), DDA(Dimethyldioctadecylammonium (Bromide Salt)) , DLin-KC2-DMA (KC2)(2-[2,2- bis[(9Z,12Z)-octadeca-9,12-dienyl]-l,3-dioxolan-4-yl]-N,N-dimethylethanamine), DLin-MC3-DMA (MC3)((6Z,9Z,28Z, 3 lZ)-heptatriacont-6, 9, 28, 31 -tetraene- 19-yl 4-(dimethylamino)butanoate), cKK- E12(3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2, 5-dione) and / or C12-200(.1,1 ‘-((2- (4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl) (2-hydroxydodecyl)amino)ethyl)piperazin-l- yl)ethyl)azanediyl)bis(dodecan-2-ol). In some cases, a cationic lipid is selected from one or more of 1 ,2-diolelyloxy-3-(trimethylamino)propane (DOTAP); N-[l -(2,3,-ditetradecyloxy)propyl]-N,N- dimethyl-N-hydroxyethylammonium bromide (DMRIE); N-[l-(2,3,-dioleyloxy)propyl]-N,N- dimethyl-N-hydroxy ethylammonium bromide (DORIE); N-[l-(2,3-dioleyloxy)propyl]-N,N,N- trimethylammonium chloride (DOTMA); 3P[N-(N',N'-dimethylaminoethane)carbamoly]cholesterol (DC-Chol); and dimethyldioctadecylammonium (DDAB). In some cases, the cationic lipid may be a neutral cationic lipid, that is, a lipid that at physiologic pH of 7.4 is predominantly, e.g., greater than 50%, neutral in charge but at a selected pH value less than physiologic pH tends to have a positive charge.

[0085] In some instances, the cationic lipid is DOTAP and may comprise 40 mole % (mol %) to 50 mol % of the total lipid present, the structural lipid is (l,2-distearoyl-sn-glycero-3- phosphorylcholine), SQDG (glycolipid), MGDG (monogalactosyldiacylglycerol)) (DSPC) and may comprise 10 mol of the total lipid present, and the lipid particle further may comprise cholesterol at 30 mol% to 40 mol % of the total lipid present, and one or more PEG-lipids at 1 mol to 2 mol% of the total lipid present. In some instances, the cationic lipid is DOTMA and may comprise 40 mol % to 50 mol % of the total lipid present, the structural lipid is DSPC and may comprise 10 mol % of the total lipid present, and the lipid particle further may comprise cholesterol at 30 mol % to 40 mol% of the total lipid present, and one or more PEG-lipids at 1 mol % to 2 mol % of the total lipid present.

[0086] In some cases, the methods for synthesizing the lipid molecules are described elsewhere in this disclosure (see, for example, Example 2).

[0087] Described herein is a method, comprising: (a) providing a cell with a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); and (iii) a cationic lipid; and (b) delivering said composition into said cell. The cationic lipid may be selected from the groupconsisting of DOTAP, DDA, DLin-KC2-DMA (KC2), DLin-MC3-DMA (MC3), cKK-E12, C12-200 and any compound of Formula (I)-(IX). The cationic lipid may be selected from the group consisting of DOTAP and any compound of Formula (IV)-(IX). The saccharide molecule may be selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate. Accordingly, provided herein is a method, comprising: (a) providing a cell with a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule, wherein the saccharide molecule is selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate; and (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP, DDA, DLin-KC2-DMA (KC2), DLin-MC3-DMA (MC3), cKK-E12, C12-200 and any compound of Formula (I)-(DC); and (b) delivering said composition into said cell. The composition used in the method may further comprise a structural lipid. Accordingly, described herein is a method, comprising: (a) providing a cell with a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); (iii) a cationic lipid; and (iv) a structural lipid; and (b) delivering said composition into said cell. The structural lipid may be selected from the group consisting of DOPE, DSPC, SQDG, MGDG, DGDG, Cyclo-PC, and DGTS. The structural lipid may be DOPE. Accordingly, provided herein is a method, comprising: (a) providing a cell with a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule, wherein the saccharide molecule is selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate; (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP, DDA, DLin-KC2-DMA (KC2), DLin-MC3-DMA (MC3), cKK-E12, C12- 200 and any compound of Formula (I)-(IX); and (iv) a structural lipid, wherein the structural lipid is selected from the group consisting of DOPE, DSPC, SQDG, MGDG, DGDG, Cyclo-PC, and DGTS; and (b) delivering said composition into said cell. Also described herein is a method, comprising: (a) providing a cell with a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule, wherein the saccharide molecule is selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate; (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP and any compound of Formula (IV)-(IX); and (iv) a structural lipid, wherein the structural lipid is DOPE; and (b) delivering said composition into said cell.

[0088] Also provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); and (iii) a cationic lipid. Provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); and (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP, DDA, DLin-KC2-DMA (KC2), DLin-MC3-DMA (MC3), cKK-E12, C12-200 and any compound of Formula (I)-(IX). The cationic lipid may be selected from the group consisting of DOTAP and any compound of Formula (IV)- (IX). Provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule, wherein the saccharide molecule is selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate; and (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP and any compound of Formula (IV)- (IX). The composition may further comprise a structural lipid. Accordingly, provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); (iii) a cationic lipid; and (iv) a structural lipid. The structural lipid may be selected from the group consisting of DOPE, DSPC, SQDG, MGDG, DGDG, Cyclo-PC, and DGTS. Provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP, DDA, DLin-KC2-DMA (KC2), DLin-MC3-DMA (MC3), cKK-E12, C12-200 and any compound of Formula (I)-(IX); and (iv) a structural lipid, wherein the structural lipid is selected from the group consisting of DOPE, DSPC, SQDG, MGDG, DGDG, Cyclo-PC, and DGTS. Provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule, wherein the saccharide molecule is selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate; (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP and any compound of Formula (IV)-(IX); and (iv) a structural lipid, wherein the structural lipid is selected from the group consisting of DOPE, DSPC, SQDG, MGDG, DGDG,Cyclo-PC, and DGTS. The structural lipid may be DOPE. Provided herein is a composition comprising (i) a nucleic acid molecule, (ii) a saccharide molecule, wherein the saccharide molecule is selected from the group consisting of sorbitan monooleate, sorbitan trioleate and mannide monooleate; (iii) a cationic lipid, wherein the cationic lipid is selected from the group consisting of DOTAP and any compound of Formula (IV)-(IX); and (iv) a structural lipid, wherein the structural lipid is DOPE.Nucleic acid

[0089] In some instances, a nucleic acid may be a nucleic acid molecule. In some cases, a nucleic acid may be a species / type of nucleic acids. A species or type of nucleic acid may share a common biological function, a mechanism of action, a structural characteristic, or a combination thereof. A species or type of nucleic acid may be a messenger ribonucleic acid (mRNA), a DNA, a micro ribonucleic acid (miRNA), a transfer ribonucleic acid (tRNA), a long non-coding RNA (IncRNA), a ribosomal ribonucleic acid (rRNA), a small nuclear RNA (snRNA), a piwi-interacting RNA (piRNA), a small nucleolar RNA (snoRNA), an extracellular RNA(exRNA), a small cajal bodyspecific RNA (scaRNA), a silencing ribonucleic acid (siRNA), self-amplifying RNA (saRNA), a YRNA (small noncoding RNA), a heterogeneous nuclear RNA (HnRNA), endless / circular RNA (eRNA), trans-amplifying RNA (ta-RNA), complementary DNA (cDNA), a transfer RNA (tRNA), a ribosomal RNA, a short-hairpin RNA (shRNA), a small activating RNA (RNAa), or a ribozyme. A nucleic acid molecule may comprise one or more modified nucleotides, such as methylated nucleotides and nucleotide analogs. In some cases, a nucleic acid molecule may comprise a polymeric form of nucleotides. In some cases, a nucleic acid molecule may comprise a polynucleotide. In some cases, a nucleic acid molecule may comprise a modified polynucleotide. In some cases, a nucleic acid molecule may comprise a canonical or non-canonical nucleotide. A canonical nucleotide may comprise adenosine (A), cytosine (C), guanine (G), thymine (T), uracil (U), or variants thereof. In some cases, a nucleic acid may be single-stranded, double-stranded or triple stranded. In some cases, a nucleic acid may be single-stranded. In some cases, a nucleic acid may be double-stranded. In some cases, a nucleic acid may be single-stranded and double-stranded.

[0090] In some cases, a nucleic acid molecule may be linear or closed linear double-stranded (e.g., a doggybone DNA). In some cases, a nucleic acid molecule may be circular. In some cases, a nucleic acid molecule may be branched. A nucleic acid molecule may comprise a deoxyribonucleic acid (DNA) or ribonucleic acid (RNA). A nucleic acid molecule may comprise a peptide nucleic acid(PNA), an unlocked nucleic acid (UNA), a locked nucleic acid (LNA), an endl ess / circular RNA. LNA may comprise a structurally rigid modification (e.g., a 2'-O, 4'-C methylene bridge). UNA may comprise a flexible modification. The modification may restrict the flexibility of the ribofuranose ring and lock the structure into a rigid bicyclic formation. LNA may have increased thermal stability and hybridization specificity relative to an unmodified nucleic acid. UNA may have a structurally flexible modification (e.g., an acyclic analogue of RNA in which the bond between the C2' and C3' atoms of the ribose ring has been cleaved). UNA may lack the C2’-C4’ bond. PNA may comprise synthetic mimics of nucleic acid in which the deoxyribose phosphate backbone or ribose phosphate backbone is replaced by a pseudo-peptide polymer to which the nucleobases are linked. A nucleic acid molecule may comprise a coding sequence. A nucleic acid molecule may comprise a noncoding sequence. A nucleic acid molecule may comprise a coding or non-coding region of a gene or gene fragment, a locus defined from linkage analysis, an exons, an intron, an intein, or any combination thereof.

[0091] In some instances, a nucleic acid molecule may be monocistronic. In some cases, a nucleic acid molecule may be polycistronic. A monocistronic nucleic acid molecule may comprise one coding sequence, the coding sequence is configured to be recognized by a ribosome for the translation. A polycistronic nucleic acid molecule may comprise at least two coding sequences, each coding sequence is configured to be recognized by a ribosome for the translation of the coding sequence. In some cases, a nucleic acid molecule may comprise a sequence of a human gene. In some cases, a nucleic acid molecule may comprise a sequence of a non-human gene.

[0092] In some instances, a nucleic acid molecule may comprise a messenger ribonucleic acid (mRNA), a DNA, a micro ribonucleic acid (miRNA), a transfer ribonucleic acid (tRNA), a long noncoding RNA (IncRNA), a ribosomal ribonucleic acid (rRNA), a small nuclear RNA (snRNA), a piwi-interacting RNA (piRNA), a small nucleolar RNA (snoRNA), an extracellular RNA(exRNA), a small cajal body-specific RNA (scaRNA), a silencing ribonucleic acid (siRNA), a small activating RNA (RNAa), a self-amplifying RNA (saRNA), a YRNA (small noncoding RNA), a heterogeneous nuclear RNA (HnRNA), an endless / circular RNA (eRNA), a trans-amplifying RNA (ta-RNA), complementary DNA (cDNA) or a combination thereof. In some instances, a nucleic acid molecule may also comprise a transfer RNA (tRNA), a ribosomal RNA, a short-hairpin RNA (shRNA), a ribozyme, a recombinant nucleic acid, a branched nucleic acid, a plasmid, a vector, an isolated DNA, an isolated RNA, or any combination thereof. In some instances, a nucleic acid molecule may comprise an mRNA, a ta-RNA, a saRNA, an eRNA, or a combination thereof. A nucleic acidmolecule may comprise a siRNA, a miRNA, a shRNA, a YRNA, or a combination thereof. A nucleic acid molecule may comprise a siRNA, a miRNA, or a combination thereof. In some cases, the vector / plasmid system may be one for being used in the animal cells. In some cases, In some cases, the vector / plasmid system may be one for being used in the plant cells.

[0093] In some instances, a nucleic acid molecule may comprise a coding RNA. A coding RNA may comprise mRNA, RNAa, saRNA, or taRNA. The coding RNA may be single-stranded. In some cases, the coding RNA, when transfected into a cell, may allow a ribosome of the cell to translate the coding sequence of the coding RNA and synthesize a protein based on the coding sequence of the coding RNA.

[0094] In some instances, an mRNA may comprise a cell type-specific gene sequence. A cell type specific-gene sequence may comprise the sequence of a gene that is expressed or specifically expressed of the cell type. The cell type may comprise an ectoderm, an endoderm, or a mesoderm. The cell type may also comprise an adipogenic, angiogenic, cardiogenic, immunogenic, chondrogenic, endothelial, epithelial, hematopoietic, hepatogenic, myogenic, neurogenic, osteogenic, parenchymal, renal, retinal cell. In some cases, a cell may be a T cell, a B cell, a natural killer cell, a neutrophil, an eosinophil, a basophil, a mast cell, a monocyte, a macrophage, or a dendritic cell. The cell type may also comprise any somatic cell, a stem cell or an immortalized cell. The stem cell or immortalized cell may comprise an induced pluripotent stem cell (iPSC), an embryonic stem cell (ESC), a mesenchymal stem cell (MSC), a satellite cell, a fibroblast.

[0095] In some instances, a coding RNA may comprise an adipogenic, angiogenic, cardiogenic, immunogenic, chondrogenic, endothelial, epithelial, hematopoietic, hepatogenic, myogenic, neurogenic, osteogenic, parenchymal, renal, retinal gene sequence, or a combination thereof. In some cases, a coding RNA may comprise an ectoderm, an endoderm, or a mesoderm gene sequence. In some cases, a coding RNA may comprise an ectoderm gene sequence. In some cases, a coding RNA may comprise an endoderm gene sequence. In some cases, a coding RNA may comprise a mesoderm gene sequence.

[0096] In some cases, a coding RNA may comprise a myogenic gene sequence. In other cases, a coding RNA may comprise MYODI. In some cases, a coding RNA may comprise MYOG. In some cases, a coding RNA may comprise MYF5. In some cases, a coding RNA may comprise MYF6. In some cases, a coding RNA may comprise PAX3. In some cases, a coding RNA may comprise PAX7. In some cases, a coding RNA may comprise MYODI, MYOG, MYF5, MYF6, PAX3, PAX7, a fragment thereof, or a variant thereof. In some cases, a coding RNA may comprise at leasttwo of MYODI, MYOG, MYF5, MYF6, PAX3, PAX7. In some cases, a coding RNA may comprise at least three of MYODI, MYOG, MYF5, MYF6, PAX3, PAX7. In some cases, a coding RNA may comprise at least four of MYODI, MYOG, MYF5, MYF6, PAX3, PAX7. In some cases, a coding RNA may comprise at least five of MYODI, MYOG, MYF5, MYF6, PAX3, PAX7. In some cases, a coding RNA may comprise MYODI, MYOG, MYF5, MYF6, PAX3, and PAX7.

[0097] In some cases, the MYODI coding RNA may comprise at least about one nucleic acid modification in the 5’ untranslated region (UTR), 3’UTR and / or the length of the polyA-tail. In some cases, the nucleic acid modification has no impact on the ultimate amino acid sequence that is translated from the coding RNA. In some cases, the MYODI coding RNA is gene optimized. In some cases, the MYODI coding RNA is codon optimized. Gene optimization may be carried out by any well-known methods in the art. Gene optimization may increase stability, reduce or inhibit degradation. Codon optimization may be carried out by any well-known methods in the art. Codon optimization may promote translation of the coding RNA. In some cases, the MYODI coding RNA is human MYODI coding RNA. In some cases, the MYODI coding RNA is non-human mammalian MYODI coding RNA (e.g. cattle, buffalo, pigs, sheep, deer, etc.), bird MYODI coding RNA (e.g. chicken, ducks, ostrich, turkey, pheasant, etc.), fish MYODI coding RNA (e.g. swordfish, salmon, tuna, sea bass, trout, catfish, etc.), invertebrate MYODI coding RNA (e.g. lobster, crab, shrimp, clams, oysters, mussels, sea urchin, etc.), reptile MYODI coding RNA (e.g. snake, alligator, turtle, etc.), or amphibian MYODI coding RNA (e.g. frogs). In some cases the MYODI coding RNA is porcine MYODI coding RNA.

[0098] In some cases, a coding RNA may comprise an adipogenic gene sequence. In some cases, a coding RNA may comprise PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, CD 137, or any combination thereof. In some cases, a coding RNA encodes PPARY. In some cases, a coding RNA encodes adiponectin. In some cases, a coding RNA encodes FATP1-6. In some cases, a coding RNA encodes FABP4. In some cases, a coding RNA encodes GLUT4. In some cases, a coding RNA encodes Leptin. In some cases, a coding RNA may comprise AdipoRl-2. In some cases, a coding RNA encodes CD137. In some cases, a coding RNA may comprise at least two of PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, CD137. In some cases, a coding RNA may comprise at least three of PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, CD 137. In some cases, a coding RNA may comprise at least four of PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, CD137. In some cases, a coding RNA may comprise at least five of PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2,CD137. In some cases, a coding RNA may comprise at least six of PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, CD 137. In some cases, a coding RNA may comprise at least seven of PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, CD137. In some cases, a coding RNA may comprise at PPARY, adiponectin, FATP1-6, FABP4, GLUT4, Leptin, AdipoRl-2, and CD137.

[0099] In some cases, a coding RNA may comprise an angiogenic gene sequence. In some cases, a coding RNA may comprise a cardiogenic gene sequence. In some cases, a coding RNA may comprise a chondrogenic gene sequence. In some cases, a coding RNA may comprise an endothelial gene sequence. In some cases, a coding RNA may comprise an epithelial gene sequence. In some cases, a coding RNA may comprise a hematopoietic gene sequence. In some cases, a coding RNA may comprise a hepatogenic gene sequence. In some cases, a coding RNA may comprise a neurogenic gene sequence. In some cases, a coding RNA may comprise an osteogenic gene sequence. In some cases, a coding RNA may comprise a parenchymal gene sequence. In some cases, a coding RNA may comprise a renal gene sequence. In some cases, a coding RNA may comprise a retinal gene sequence.

[0100] In some instances, a coding RNA may be monocistronic. In some cases, a coding RNA may be polycistronic. A monocistronic coding RNA may comprise one coding sequence, the coding sequence is configured to be recognized by a ribosome for the translation. A polycistronic coding RNA may comprise at least two coding sequences, each coding sequence is configured to be recognized by a ribosome for the translation of the coding sequence. In some instances, a coding RNA may comprise an RNA-regulatory element. In some cases, a coding RNA comprising an RNA- regulatory element may reduce or inhibit degradation of the coding RNA at least about 5 %, at least about 10 %, at least about 15 %, at least about 20 %, at least about 25 %, at least about 30 %, at least about 35 %, at least about 40 %, at least about 45 %, at least about 50 %, at least about 55 %, at least about 60 %, at least about 65 %, at least about 70 %, at least about 75 %, at least about 80 %, at least about 85 %, at least about 90 %, or at least about 95 %, relative to a coding RNA without the RNA- regulatory element.

[0101] In some cases, an RNA-regulatory element may comprise a transcriptional regulatory element, a co-transcriptional regulatory element, a post-transcriptional regulatory element, a translational regulatory element, or any combination thereof. In some cases, an RNA-regulatory element may comprise a 5 ’-cap, 5’ UTRs, a 3’-UTR, a poly- A tail modification, or any combination thereof. In some cases, an RNA-regulatory element may comprise a 5 ’-cap. In some cases, anRN A- regulatory element may comprise a 5’ UTR. In some cases, an RNA-regulatory element may comprise a 3 ’-UTR. In some cases, an RNA-regulatory element may comprise a poly- A tail modification. In some cases, an RNA-regulatory element may comprise at least two of a 5 ’-cap, 5’ UTRs, 3’-UTRs, or a poly- A tail modification. In some cases, an RNA-regulatory element may comprise a 5 ’-cap, 5’ UTRs, a 3 ’-UTR, and a poly- A tail modification. Other RNA-regulatory elements may comprise an intron sequence, a stop codon sequence, a translation start site, an RNA- localization sequence, or any combination thereof.

[0102] In some instances, a nucleic acid molecule may comprise an inhibitory RNA. An inhibitory RNA can comprise miRNA, siRNA, or shRNA. In some cases, an inhibitory RNA may comprise a polynucleotide sequence that facilitates a reduction of pluripotency of a cell. In some cases, an inhibitory RNA may comprise a polynucleotide sequence that facilitates differentiation by reducing pluripotency of a cell. In some cases, a polynucleotide sequence may comprise a polynucleotide sequence that facilitates knockdown / reduction of a pluripotency of a cell by POUF51 (OCT3 / 4), KLF4, or SOX2, or a complementary sequence thereof.

[0103] In some cases, siRNA may be a class of short, double-stranded RNA non-coding RNA molecules which may interfere with the expression of specific genes with complementary nucleotide sequences. In some cases, siRNA may interfere with gene expression by degrading mRNA after transcription or preventing translation. In some cases, siRNAs may be 10-30 base pairs in length with phosphorylated 5’ ends and hydroxylated 3’ ends. In some cases, siRNAs may target complementary mRNA for degradation, thus preventing translation. In some cases,

[0104] In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of POU5F1 (OCT3 / 4), SOX2, nanog, SSEA-4, KLF4 and / or TRA-1-60. In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of POU5F1 (OCT3 / 4). In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of SOX2. In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of nanog. In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of SSEA-4. In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of KLF4. In some cases, an inhibitory RNA may comprise a sequence or complementary sequence of TRA-1-60. In some cases, an inhibitory RNA may comprise at least two sequences or complementary sequences of POU5F1 (OCT3 / 4), SOX2, nanog, SSEA-4, KLF4 or TRA-1-60. In some cases, an inhibitory RNA may comprises at least three sequences or complementary sequences of POU5F1 (OCT3 / 4), SOX2, nanog, SSEA-4, KLF4 or TRA-1-60. In some cases, an inhibitory RNA may comprises at least foursequences or complementary sequences of POU5F1 (OCT3 / 4), SOX2, nanog, SSEA-4, KLF4 or TRA-1-60. In some cases, an inhibitory RNA may comprises sequences or complementary sequences of POU5F1 (OCT3 / 4), SOX2, nanog, SSEA-4, KLF4 and TRA-1-60.

[0105] A shRNA, in some cases, may be converted to a siRNA by a cell. A miRNA can be a small non-coding RNA molecule that functions in RNA silencing and post-transcriptional regulation of gene expression. In some cases, miRNAs base-pair with complementary sequences within mRNA molecules, silencing the mRNA molecules. In some cases, silencing may be achieved upon binding of the miRNA to the 3’UTR of the target mRNA through cleavage of the mRNA strand into two pieces, destabilization of mRNA through shortening the poly-A tail, or through inefficient translation of the mRNA into proteins by ribosomes. In some cases, modulation of myogenic gene expression may occur through miRNAs. miRNAs that may modulate myogenic gene expression may comprise miR-1, miR-24, miR-26a, miR-27b, miR-29b / c, miR-125b, miR-133, miR-181, miR-206, miR- 208b / 499, miR-214, miR-221 / 222, miR-322 / 424, mi486, or miR-503. In some cases, a miRNA may be expressed in a cell type. In some cases, a miRNA may be specifically expressed in a cell type. miRNAs may be specifically expressed in cardiac and skeletal muscles under the control of the myogenic transcription factors SRF, MyoD or MEF2 where they may regulate processes of skeletal myogenesis such as myoblast / satellite cell proliferation and differentiation.

[0106] In some cases, one or more genes may be targeted and modulated with one, two, or a plurality of nucleic acid molecules. In some cases, one or more genes may comprise greater or equal to about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 genes, or more. In some cases, modulating expression of one or more genes in a cell may comprise enhancing expression of a first gene of the at least about two genes, and inhibiting expression of a second gene of the at least about two genes.

[0107] In some instances, a nucleic acid molecule may be modified. In some cases, a nucleic acid molecule may be chemically modified (i.e., the nucleic acid molecule comprises a chemical modification). In some instances, chemically modified nucleic acid molecule may reduce or inhibit degradation of the nucleic acid molecule at least about 5 %, at least about 10 %, at least about 15 %, at least about 20 %, at least about 25 %, at least about 30 %, at least about 35 %, at least about 40 %, at least about 45 %, at least about 50 %, at least about 55 %, at least about 60 %, at least about 65 %, at least about 70 %, at least about 75 %, at least about 80 %, at least about 85 %, at least about 90 %, at least about 95 % or more, relative to a nucleic acid molecule not chemically modified.

[0108] In some instances, a nucleic acid may have a length of at least about 5 nucleotides, 10 nucleotides, 50 nucleotides, 100 nucleotides, 500 nucleotides, 1000 nucleotides, 5000 nucleotides,10000 nucleotides, or more. In some instances, a nucleic acid may have a length of at most about 5 nucleotides, 10 nucleotides, 50 nucleotides, 100 nucleotides, 500 nucleotides, 1000 nucleotides, 5000 nucleotides, or 10000 nucleotides.Lipoplex / transfection complex

[0109] In some instances, a saccharide, a lipid, or a lipoplex may facilitate an uptake of a nucleic acid by a cell. In some cases, a saccharide, a lipid, or a polymer comprising a positive charge may form an ionic bond with a nucleic acid molecule. A nucleic acid molecule bound by a saccharide or associated into a lipoplex may have a reduced negative charge. A nucleic acid molecule bound by a saccharide or associated into a lipoplex may have a neutral charge. A nucleic acid molecule bound by a saccharide or associated into a lipoplex may have a positive charge. The reduced negative charge, neutral charge, or positive charge of a nucleic acid molecule (and the saccharide or the lipoplex) may facilitate its uptake by a cell. In some cases, a saccharide may bind a nucleic acid via coacervation. For example, reducing the net negative charge of a composition comprising a nucleic acid molecule — via (1) the binding or association of the nucleic acid molecule with a saccharide; (2) formation of a lipoplex from the nucleic acid molecule and the saccharide; (3) a combination thereof — may facilitate the update of the nucleic acid molecule by the cell, wherein a plasma membrane may carry a negative charge. Association of a nucleic acid molecule with a saccharide or formation of a lipoplex from the nucleic acid molecule and the saccharide may reduce the amount (number, weight, of mass) of the nucleic acid molecule to be used for contacting the cell (to allow for the cell to uptake the nucleic acid molecule), relative to the amount of the nucleic acid molecule to be used for contacting the cell without the saccharide or formation of the lipoplex.

[0110] In some instances, a saccharide molecule, a lipid molecule, and a nucleic acid molecule as described herein are configured to self-assemble with each other. In some cases, a saccharide molecule, a lipid molecule, and a nucleic acid molecule are configured to associate with each other. In some cases, a saccharide may bind a nucleic acid molecule. In some cases, a saccharide molecule, a lipid molecule, and a nucleic acid molecule are configured to form a transfection complex or transfection complex.

[0111] Self-assembly of two molecular entities may comprise the initiation and formation of a distinct molecular entity from the two molecular entities without an assistance of other molecular entity (e.g., a catalyst). In some cases, a saccharide molecule, a lipid molecule, and a nucleic acid molecule as described herein may associate with each other based at least on the electrostaticinteraction between them. For example, the nucleic acid molecule may be anionic and the saccharide and / or lipid may be cationic. The electrostatic interaction between the anionic charge of the nucleic acid molecule and the cationic charge of the saccharide / lipid may facilitate the self-assembly of the nucleic acid molecule and the saccharide into a complex. The electrostatic interaction between the anionic charge of the nucleic acid molecule and the cationic charge of the saccharide / lipid may facilitate the formation of a transfection complex. Adjusting the charges of a saccharide / lipid (e.g., via modification, functionalization, or derivatization of the saccharide) may control the association, self-assembly, and / or formation of the transfection complex.

[0112] In some cases, a nucleic acid molecule is configured to be at or near a surface of a transfection complex. The nucleic acid molecule is at or near the surface of the transfection complex. A nucleic acid molecule is configured to be adsorbed onto a surface of the saccharides / lipids of a transfection complex. The nucleic acid molecule may be adsorbed onto the saccharides / lipids of a surface a transfection complex. In some cases, a nucleic acid molecule is configured to be encapsulated within a transfection complex. A nucleic acid molecule may be encapsulated within a transfection complex. In some cases, a nucleic acid molecule is present inside a transfection complex. In some cases, a nucleic acid molecule is present outside a transfection complex. In some cases, a nucleic acid molecule is present inside and outside a transfection complex. The nucleic acid molecule may also be intercalated with other molecules, for example, the saccharide / lipid, of the transfection complex. The configuration of the nucleic acid molecule and the saccharide / lipid of a transfection complex may be random. The configuration of the nucleic acid molecule and the saccharide / lipid of a transfection complex may be ordered. In some cases, a transfection complex is regularly shaped, irregularly shaped, spherical, linear, or branched. In some cases, a transfection complex is regularly shaped, irregularly shaped or branched. In some cases, a transfection complex is regularly shaped. In some cases, a transfection complex is irregularly shaped. In some cases, a transfection complex is branched. In some cases, a transfection complex is spherical or linear. In some cases, a transfection complex is spherical. In some cases, a transfection complex is linear. A regularly shape may comprise any orthogonal shapes. A regularly shape may comprise any polygonal shapes (comprising 3, 4, 5, 6, 7, 8, 9, 10 or more edges). An irregular shape may comprise shapes that are not regular shapes.

[0113] In some instances, a size of a transfection complex may be measured by a diameter of the transfection complex. In some cases, a diameter of a transfection complex may comprise an apparent diameter of the transfection complex. In some cases, an apparent diameter of a transfection complexmay comprise an average diameter of a distribution of the diameters of a plurality of transfection complexes. In some cases, an apparent diameter of a transfection complex may comprise a representative diameter of a plurality of transfection complexes. In some cases, an apparent diameter of a transfection complex may comprise a mean diameter of a plurality of transfection complexes. In some cases, an apparent diameter of a transfection complex may comprise a median diameter of a plurality of transfection complexes. In some cases, an apparent diameter of a transfection complex may comprise a mode diameter of a plurality of transfection complexes. The representative diameter may also comprise a quantile diameter or a range diameter of a plurality of transfection complexes. The representative diameter may also be described as a distribution of a plurality of diameters of a plurality of transfection complexes.

[0114] In some cases, a diameter of a transfection complex may be measured by dynamic light scattering (DLS), gel filtration chromatography, analytical ultracentrifugation or microscopy. In some cases, a diameter of a transfection complex formed in the compositions may be measured by a Dynamic light scattering (DLS). DLS may determine the size of a particle (e.g., a transfection complex) by measuring the random changes in the intensity of light scattered from a suspension or solution containing the molecule. DLS may measure the particle size by sensing the Brownian motion of particles. Since the Brownian motion velocities are higher for smaller particles, the Doppler spectral broadening of the scattered light may be size dependent. DLS may have a resolution of about 0.01 nanometer (nm), 0.1 nm, or 1 nm. In other instances, a transfection complex formed is observed and / or measured by cryo-electron microscopy. In some cases, a transfection complex may be loaded in the form of a thin membrane on a grid and frozen at -80 °C, -90 °C, -100 °C, -110 °C, -120 °C, - 130 °C, -140 °C, -150 °C, -160 °C, -170 °C, -180 °C, -190 °C, -200 °C or lower temperature.

[0115] A nucleic acid molecule described herein may be present at a concentration at least about 1 picomolar (pM), 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 nanomolar (nM), 2 nM, 5 nM, 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 500 nM, 1 micromolar (pM), 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 millimolar (mM), 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 100 mM, 200 mM, 500 mM, 1 molar (M) or more within the composition. A nucleic acid molecule described herein described herein may be present at a concentration at most about 1 pM, 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 nM, 2 nM, 5 nM, 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 500 nM, 1 pM, 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500pM, 1 mM, 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 100 mM, 200 mM, 500 mM, or 1 M within the composition.

[0116] A saccharide described herein may be present at a concentration at least about 1 picomolar (pM), 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 nanomolar (nM), 2 nM, 5 nM, 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 500 nM, 1 micromolar (pM), 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 millimolar (mM), 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 100 mM, 200 mM, 500 mM, 1 molar (M) or more within the composition. A saccharide described herein may be present at a concentration at most about 1 pM, 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 nM, 2 nM, 5 nM, 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 500 nM, 1 pM, 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 mM, 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 100 mM, 200 mM, 500 mM, or 1 M within the composition.

[0117] A lipid molecule described herein may be present at a concentration at least about 1 picomolar (pM), 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 nanomolar (nM), 2 nM, 5 nM, 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 500 nM, 1 micromolar (pM), 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 millimolar (mM), 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 100 mM, 200 mM, 500 mM, 1 molar (M) or more within the composition. A lipid molecule described herein may be present at a concentration at most about 1 pM, 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 nM, 2 nM, 5 nM, 10 nM, 20 nM, 50 nM, 100 nM, 200 nM, 500 nM, 1 pM, 2 pM, 5 pM, 10 pM, 20 pM, 50 pM, 100 pM, 200 pM, 500 pM, 1 mM, 2 mM, 5 mM, 10 mM, 20 mM, 50 mM, 100 mM, 200 mM, 500 mM, or 1 M within the composition.

[0118] The saccharide molecule and the nucleic acid molecule within a composition (such as a transfection complex), as described herein, may be present in a ratio, by mass, of at least about 0.001, 0.002, 0.003, 0.004, 0.005, 0.006, 0.007, 0.008, 0.009, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 200, 300, 400, 500, 600, 700, 800, 900, or 1000. The saccharide molecule and the nucleic acid molecule within a composition (such as a transfection complex), as described herein, may be present in a ratio, by mass, of at most about 0.001, 0.002, 0.003, 0.004, 0.005, 0.006, 0.007, 0.008, 0.009, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45,46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 200, 300, 400, 500, 600, 700, 800, 900, or 1000. The lipid molecule and the nucleic acid molecule within a composition (such as a transfection complex), as described herein, may be present in a ratio, by mass, of at least about 0.001, 0.002, 0.003, 0.004, 0.005, 0.006, 0.007, 0.008, 0.009, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68,69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95,96, 97, 98, 99, 100, 200, 300, 400, 500, 600, 700, 800, 900, or 1000. The lipid molecule and the nucleic acid molecule within a composition (such as a transfection complex), as described herein, may be present in a ratio, by mass, of at most about 0.001, 0.002, 0.003, 0.004, 0.005, 0.006, 0.007, 0.008, 0.009, 0.01, 0.02, 0.03, 0.04, 0.05, 0.06, 0.07, 0.08, 0.09, 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64,65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91,92, 93, 94, 95, 96, 97, 98, 99, 100, 200, 300, 400, 500, 600, 700, 800, 900, or 1000. The ratio between any two components of a composition as described herein can be determined by using the methods described elsewhere in this disclosure for measuring the transfection efficiency of a transfection complex (or the nucleic acid molecule contained therewithin) with a particular cell type (see, for example, Example 1).

[0119] In some cases, when preparing for the transfection complex, the saccharide molecule, the lipid molecule, and / or the nucleic acid molecule may be dissolved or maintained in a solvent. The solvent may be an organic solvent. The solvent may be a hydrophobic solvent. The solvent may comprise a polar organic solvent. The solvent may comprise a non-polar organic solvent. The solvent may be miscible with water. For example, the solvent may comprise a polar organic solvent miscible with water. The hydrophobicity of the solvent described herein may have a beneficial advantage for facilitating the homogenization of the lipid molecule (or the lipid moiety of the saccharide molecule). Homogenization of the lipid molecule may facilitate the formation of the transfection complex. In some cases, the solvent may comprise an inorganic solvent. The solvent may be an inorganic solvent. The solvent may comprise a polar inorganic solvent. The solvent may comprise a non-polar inorganic solvent. In some cases, the solvent may be a hydrophilic solvent. Insome cases, the hydrophilicity of the solvent may have a beneficial advantage for dissolving the nucleic acid or saccharide. Dissolution of the nucleic acid molecule or saccharide may facilitate the formation of the transfection complex. In some cases, the solvent may be a food-grade substance or safe for consumption by a human subject. In some cases, the solvent may be insert. In some cases, the solvent may be safe for therapeutic uses. The solvent may be an alcohol-based solvent. The solvent may be DMSO, DMF, ethanol, propanol, 2-propanol, ethylene glycol, a functional variant thereof, a derivative thereof, or any combination thereof. The solvent may be DMSO, a functional variant thereof, or a derivative thereof. The solvent may be DMF, a functional variant thereof, or a derivative thereof. The solvent may be ethanol, a functional variant thereof, or a derivative thereof. The solvent may be propanol, a functional variant thereof, or a derivative thereof. The solvent may be 2-propanol, a functional variant thereof, or a derivative thereof. The solvent may be ethylene glycol, a functional variant thereof, or a derivative thereof. The solvent may be DMSO. The solvent may be DMF. The solvent may be an ethanol. The solvent may be propanol. The solvent may be 2- propanol. The solvent may be ethylene glycol.

[0120] A solvent described herein may be present in an amount, by weight, of at least about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %,25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %, 34 %, 35 %, 36 %, 37 %, 38 %, 39 %, 40%, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49 %, 50 %, 51 %, 52 %, 53 %, 54 %, 55 %,56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %, 65 %, 66 %, 67 %, 68 %, 69 %, 70 %, 71%, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80 %, 81 %, 82 %, 83 %, 84 %, 85 %, 86 %,87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97 %, 98 %, 99 %, or 100 % of the composition as described herein. A solvent described herein may be present in an amount, by weight, of at most about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %,20 %, 21 %, 22 %, 23 %, 24 %, 25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %, 34 %, 35%, 36 %, 37 %, 38 %, 39 %, 40 %, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49 %, 50 %,51 %, 52 %, 53 %, 54 %, 55 %, 56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %, 65 %, 66%, 67 %, 68 %, 69 %, 70 %, 71 %, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80 %, 81 %,82 %, 83 %, 84 %, 85 %, 86 %, 87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97%, 98 %, 99 %, or 100 % of the composition as described herein. A solvent described herein may be present in an amount, by volume, of at least about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %,0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %, 25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31%, 32 %, 33 %, 34 %, 35 %, 36 %, 37 %, 38 %, 39 %, 40 %, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %,47 %, 48 %, 49 %, 50 %, 51 %, 52 %, 53 %, 54 %, 55 %, 56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62%, 63 %, 64 %, 65 %, 66 %, 67 %, 68 %, 69 %, 70 %, 71 %, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %,78 %, 79 %, 80 %, 81 %, 82 %, 83 %, 84 %, 85 %, 86 %, 87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93%, 94 %, 95 %, 96 %, 97 %, 98 %, 99 %, or 100 % of the composition as described herein. A solvent described herein may be present in an amount, by volume, of at most about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %, 25 %,26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %, 34 %, 35 %, 36 %, 37 %, 38 %, 39 %, 40 %, 41%, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49 %, 50 %, 51 %, 52 %, 53 %, 54 %, 55 %, 56 %,57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %, 65 %, 66 %, 67 %, 68 %, 69 %, 70 %, 71 %, 72%, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80 %, 81 %, 82 %, 83 %, 84 %, 85 %, 86 %, 87 %,88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97 %, 98 %, 99 %, or 100 % of the composition as described herein.

[0121] Ethanol described herein may be present in an amount, by weight, of at least about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %,25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %, 34 %, 35 %, 36 %, 37 %, 38 %, 39 %, 40%, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49 %, 50 %, 51 %, 52 %, 53 %, 54 %, 55 %,56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %, 65 %, 66 %, 67 %, 68 %, 69 %, 70 %, 71%, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80 %, 81 %, 82 %, 83 %, 84 %, 85 %, 86 %,87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97 %, 98 %, 99 %, or 100 % of the composition as described herein. Ethanol may be present in an amount, by weight, of at most about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %, 25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %, 34 %, 35 %, 36 %, 37 %, 38 %, 39%, 40 %, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49 %, 50 %, 51 %, 52 %, 53 %, 54 %,55 %, 56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %, 65 %, 66 %, 67 %, 68 %, 69 %, 70%, 71 %, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80 %, 81 %, 82 %, 83 %, 84 %, 85 %,86 %, 87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97 %, 98 %, 99 %, or 100 %of the composition as described herein. Ethanol described herein may be present in an amount, by volume, of at least about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %, 25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %, 34 %, 35%, 36 %, 37 %, 38 %, 39 %, 40 %, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49 %, 50 %,51 %, 52 %, 53 %, 54 %, 55 %, 56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %, 65 %, 66%, 67 %, 68 %, 69 %, 70 %, 71 %, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80 %, 81 %,82 %, 83 %, 84 %, 85 %, 86 %, 87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %, 96 %, 97%, 98 %, 99 %, or 100 % of the composition as described herein. Ethanol may be present in an amount, by volume, of at most about, 0.1 %, 0.2 %, 0.3 %, 0.4 %, 0.5 %, 0.6 %, 0.7 %, 0.8 %, 0.9 %, 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %, 25 %, 26 %, 27 %, 28 %, 29 %, 30 %, 31 %, 32 %, 33 %,34 %, 35 %, 36 %, 37 %, 38 %, 39 %, 40 %, 41 %, 42 %, 43 %, 44 %, 45 %, 46 %, 47 %, 48 %, 49%, 50 %, 51 %, 52 %, 53 %, 54 %, 55 %, 56 %, 57 %, 58 %, 59 %, 60 %, 61 %, 62 %, 63 %, 64 %,65 %, 66 %, 67 %, 68 %, 69 %, 70 %, 71 %, 72 %, 73 %, 74 %, 75 %, 76 %, 77 %, 78 %, 79 %, 80%, 81 %, 82 %, 83 %, 84 %, 85 %, 86 %, 87 %, 88 %, 89 %, 90 %, 91 %, 92 %, 93 %, 94 %, 95 %,96 %, 97 %, 98 %, 99 %, or 100 % of the composition as described herein.

[0122] A transfection complex may be maintained in a buffer. The buffer may have a pH of at least about 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, 9 or more. The buffer may have a pH of at most about 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8, 8.1, 8.2, 8.3, 8.4, 8.5, 8.6, 8.7, 8.8, 8.9, or 9. The buffer The buffer may comprise a saline. The buffer may comprise phosphate-buffered saline (PBS). The buffer may also comprise HEPES or Tris-buffered saline (TBS). The buffer may also be the alcohol-based solvent. The buffer may be ethanol.

[0123] A nucleic acid, a saccharide, a lipid, or a combination thereof may be incubated at about -80 °C, -50 °C, -20 °C, -10 °C, -9 °C, -8 °C, -7 °C, -6 °C, -5 °C, -4 °C, -3 °C, -2 °C, -1 °C, 0 °C, 1 °C, 2 °C, 3 °C, 4 °C, 5 °C, 6 °C, 7 °C, 8 °C, 9 °C, 10 °C, 11 °C, 12 °C, 13 °C, 14 °C, 15 °C, 16 °C, 17 °C,18 °C, 19 °C, 20 °C, 21 °C, 22 °C, 23 °C, 24 °C, 25 °C, 26 °C, 27 °C, 28 °C, 29 °C, 30 °C, 31 °C, 32°C, 33 °C, 34 °C, 35 °C, 36 °C, 37 °C or higher to form a transfection complex. A nucleic acid, a saccharide, a lipid, or a combination thereof may be incubated for at about 1 minute, 2 minutes, 3 minutes, 4 minutes, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, 10 minutes, 11 minutes,12 minutes, 13 minutes, 14 minutes, 15 minutes, 16 minutes, 17 minutes, 18 minutes, 19 minutes, 20 minutes, 21 minutes, 22 minutes, 23 minutes, 24 minutes, 25 minutes, 26 minutes, 27 minutes, 28 minutes, 29 minutes, 30 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours or more to form a transfection complex.

[0124] In some instances, an apparent diameter of a transfection complex is at least about 5.0 nanometer (nm). In some instances, an apparent diameter of a transfection complex is at least about 10.0 nm. In some instances, an apparent diameter of a transfection complex is at least about 1.0 nm, at least about 1.5 nm, at least about 2.0 nm, at least about 2.5 nm, at least about 3.0 nm, at least about3.5 nm, at least about 4.0 nm, at least about 4.5 nm, at least about 5.0 nm, at least about 5.5 nm, at least about 6.0 nm, at least about 6.5 nm, at least about 7.0 nm, at least about 7.5 nm, at least about 8.0 nm, at least about 8.5 nm, at least about 9.0, at least about 9.5 nm, at least about 10.0 nm, at least about 10.5 nm, at least about 11.0 nm, at least about 11.5 nm, at least about 12.0 or more. In some instances, an apparent diameter of a transfection complex is at most about 5000 nm. In some instances, an apparent diameter of a transfection complex is at most about 800 nm. In some instances, an apparent diameter of a transfection complex is at most about 600 nm. In some instances, an apparent diameter of a transfection complex is at most about 7000 nm, at most about 6000 nm, at most about 5000 nm, at most about 4000 nm, at most about 3000 nm, at most about 2000 nm, at most about 1000 nm, at most about 900 nm, at most about 800 nm, at most about 700 nm, at most about 600 nm, at most about 500 nm, at most about 400 nm, at most about 300 nm, at most about 200 nm, or at most about 100 nm.

[0125] In some instances, a transfection complex may comprise a nanoparticle comprising the saccharide molecule, the lipid molecule (for example, at least two different lipid molecules as described herein), and the nucleic acid molecule as described herein. In some cases, a nanoparticle may be at least about 5.0 nm. In some cases, a nanoparticle may be at least about 10.0 nm. In some cases, a nanoparticle may be at least about 1.0 nm, at least about 1.5 nm, at least about 2.0 nm, at least about 2.5 nm, at least about 3.0 nm, at least about 3.5 nm, at least about 4.0 nm, at least about4.5 nm, at least about 5.0 nm, at least about 5.5 nm, at least about 6.0 nm, at least about 6.5 nm, at least about 7.0 nm, at least about 7.5 nm, at least about 8.0 nm, at least about 8.5 nm, at least about 9.0, at least about 9.5 nm, at least about 10.0 nm, at least about 10.5 nm, at least about 11.0 nm, at least about 11.5 nm, at least about 12.0 or more. In some cases, a nanoparticle may be at most about 10.0 nm. In some cases, a nanoparticle may be at most about 1.0 nm, at most about 1.5 nm, at most about 2.0 nm, at most about 2.5 nm, at most about 3.0 nm, at most about 3.5 nm, at most about 4.0nm, at most about 4.5 nm, at most about 5.0 nm, at most about 5.5 nm, at most about 6.0 nm, at most about 6.5 nm, at most about 7.0 nm, at most about 7.5 nm, at most about 8.0 nm, at most about 8.5 nm, at most about 9.0, at most about 9.5 nm, at most about 10.0 nm, at most about 10.5 nm, at most about 11.0 nm, at most about 11.5 nm, or at most about 12.0.

[0126] In some cases, a nanoparticle may be a colloidal nanoparticle. The colloidal lipid particles used herein may be at least substantially non-toxic to a subject or a cell. In some cases, the lipid particles have a mean diameter of from about 40 nm to about 150 nm. In some cases, the diameter is from about 50 nm to about 140 nm, about 60 nm to about 130 nm, or to about 70 nm to about 120 nm. In some cases, a colloidal nanoparticle may be at most about 5000 nm. In some cases, a colloidal nanoparticle may be at most about 800 nm. In some cases, a colloidal nanoparticle may be at most about 600 nm. In some cases, a colloidal nanoparticle may be at most about 7000 nm, at most about 6000 nm, at most about 5000 nm, at most about 4000 nm, at most about 3000 nm, at most about 2000 nm, at most about 1000 nm, at most about 900 nm, at most about 800 nm, at most about 700 nm, at most about 600 nm, at most about 500 nm, at most about 400 nm, at most about 300 nm, at most about 200 nm, or at most about 100 nm. In some cases, a colloidal nanoparticle may be at least about 600 nm. In some cases, a colloidal nanoparticle may be at least about 7000 nm, at least about 6000 nm, at least about 5000 nm, at least about 4000 nm, at least about 3000 nm, at least about 2000 nm, at least about 1000 nm, at least about 900 nm, at least about 800 nm, at least about 700 nm, at least about 600 nm, at least about 500 nm, at least about 400 nm, at least about 300 nm, at least about 200 nm, or at least about 100 nm.

[0127] In some cases, the corresponding cationic charge density of a nanoparticle described herein may be at least about 0.5 mequiv / g, at least about 1 mequiv / g, at least about 1.5 mequiv / g, at least about 2 mequiv / g, at least about 2.5 mequiv / g, at least about 3 mequiv / g, at least about 3.5 mequiv / g, at least about 4 mequiv / g, at least about 4.5 mequiv / g, at least about 5 mequiv / g, at least about 5.5 mequiv / g, at least about 6 mequiv / g, at least about 6.5 mequiv / g, at least about 7 mequiv / g, at least about 7.5 mequiv / g, at least about 8 mequiv / g, at least about 8.5 mequiv / g, at least about 9 mequiv / g, at least about 9.5 mequiv / g, at least about 10 mequiv / g, at least about 10.5 mequiv / g, at least about 11 mequiv / g, at least about 11.5 mequiv / g, at least about 12 mequiv / g, at least about 12.5 mequiv / g, at least about 13 mequiv / g, at least about 13.5 mequiv / g, at least about 14 mequiv / g, at least about 14.5 mequiv / g, at least about 15 mequiv / g, at least about 15.5 mequiv / g, at least about 16 mequiv / g, at least about 16.5 mequiv / g, at least about 17 mequiv / g, at least about 17.5 mequiv / g, at least about 18 mequiv / g, at least about 18.5 mequiv / g, at least about 19 mequiv / g, at least about 19.5 mequiv / g,or at least about 20 mequiv / g. In some cases, the corresponding cationic charge density of a nanoparticle described herein may be at most about 0.5 mequiv / g, at most about 1 mequiv / g, at most about 1.5 mequiv / g, at most about 2 mequiv / g, at most about 2.5 mequiv / g, at most about 3 mequiv / g, at most about 3.5 mequiv / g, at most about 4 mequiv / g, at most about 4.5 mequiv / g, at most about 5 mequiv / g, at most about 5.5 mequiv / g, at most about 6 mequiv / g, at most about 6.5 mequiv / g, at most about 7 mequiv / g, at most about 7.5 mequiv / g, at most about 8 mequiv / g, at most about 8.5 mequiv / g, at most about 9 mequiv / g, at most about 9.5 mequiv / g, at most about 10 mequiv / g, at most about 10.5 mequiv / g, at most about 11 mequiv / g, at most about 11.5 mequiv / g, at most about 12 mequiv / g, at most about 12.5 mequiv / g, at most about 13 mequiv / g, at most about 13.5 mequiv / g, at most about 14 mequiv / g, at most about 14.5 mequiv / g, at most about 15 mequiv / g, at most about 15.5 mequiv / g, at most about 16 mequiv / g, at most about 16.5 mequiv / g, at most about 17 mequiv / g, at most about 17.5 mequiv / g, at most about 18 mequiv / g, at most about 18.5 mequiv / g, at most about 19 mequiv / g, at most about 19.5 mequiv / g, or at most about 20 mequiv / g. In some cases, the corresponding cationic charge density of a nanoparticle described herein may be from 0.005 to 2000 mequiv / g. In some cases, the corresponding cationic charge density of a nanoparticle described herein may be from 0.05 to 200 mequiv / g. In some cases, the corresponding cationic charge density of a nanoparticle described herein may be from 0.5 to 20 mequiv / g.

[0128] In some instances, a composition described herein may comprise a gold particle, a calcium phosphate nanoparticle, an iron oxide particle, or a combination thereof. In some cases, a calcium phosphate (Cap) particle may comprise a silica shell, a PEI (poly(ethyleneimine)) shell, or a combination thereof. In some cases, calcium phosphate particle may comprise a triple shell calcium phosphate particle (cap-nucleic acid-CaP-PEI), a calcium phosphate particle with a silica shell (Cap-nucleic acid-PEI-SiO2-SH), a calcium phosphate with a single-shell Cap-PEI, or a combination thereof (see, for example, Neuhaus B et al., RSV Adv., 2016, 6, 18102, which is herein incorporated by reference in its entirety).

[0129] In some instances, a molar ratio of a transfection complex may be measured. In some cases, the molar ratio of a transfection complex may be a molar ratio of polycation amino groups of the transfection complex to nucleic acid phosphate groups of the transfection complex. In some cases, a molar ratio of a transfection complex is at least about 1, at least about 2, at least about 3, at least about 4, at least about 5, at least about 6, at least about 7, at least about 8, at least about 9, at least about 10, at least about 15, at least about 20, at least about 25, at least about 30, at least about 35, at least about 40, at least about 45, at least about 50, at least about 51, at least about 52, at least about53, at least about 54, at least about 55, at least about 56, at least about 57, at least about 58, at least about 59, or at least about 60. In some cases, a molar ratio of a transfection complex is at most about 1, at most about 2, at most about 3, at most about 4, at most about 5, at most about 6, at most about 7, at most about 8, at most about 9, at most about 10, at most about 15, at most about 20, at most about 25, at most about 30, at most about 35, at most about 40, at most about 45, at most about 50, at most about 51, at most about 52, at most about 53, at most about 54, at most about 55, at most about 56, at most about 57, at most about 58, at most about 59, or at most about 60. In some cases, a molar ratio of a transfection complex is at least about 1. In some cases, a molar ratio of a transfection complex is at most about 60. In some cases, a molar ratio of a transfection complex is from about 1 to about 60.

[0130] In some cases, a nanoparticles can be stored at about 4 °C. In some cases, a nanoparticles can be stored at about -196°C, -150°C, -80 °C, -50 °C, -20 °C, -15 °C, -10 °C, -5 °C, 0 °C, 1 °C, 2 °C, 3 °C, 4 °C, 5 °C, 6 °C, 7 °C, 8 °C, 9 °C, 10 °C, 15 °C, 20 °C, 25 °C, 30 °C, 37 °C, 40 °C or higher. In some cases, a nanoparticles can be stored from about 7 top about 14 days. In some cases, a nanoparticles can be stored at about 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, 8 days, 9 days, 10 days, 11 days, 12 days, 13 days, 14 days, 1 month, 6 months, 1 year, or 5 years. In some cases, a nanoparticle may be lyophilized.

[0131] In some instances, a saccharide, a lipid, or a transfection complex is configured to inhibit or reduce or inhibit degradation of a nucleic acid molecule. In some instances, a saccharide, a lipid, or a transfection complex is configured to inhibit or reduce nuclease degradation of a nucleic acid molecule. In some cases, a saccharide (or the saccharide within a transfection complex) may bind to a nucleic acid molecule to protect a nucleic acid against cleavage by nuclease. In some cases, a nuclease may comprise an exonuclease or an endonuclease. In some instances, a nuclease may comprise deoxyribonuclease or ribonuclease. In some cases, a nuclease may comprise topoisomerases, recombinases, ribozymes, and RNA splicing enzymes. In some cases, protection of nucleic acid molecules against degradation increases the amount of the nucleic acid molecules available for contacting the cell or being taken up by the cell.

[0132] In some cases, a saccharide, a lipid, or a transfection complex may reduce or inhibit degradation of a nucleic acid molecule at least about 10 %, relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. In some cases, a saccharide, a lipid, or a transfection complex may reduce or inhibit degradation of a nucleic acid molecule at least about 50 %, relative to a nucleic acid molecule not bound by the saccharide. Insome cases, a saccharide may reduce or inhibit degradation of a nucleic acid molecule at least about 90 %, relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. In some cases, a saccharide or transfection complex may reduce or inhibit degradation of a nucleic acid molecule at least about 5 %, at least about 10 %, at least about 15 %, at least about 20 %, at least about 25 %, at least about 30 %, at least about 35 %, at least about 40 %, at least about 45 %, at least about 50 %, at least about 55 %, at least about 60 %, at least about 65 %, at least about 70 %, at least about 75 %, at least about 80 %, at least about 85 %, at least about 90 %, or at least about 95 %, relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. In some cases, a saccharide or transfection complex may reduce or inhibit degradation of a nucleic acid molecule at most about 10 %, relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. In some cases, a saccharide or transfection complex may reduce or inhibit degradation of a nucleic acid molecule at most about 50 %, relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. In some cases, a saccharide or transfection complex may reduce or inhibit degradation of a nucleic acid molecule at most about 90 %, relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. In some cases, a saccharide or transfection complex may reduce or inhibit degradation of a nucleic acid molecule at most about 5 %, at most about 10 %, at most about 15 %, at most about 20 %, at most about 25 %, at most about 30 %, at most about 35 %, at most about 40 %, at most about 45 %, at most about 50 %, at most about 55 %, at most about 60 %, at most about 65 %, at most about 70 %, at most about 75 %, at most about 80 %, at most about 85 %, at most about 90 %, at most about 95 %, or 100 % relative to a nucleic acid molecule not bound by the saccharide or not associated with the transfection complex. The reduction or inhibition of degradation of a nucleic acid molecule may be measured by the amount of nucleic acid molecule present / absent or by the amount of the time it takes for a nucleic acid molecule to be degraded.Conditions

[0133] In some cases, a method may comprise contacting a cell with a nucleic acid molecule, saccharide, lipid, or transfection complex as described herein in a condition sufficient for the cell to uptake the nucleic acid molecule, saccharide, lipid, or transfection complex as described herein.

[0134] In some cases, the method may comprise contacting a cell that is maintained in a cell culture. Thus, the method may comprise contacting a cell in vitro or ex vivo. In some cases, the sufficientcondition may comprise growing a cell in a 2D or 3D culture or environment. In some cases, the cell may be grown in the culture without a scaffold. In other cases, the cell may also be grown in the culture with a scaffold. In some cases, a cell may be viable. A cell may also be healthy. In some cases, a sufficient condition may comprise a population of viable cells before the contacting. The population of cells may comprise at least about 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 99 % or more viable cells. The population of cells may also comprise from about 80 % to about 90 % viable cells. The population of cells may comprise at most about 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, or 100 % viable cells.

[0135] In some cases, the method may comprise contacting a cell or an organism comprising the cell. Thus, the contacting may comprise contacting a cell in vivo. In some cases, a lipoplex may be prepared immediately prior to the contacting. In some cases, a lipoplex may be prepared at most about 5 hours, 4 hours, 3 hours, 2 hours, 1 hour, 45 minutes, 30 minutes, 15 minutes, or 5 minutes before the contacting. In some cases, a nucleic acid molecule may be kept at a temperature at most about -196°C, -150°C, -80 °C, -50 °C, -20 °C, -15 °C, -10 °C, -5 °C, 0 °C, 1 °C, 2 °C, 3 °C, 4 °C, 5 °C, 6 °C, 7 °C, 8 °C, 9 °C, 10 °C prior to the formation of the lipoplex.

[0136] In some cases, a sufficient condition may comprise contacting a cell or the organism comprising the cell with a nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof for at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 hours or more. In some cases, a sufficient condition may comprise contacting a cell or the organism comprising the cell with a nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof for at least about 1, 2, 3, 4, 5 days or more. In some cases, the contacting may be carried out in a 2D or 3D culture condition.

[0137] In some cases, the cell may be cultured in a first media for at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours after the contacting. In some cases, the cell may be cultured in a first media for at least about 1, 2, 3, 4, 5 days after the contacting. In some cases, the cell may be cultured in a second media at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24 hours after the cell is cultured in the first media. In some cases, the cell may be cultured in a second media at least about 1, 2, 3, 4, 5 days after the cell is cultured in the first media. In some cases, the cell may be cultured in a second media for at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54,55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96 hours or more. In some cases, the cell may be cultured in a second media for at least about 5, 6, 7, 8, 9, 10 days or more. In some cases, the first media may be the same as the second media. In some cases, the first media may be different from the second media. In some cases, the first or the second media may be any media described herein.

[0138] A sufficient condition may comprise contacting or incubating the cell or the organism comprising the cell with the nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof with at most about 25 %, 24 %, 23 %, 22 %, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or lower atmospheric oxygen. A sufficient condition may comprise contacting or incubating the cell with the nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof with at least about 1 %, 2 %, 3 %, 4 %, 5 %, 6 %, 7 %, 8 %, 9 %, 10 %, 11 %, 12 %, 13 %, 14 %, 15 %, 16 %, 17 %, 18 %, 19 %, 20 %, 21 %, 22 %, 23 %, 24 %, 25 % or more atmospheric oxygen.

[0139] A sufficient condition may comprise contacting or incubating the cell or the organism comprising the cell with the nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof with at most about 1 %, 1.5 %, 2 %, 2.5 %, 3 %, 3.5 %, 4 %, 4.5 %, 5 %, 5.5 %, 6 %, 6.5 %, 7 %, 7.5 %, 8 %, 8.5 %, 9 %, 9.5 %, 10 %, 10.5 %, 11 %, 11.5 %, 12 %, 12.5 %, 13 %, 13.5 %, 14 %, 14.5 %, or 15 % atmospheric carbon dioxide (CO2).

[0140] The methods provided herein may comprise contacting or incubating the cell or the organism comprising the cell with the nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof within a cell culture container. The cell culture container may comprise a 35mm culture dish, a 60mm culture dish, a 100mm culture dish, a 150mm culture dish, a 6- well culture plate, a 12- well culture plate, a 24-well culture plate, a 48-well culture plate, a 96-well culture plate, a T-25 flask, a T-75 flask, a T-175 flask, or a T-225 flask.

[0141] The cell culture container may have a cell growth surface area. The cell growth surface area may have an area of about 0.1 square centimeter (cmA2), 0.2 cmA2, 0.3 cmA2, 0.4 cmA2, 0.5 cmA2, 0.6 cmA2, 0.7 cmA2, 0.8 cmA2, 0.9 cmA2, 1 cmA2, 1.1 cmA2, 1.2 cmA2, 1.3 cmA2, 1.4 cmA2, 1.5 cmA2, 1.6 cmA2, 1.7 cmA2, 1.8 cmA2, 1.9 cmA2, 2 cmA2, 2.1 cmA2, 2.2 cmA2, 2.3 cmA2, 2.4 cmA2,2.5 cmA2, 2.6 cmA2, 2.7 cmA2, 2.8 cmA2, 2.9 cmA2, 3 cmA2, 3.1 cmA2, 3.2 cmA2, 3.3 cmA2, 3.4 cmA2, 3.5 cmA2, 3.6 cmA2, 3.7 cmA2, 3.8 cmA2, 3.9 cmA2, 4 cmA2, 4.1 cmA2, 4.2 cmA2, 4.3 cmA2,4.4 cmA2, 4.5 cmA2, 4.6 cmA2, 4.7 cmA2, 4.8 cmA2, 4.9 cmA2, 5 cmA2, 6 cmA2, 7 cmA2, 8 cmA2, 9cmA2, 10 cmA2, 20 cmA2, 30 cmA2, 40 cmA2, 50 cmA2, 60 cmA2, 70 cmA2 , 80 cmA2 , 90 cmA2 , 100 cmA2, 110 cmA2, 120 cmA2, 130 cmA2, 140 cmA2 , 150 cmA2 , 200 cmA2, 500 cmA2 , 1000 cmA2 or more.

[0142] The methods provided herein may comprise contacting a cell population with the nucleic acid molecule, saccharide, lipid, or transfection complex as described herein, or a combination thereof. The cell population may comprise at least about 1x10A3 cells, 2x10A3 cells, 3x10A3 cells, 4x10A3 cells, 5xl0A3 cells, 6xlOA3 cells, 7xlOA3 cells, 8xl0A3 cells, 9xlOA3 cells, lxlOA4 cells,2x10A4 cells, 3x10A4 cells, 4x10A4 cells, 5x10A4 cells, 6x10A4 cells, 7x10A4 cells, 8x10A4 cells,9x10A4 cells, 1x10A5 cells, 2x10A5 cells, 3x10A5 cells, 4x10A5 cells, 5x10A5 cells, 6x10A5 cells,7x10A5 cells, 8x10A5 cells, 9x10A5 cells, 1x10A6 cells, 2x10A6 cells, 3x10A6 cells, 4x10A6 cells,5x10A6 cells, 6x10A6 cells, 7x10A6 cells, 8x10A6 cells, 9x10A6 cells, 1x10A7 cells, 2x10A7 cells,3x10A7 cells, 4x10A7 cells, 5x10A7 cells, 6x10A7 cells, 7x10A7 cells, 8x10A7 cells, 9x10A7 cells,1x10A8 cells, 2xlOA8 cells, 3xl0A8 cells, 4xlOA8 cells, 5xl0A8 cells, 6xlOA8 cells, 7xlOA8 cells,8x10A8 cells, 9xlOA8 or more cells. The cell population may comprise at most about lxl0A3 cells, 2x10A3 cells, 3x10A3 cells, 4x10A3 cells, 5x10A3 cells, 6x10A3 cells, 7x10A3 cells, 8x10A3 cells,9x10A3 cells, 1x10A4 cells, 2x10A4 cells, 3x10A4 cells, 4x10A4 cells, 5x10A4 cells, 6x10A4 cells,7x10A4 cells, 8x10A4 cells, 9x10A4 cells, 1x10A5 cells, 2x10A5 cells, 3x10A5 cells, 4x10A5 cells,5x10A5 cells, 6x10A5 cells, 7x10A5 cells, 8x10A5 cells, 9x10A5 cells, 1x10A6 cells, 2x10A6 cells,3x10A6 cells, 4x10A6 cells, 5x10A6 cells, 6x10A6 cells, 7x10A6 cells, 8x10A6 cells, 9x10A6 cells,1x10A7 cells, 2xlOA7 cells, 3xlOA7 cells, 4xlOA7 cells, 5xlOA7 cells, 6xlOA7 cells, 7xlOA7 cells,8x10A7 cells, 9x10A7 cells, 1x10A8 cells, 2x10A8 cells, 3x10A8 cells, 4x10A8 cells, 5x10A8 cells,6x10A8 cells, 7x10A8 cells, 8x10A8 cells, or 9x10A8 cells.

[0143] The sufficient conditions for the methods provide herein may comprise a desirable cell density. The desirable cell density may be at least about 1x10A2 cells per cmA2 cell growth area, 2.5xlOA2 cells per cmA2 cell growth area, 5xlOA2 cells per cmA2 cell growth area, lxl0A3 cells per cmA2 cell growth area, 2.5xlOA3 cells per cmA2 cell growth area, 5x10A3 cells per cmA2 cell growth area, lxlOA4 cells per cmA2 cell growth area, 2.5xlOA4 cells per cmA2 cell growth area, 5xlOA4 cells per cmA2 cell growth area, 1x10A5 cells per cmA2 cell growth area, 2.5xlOA5 cells per cmA2 cell growth area, 5x10A5 cells per cmA2 cell growth area, 1x10A6 cells per cmA2 cell growth area, 2.5xlOA6 cells per cmA2 cell growth area, 5xlOA6 cells per cmA2 cell growth area, lxlOA7 cells per cmA2 cell growth area, 2.5xlOA7 cells per cmA2 cell growth area, 5x10A7 cells per cmA2 cell growth area, lxl0A8 cells per cmA2 cell growth area, 2.5xlOA8 cells per cmA2 cell growth area, 5xl0A8 cellsper cmA2 cell growth area or more. The desirable cell density may be at most about 1x10A2 cells per cmA2 cell growth area, 2.5xlOA2 cells per cmA2 cell growth area, 5x10A2 cells per cmA2 cell growth area, lxlOA3 cells per cmA2 cell growth area, 2.5xlOA3 cells per cmA2 cell growth area, 5xlOA3 cells per cmA2 cell growth area, 1x10A4 cells per cmA2 cell growth area, 2.5xlOA4 cells per cmA2 cell growth area, 5x10A4 cells per cmA2 cell growth area, 1x10A5 cells per cmA2 cell growth area, 2.5xlOA5 cells per cmA2 cell growth area, 5xlOA5 cells per cmA2 cell growth area, lxlOA6 cells per cmA2 cell growth area, 2.5xlOA6 cells per cmA2 cell growth area, 5x10A6 cells per cmA2 cell growth area, 1x10A7 cells per cmA2 cell growth area, 2.5xlOA7 cells per cmA2 cell growth area, 5x10A7 cells per cmA2 cell growth area, 1x10A8 cells per cmA2 cell growth area, 2.5xlOA8 cells per cmA2 cell growth area, or 5xlOA8 cells per cmA2 cell growth area.

[0144] In some cases, a nucleic acid and a saccharide, when used for contacting or transfecting a cell or the organism comprising the cell, may have a mass ratio of at least about 10000:1, 5000: 1, 2000:1, 1000:1, 500:1, 200:1, 100:1, 50:1, 20:1, 10:1, 5:1, 2:1, 1:1, 1:2, 1:5, 1:10, 1:20, 1:50, 1:100, 1:200, 1:500, 1:1000, 1:2000, 1:5000, or 1:10000. In some cases, a nucleic acid and a saccharide, when used for contacting or transfecting a cell, may have a mass ratio of at most about 10000: 1, 5000:1, 2000:1, 1000:1, 500:1, 200:1, 100:1, 50:1, 20:1, 10:1, 5:1, 2:1, 1:1, 1:2, 1:5, 1:10, 1:20, 1:50, 1:100, 1:200, 1:500, 1:1000, 1:2000, 1:5000, or 1:10000. In some cases, a nucleic acid and a lipid, when used for contacting or transfecting a cell or the organism comprising the cell, may have a mass ratio of at least about 10000:1, 5000:1,2000:1, 1000:1, 500:1,200:1, 100:1, 50:1, 20:1, 10:1, 5:1, 2:1, 1:1, 1:2, 1:5, 1:10, 1:20, 1:50, 1:100, 1:200, 1:500, 1:1000, 1:2000, 1:5000, or 1:10000. In some cases, a nucleic acid and a lipid, when used for contacting or transfecting a cell, may have a mass ratio of at most about 10000:1, 5000:1,2000:1, 1000:1, 500:1,200:1, 100:1, 50:1,20:1, 10:1, 5:1, 2:1, 1:1, 1:2, 1:5, 1:10, 1:20, 1:50, 1:100, 1:200, 1:500, 1:1000, 1:2000, 1:5000, or 1:10000. In some cases, a nucleic acid, a saccharide, and a lipid, when used for contacting or transfecting a cell or the organism comprising the cell, may have a mass ratio of about 1:2:1, 1:2:, 2, 1:2:3, 1:2:4, 1:2:5, 1:2:10, 1:2:20, 1:4:4, 1:10:10, 1:10:100, 1:100:100, 1:300:300, 1:350:300, 1:400:300, 1:450:300, 1:500:300; 1:550:300, 1:600:300, 1:650:300, 1:700:300, 1:1000:300, 5:10:100, 5:100:100, 5:300:300, 5:350:300, 5:400:300, 5:450:300, 5:500:300; 5:550:300, 5:600:300, 5:650:300, 5:700:300, 5:1000:300, 10:10:100, 10:100:100, 10:300:300, 10:350:300, 10:400:300, 10:450:300, 10:500:300; 10:550:300, 10:600:300, 10:650:300, 10:700:300, 10:1000:300, 1:500:1, 1:500:10, 1:500:100, 1:500:150, 1:500:200, 1:500:250, 1:500:300, 1:500:350, 1:500:400, 1:500:450, or 1:500:500.

[0145] In some cases, a nucleic acid may be at least about 0.001 ng / 10000 cells, 0.002 ng / 10000 cells, 0.005 ng / 10000 cells, 0.01 ng / 10000 cells, 0.02 ng / 10000 cells, 0.05 ng / 10000 cells, 0.1 ng / 10000 cells, 0.2 ng / 10000 cells, 0.5 ng / 10000 cells, 1 ng / 10000 cells, 2 ng / 10000 cells, 5 ng / 10000 cells, 10 ng / 10000 cells, 20 ng / 10000 cells, 50 ng / 10000 cells, 100 ng / 10000 cells, 200 ng / 10000 cells, 500 ng / 10000 cells, 1000 ng / 10000 cells, 5000 ng / 10000 cells, 10000 ng / 10000 cells or more, when used for contacting or transfecting a cell. In some cases, a nucleic acid may be at most about 0.001 ng / 10000 cells, 0.002 ng / 10000 cells, 0.005 ng / 10000 cells, 0.01 ng / 10000 cells, 0.02 ng / 10000 cells, 0.05 ng / 10000 cells, 0.1 ng / 10000 cells, 0.2 ng / 10000 cells, 0.5 ng / 10000 cells, 1 ng / 10000 cells, 2 ng / 10000 cells, 5 ng / 10000 cells, 10 ng / 10000 cells, 20 ng / 10000 cells, 50 ng / 10000 cells, 100 ng / 10000 cells, 200 ng / 10000 cells, 500 ng / 10000 cells, 1000 ng / 10000 cells, 5000 ng / 10000 cells, or 10000 ng / 10000 cells, when used for contacting or transfecting a cell.

[0146] In some cases, a saccharide may be at least about 0.001 ng / 10000 cells, 0.002 ng / 10000 cells, 0.005 ng / 10000 cells, 0.01 ng / 10000 cells, 0.02 ng / 10000 cells, 0.05 ng / 10000 cells, 0.1 ng / 10000 cells, 0.2 ng / 10000 cells, 0.5 ng / 10000 cells, 1 ng / 10000 cells, 2 ng / 10000 cells, 5 ng / lOsOOO cells, 10 ng / 10000 cells, 20 ng / 10000 cells, 50 ng / 10000 cells, 100 ng / 10000 cells, 200 ng / 10000 cells, 500 ng / 10000 cells, 1000 ng / 10000 cells, 2000 ng / 10000 cells, 5000 ng / 10000 cells, 10000 ng / 10000 cells, 20 pg / 10000 cells, 50 pg / 10000 cells, 100 pg / 10000 cells, 200 pg / 10000 cells, 500 pg / 10000 cells, 1000 pg / 10000 cells, 2000 pg / 10000 cells, 5000 pg / 10000 cells, 10000 pg / 10000 cells or more, when used for contacting or transfecting a cell. In some cases, a saccharide may be at most about 0.001 ng / 10000 cells, 0.002 ng / 10000 cells, 0.005 ng / 10000 cells, 0.01 ng / 10000 cells, 0.02 ng / 10000 cells, 0.05 ng / 10000 cells, 0.1 ng / 10000 cells, 0.2 ng / 10000 cells, 0.5 ng / 10000 cells, 1 ng / 10000 cells, 2 ng / 10000 cells, 5 ng / 10000 cells, 10 ng / 10000 cells, 20 ng / 10000 cells, 50 ng / 10000 cells, 100 ng / 10000 cells, 200 ng / 10000 cells, 500 ng / 10000 cells, 1000 ng / 10000 cells, 2000 ng / 10000 cells, 5000 ng / 10000 cells, 10000 ng / 10000 cells, 20 pg / 10000 cells, 50 pg / 10000 cells, 100 pg / 10000 cells, 200 pg / 10000 cells, 500 pg / 10000 cells, 1000 pg / 10000 cells, 2000 pg / 10000 cells, 5000 pg / 10000 cells, or 10000 pg / 10000 cells, when used for contacting or transfecting a cell.

[0147] In some cases, a lipid may be at least about 0.001 ng / 10000 cells, 0.002 ng / 10000 cells, 0.005 ng / 10000 cells, 0.01 ng / 10000 cells, 0.02 ng / 10000 cells, 0.05 ng / 10000 cells, 0.1 ng / 10000 cells, 0.2 ng / 10000 cells, 0.5 ng / 10000 cells, 1 ng / 10000 cells, 2 ng / 10000 cells, 5 ng / 10000 cells, 10 ng / 10000 cells, 20 ng / 10000 cells, 50 ng / 10000 cells, 100 ng / 10000 cells, 200 ng / 10000 cells, 500 ng / 10000 cells, 1000 ng / 10000 cells, 2000 ng / 10000 cells, 5000 ng / 10000 cells, 10000 ng / 10000cells, 20 pg / 10000 cells, 50 pg / 10000 cells, 100 pg / 10000 cells, 200 pg / 10000 cells, 500 pg / 10000 cells, 1000 pg / 10000 cells, 2000 pg / 10000 cells, 5000 pg / 10000 cells, 10000 pg / 10000 cells or more, when used for contacting or transfecting a cell. In some cases, a lipid may be at most about 0.001 ng / 10000 cells, 0.002 ng / 10000 cells, 0.005 ng / 10000 cells, 0.01 ng / 10000 cells, 0.02 ng / 10000 cells, 0.05 ng / 10000 cells, 0.1 ng / 10000 cells, 0.2 ng / 10000 cells, 0.5 ng / 10000 cells, 1 ng / 10000 cells, 2 ng / 10000 cells, 5 ng / 10000 cells, 10 ng / 10000 cells, 20 ng / 10000 cells, 50 ng / 10000 cells, 100 ng / 10000 cells, 200 ng / 10000 cells, 500 ng / 10000 cells, 1000 ng / 10000 cells, 2000 ng / 10000 cells, 5000 ng / 10000 cells, 10000 ng / 10000 cells, 20 pg / 10000 cells, 50 pg / 10000 cells, 100 pg / 10000 cells, 200 pg / 10000 cells, 500 pg / 10000 cells, 1000 pg / 10000 cells, 2000 pg / 10000 cells, 5000 pg / 10000 cells, or 10000 pg / 10000 cells, when used for contacting or transfecting a cell.

[0148] In some cases, a nucleic acid may be at least about 0.001 mg / 1 kg of an organism mass, 0.002 mg / 1 kg of an organism mass, 0.005 mg / 1 kg of an organism mass, 0.01 mg / 1 kg of an organism mass, 0.02 mg / 1 kg of an organism mass, 0.05 mg / 1 kg of an organism mass, 0.1 mg / 1 kg of an organism mass, 0.2 mg / 1 kg of an organism mass, 0.5 mg / 1 kg of an organism mass, 1 mg / 1 kg of an organism mass, 2 mg / 1 kg of an organism mass, 5 mg / 1 kg of an organism mass, 10 mg / 1 kg of an organism mass, 20 mg / 1 kg of an organism mass, 50 mg / 1 kg of an organism mass, 100 mg / 1 kg of an organism mass, 200 mg / 1 kg of an organism mass, 500 mg / 1 kg of an organism mass, 1000 mg / 1 kg of an organism mass, 5000 mg / 1 kg of an organism mass, 10000 mg / 1 kg of an organism mass or more, when used for contacting or transfecting the organism. In some cases, a nucleic acid may be at most about 0.001 mg / 1 kg of an organism mass, 0.002 mg / 1 kg of an organism mass, 0.005 mg / 1 kg of an organism mass, 0.01 mg / 1 kg of an organism mass, 0.02 mg / 1 kg of an organism mass, 0.05 mg / 1 kg of an organism mass, 0.1 mg / 1 kg of an organism mass, 0.2 mg / 1 kg of an organism mass, 0.5 mg / 1 kg of an organism mass, 1 mg / 1 kg of an organism mass, 2 mg / 1 kg of an organism mass, 5 mg / 1 kg of an organism mass, 10 mg / 1 kg of an organism mass, 20 mg / 1 kg of an organism mass, 50 mg / 1 kg of an organism mass, 100 mg / 1 kg of an organism mass, 200 mg / 1 kg of an organism mass, 500 mg / 1 kg of an organism mass, 1000 mg / 1 kg of an organism mass, 5000 mg / 1 kg of an organism mass, or 10000 mg / 1 kg of an organism mass, when used for contacting or transfecting the organism.

[0149] In some cases, a saccharide may be at least about 0.001 mg / 1 kg of an organism mass, 0.002 mg / 1 kg of an organism mass, 0.005 mg / 1 kg of an organism mass, 0.01 mg / 1 kg of an organism mass, 0.02 mg / 1 kg of an organism mass, 0.05 mg / 1 kg of an organism mass, 0.1 mg / 1 kg of anorganism mass, 0.2 mg / 1 kg of an organism mass, 0.5 mg / 1 kg of an organism mass, 1 mg / 1 kg of an organism mass, 2 mg / 1 kg of an organism mass, 5 mg / 1 kg of an organism mass, 10 mg / 1 kg of an organism mass, 20 mg / 1 kg of an organism mass, 50 mg / 1 kg of an organism mass, 100 mg / 1 kg of an organism mass, 200 mg / 1 kg of an organism mass, 500 mg / 1 kg of an organism mass, 1000 mg / 1 kg of an organism mass, 2000 mg / 1 kg of an organism mass, 5000 mg / 1 kg of an organism mass, 10000 mg / 1 kg of an organism mass or more, when used for contacting or transfecting the organism. In some cases, a saccharide may be at most about 0.001 mg / 1 kg of an organism mass, 0.002 mg / 1 kg of an organism mass, 0.005 mg / 1 kg of an organism mass, 0.01 mg / 1 kg of an organism mass, 0.02 mg / 1 kg of an organism mass, 0.05 mg / 1 kg of an organism mass, 0.1 mg / 1 kg of an organism mass, 0.2 mg / 1 kg of an organism mass, 0.5 mg / 1 kg of an organism mass, 1 mg / 1 kg of an organism mass, 2 mg / 1 kg of an organism mass, 5 mg / 1 kg of an organism mass, 10 mg / 1 kg of an organism mass, 20 mg / 1 kg of an organism mass, 50 mg / 1 kg of an organism mass, 100 mg / 1 kg of an organism mass, 200 mg / 1 kg of an organism mass, 500 mg / 1 kg of an organism mass, 1000 mg / 1 kg of an organism mass, 2000 mg / 1 kg of an organism mass, 5000 mg / 1 kg of an organism mass, or 10000 mg / 1 kg of an organism mass, when used for contacting or transfecting the organism.

[0150] In some cases, a lipid may be at least about 0.001 mg / 1 kg of an organism mass, 0.002 mg / 1 kg of an organism mass, 0.005 mg / 1 kg of an organism mass, 0.01 mg / 1 kg of an organism mass, 0.02 mg / 1 kg of an organism mass, 0.05 mg / 1 kg of an organism mass, 0.1 mg / 1 kg of an organism mass, 0.2 mg / 1 kg of an organism mass, 0.5 mg / 1 kg of an organism mass, 1 mg / 1 kg of an organism mass, 2 mg / 1 kg of an organism mass, 5 mg / 1 kg of an organism mass, 10 mg / 1 kg of an organism mass, 20 mg / 1 kg of an organism mass, 50 mg / 1 kg of an organism mass, 100 mg / 1 kg of an organism mass, 200 mg / 1 kg of an organism mass, 500 mg / 1 kg of an organism mass, 1000 mg / 1 kg of an organism mass, 2000 mg / 1 kg of an organism mass, 5000 mg / 1 kg of an organism mass, 10000 mg / 1 kg of an organism mass or more, when used for contacting or transfecting the organism. In some cases, a lipid may be at most about 0.001 mg / 1 kg of an organism mass, 0.002 mg / 1 kg of an organism mass, 0.005 mg / 1 kg of an organism mass, 0.01 mg / 1 kg of an organism mass, 0.02 mg / 1 kg of an organism mass, 0.05 mg / 1 kg of an organism mass, 0.1 mg / 1 kg of an organism mass, 0.2 mg / 1 kg of an organism mass, 0.5 mg / 1 kg of an organism mass, 1 mg / 1 kg of an organism mass, 2 mg / 1 kg of an organism mass, 5 mg / 1 kg of an organism mass, 10 mg / 1 kg of an organism mass, 20 mg / 1 kg of an organism mass, 50 mg / 1 kg of an organism mass, 100 mg / 1 kg of an organism mass, 200 mg / 1 kg of an organism mass, 500 mg / 1 kg of an organism mass, 1000 mg / 1 kg of an organismmass, 2000 mg / 1 kg of an organism mass, 5000 mg / 1 kg of an organism mass, or 10000 mg / 1 kg of an organism mass, when used for contacting or transfecting the organism.

[0151] Any compositions herein may be biodegradable. Any compositions herein may be biocompatible with a cell that it contacts. A composition, saccharide, lipid, nucleic acid molecule, or a combination thereof, described herein, may be biodegradable or biocompatible with a cell.Biocompatible compositions, saccharides, lipids, nucleic acid molecules, polymeric materials, or a combination thereof may be beneficial. Biocompatible materials may have immunostimulatory activity, anticoagulant activity, wound-healing properties, anti-microbial properties, or a combination thereof. In some cases, biocompatible composition, saccharide, lipid, nucleic acid molecule, or a combination thereof may be non-toxic, non-hemolytic, non-immunogenic, slowly biodegradable, and / or nuclease resistant. In some cases, a saccharide or lipid may increase transcellular and paracellular transport. Thus, using any biocompatible materials described herein may increase the number of cells that has taken up any nucleic acids described herein, relative to using non-biocompatible materials.

[0152] Biodegradable compositions, saccharides, lipids, nucleic acid molecules, or a combination thereof may be beneficial. For example, even if a biodegradable material may have undesirable effects to a cell (cytotoxicity, carcinogenic activities, immunogenic activities, or a combination thereof), since the material is biodegradable, the undesirable effects generated by the material is minimized, relative to the non-biodegradable counterparts. Biodegradability may be mediated by various enzymes of the cell.Protein expression or differentiation

[0153] In some instances, the nucleic acid molecule may be configured to facilitate protein expression of a cell. A nucleic acid molecule may facilitate alteration of protein expression of a cell. A nucleic acid molecule, or a combination thereof may alter the protein expression of a cell. In some instances, a nucleic acid molecule may be configured to alter the protein expression of a cell. In some instances, a nucleic acid molecule may be configured to promote differentiation of a cell. In some instances, a nucleic acid molecule may be configured to facilitate differentiation of a cell.

[0154] In some instances, an alteration in protein expression within a cell may initiate or facilitate the process of a conversion of a cell type into another cell type. Initiation or alteration of protein expression within a cell may alter the shared structural or functional characteristics of a cell, leading to conversion of a cell type into another cell type. In some case, a change in protein expressionwithin a cell may facilitate or promote differentiation of the cell. In some instances, protein expression may result in a differentiation which may comprise transdifferentiation. In some cases, a transdifferentiation may comprise a differentiation of a cell type into another cell type without reaching a pluripotent cell state during the differentiation process. In some instances, a differentiation may comprise transdifferentiation of somatic cells or directed differentiation of naive cells.

[0155] In some instances, a nucleic acid molecule may alter the protein expression of a cell and may promote or facilitate differentiation of a cell. In some instances, a saccharide may promote or facilitate differentiation of a cell. In some instances, a cationic saccharide may promote or facilitate a differentiation of a cell. In some instances, a lipid may promote or facilitate a differentiation of a cell. In some instances, a nanoparticle may promote or facilitate a differentiation of a cell. In some cases, a saccharide / lipid may facilitate or promote the differentiation of a cell by enhancing the uptake of a nucleic acid molecule by the cell. The terms “promote,” “promotion,” “facilitate,” “facilitation,” or grammatically equivalent, when used herein referring to a cellular process (e.g., protein expression or differentiation), refers to lowering the threshold or increasing the occurrence of the cellular process. In some cases, if a molecular entity or a group of molecular entities facilitate(s) a cellular process of a cell, the cellular process has an increased probability to occur in the cell comprising the molecular entity or the group of molecular entities, relative to the cell not comprising the molecular entity or the group of molecular entities. For example, if a molecular entity or a group of molecular entities facilitate(s) a cellular process of a cell, the cellular process may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more likely to occur in the cell comprising the molecular entity or the group of molecular entities, relative to the cell not comprising the molecular entity or the group of molecular entities.

[0156] In some cases, if a molecular entity or a group of molecular entities facilitate(s) a cellular process of a group of cells, the cellular process has an increased probability to occur or an increased number of cells with the occurrence of the cellular process in the group of cells comprising the molecular entity or the group of molecular entities, relative to the group of cells not comprising the molecular entity or the group of molecular entities. For example, if a molecular entity or a group of molecular entities facilitate(s) a cellular process of a group cells, the cellular process may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times,2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more likely to occur in the group of cells comprising the molecular entity or the group of molecular entities, relative to the group of cells not comprising the molecular entity or the group of molecular entities. In other cases, if a molecular entity or a group of molecular entities facilitate(s) a cellular process of a group cells, the cellular process may occur in at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more cells in the group of cells comprising the molecular entity or the group of molecular entities, relative to the group of cells not comprising the molecular entity or the group of molecular entities.

[0157] The protein expression within a cell facilitated or promoted by a nucleic acid molecule may comprise a polypeptide or protein encoded by the nucleic acid molecule. In some cases, the nucleic acid molecule taken up by the cell may alter the expression of other proteins. For example, the nucleic acid molecule that contacts the cell may be translated by the cell into a polypeptide or protein. The polypeptide or protein may interact with other cellular components and facilitate or promote an alteration of expressions of other proteins, transcripts, nucleic acids, and / or other metabolites. In some cases, the nucleic acid molecule that contacts the cell may not be translated into a protein or polypeptide by the cell. The nucleic acid molecule, without itself being translated into a protein or polypeptide, may facilitate or promote expression of other cellular proteins. The nucleic acid molecule may interact with other cellular components and facilitate or promote an alteration of expressions of other proteins, transcripts, nucleic acids, and / or other metabolites.

[0158] In some cases, the facilitation or promotion of an alteration of expression of a protein within a cell by a nucleic acid molecule described herein may occur at the transcriptional level (e.g., an alteration of transcription of the gene encoding the protein), post- transcriptional level (e.g., an alteration of transcript stability, transport, RNA modification, or a combination thereof of the transcript encoding the protein), translational level (e.g., an alteration of translation of the transcript encoding the protein), post-translational level (e.g., an alteration of post-translational modifications of the proteins), or a combination thereof.

[0159] In some instances, promoting or facilitating an expression of a protein in a cell may comprise an increase in the expression level of the protein. The terms “cell(s)”, “cell population(s),” or “population(s) of cells,” or any grammatically equivalent terms may be used interchangeably. Insome cases, the increase in the expression level of the protein may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more than the expression level of the protein without the increase. In some instances, promoting or facilitating an expression of a protein in a cell may comprise an increase in the number of cells in population of cells that may express the proteins. The increase in the number of cells that expressed the protein, with the aid of the compositions or methods of the present disclosure, may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times greater than the number of cells that expressed the protein in the absence of the compositions or methods of the present disclosure. In some instances, promoting or facilitating an alteration of an expression of a protein in a cell may comprise a decrease in the expression level of the protein. In some cases, the decrease in the expression level of the protein may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 95 %, 99 % or lower than the expression level of the protein without the decrease. In some instances, promoting or facilitating an expression of a protein in a cell may comprise a decrease in the number of cells in population of cells that may express the proteins. The decrease in the number of cells that expressed the protein, with the aid of the compositions or methods of the present disclosure, may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 95 %, 99 % or fewer than the number of cells that expressed the protein in the absence of the compositions or methods of the present disclosure.

[0160] Cell that has contacted a composition provided herein comprising a nucleic acid molecule, or a progeny derive from the cell, may express at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more the nucleic acid molecule, the transcript or derivative of the nucleic molecule, the protein or polypeptide encoded by the nucleic acid molecule, or a combination thereof, relative to the cell that has not contacted the composition or a progeny thereof. A population of cells that has contacted a composition provided herein comprising a nucleic acid molecule, or a progeny derive from the population of cells, may have at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more cells that express the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the population of cells that has not contacted the composition or a progeny thereof.

[0161] Cell that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the cell, may express at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more of a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the cell that has not contacted the composition or a progeny thereof. A population of cells that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the population of cells, may have at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more cells that express a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the population of cells that has not contacted the composition or a progeny thereof. Cell that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the cell, may express at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 95 %, 99 % or less of a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the cell that has not contacted the composition or a progeny thereof. A population of cells that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the population of cells, may have at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 95 %, 99 % or less cells that express a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or acombination thereof, relative to the population of cells that has not contacted the composition or a progeny thereof.

[0162] Cell that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the cell, may express at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times,6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more of a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the cell that has only contacted the first nucleic acid molecule without the other components of the compositions described herein or a progeny thereof. A population of cells that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the population of cells, may have at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times,7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times or more cells that express a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the population of cells that has only contacted the first nucleic acid molecule without the other components of the compositions described herein or a progeny thereof.

[0163] Cell that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the cell, may express at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 95 %, 99 % or less of a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combination thereof, relative to the cell that has only contacted the first nucleic acid molecule without the other components of the compositions described herein or a progeny thereof. A population of cells that has contacted a composition provided herein comprising a first nucleic acid molecule, or a progeny derive from the population of cells, may have at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 95 %, 99 % or less cells that express a second nucleic acid molecule different from the first nucleic acid molecule, the transcript or derivative of the second nucleic molecule, the protein or polypeptide encoded by the second nucleic acid molecule, or a combinationthereof, relative to the population of cells that has not only contacted the first nucleic acid molecule without the other components of the compositions described herein or a progeny thereof.

[0164] The alteration of protein expression may be measured at least about 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours,25 hours, 26 hours, 27 hours, 28 hours, 29 hours, 30 hours, 31 hours, 32 hours, 33 hours, 34 hours,35 hours, 36 hours, 37 hours, 38 hours, 39 hours, 40 hours, 41 hours, 42 hours, 43 hours, 44 hours,45 hours, 46 hours, 47 hours, 48 hours, 3 days, 4 days, 5 days, 6 days, 7 days, 1 week, 2 weeks, 3 weeks, 4 weeks or more, subsequent to a cell is contacted with a nucleic acid, a saccharide, or a combination thereof, or a control thereof. The increase or decrease of protein expression may be measured at most about 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours,20 hours, 21 hours, 22 hours, 23 hours, 24 hours, 25 hours, 26 hours, 27 hours, 28 hours, 29 hours,30 hours, 31 hours, 32 hours, 33 hours, 34 hours, 35 hours, 36 hours, 37 hours, 38 hours, 39 hours,40 hours, 41 hours, 42 hours, 43 hours, 44 hours, 45 hours, 46 hours, 47 hours, 48 hours, 3 days, 4 days, 5 days, 6 days, 7 days, 1 week, 2 weeks, 3 weeks, or 4 weeks, subsequent to a cell is contacted with a nucleic acid, a saccharide, or a combination thereof, or a control thereof.

[0165] In some cases, the alteration of protein expression of a cell may comprise a difference of protein expressed (or not express) or a difference of proteins expressed at a pre-determined level. Alteration of protein expression of a cell may comprise at least about 1, 5, 10, 50, 100, 500, 1000, 5000, 10000 or more proteins that are expressed or not expressed. Alteration of protein expression of a cell may comprise at least about 1, 5, 10, 50, 100, 500, 1000, 5000, 10000 or more proteins that are expressed or not expressed at a pre-determined level. The pre-determined level may be a level relative to a pre- determined control proteins. The control proteins may be the proteins expressed at a constant level by the cells in various cellular states or culture conditions. The control proteins may be the proteins expressed by house-keeping genes. The levels of a control protein may be within at most about 1 %, 2 %, 3 %, 4 %, 5 %, 7 %, 8 %, 9 %, 10 %, 15 %, 20 %, 25 %, or 30 % when measured in the cells in two distinct cellular states or culture conditions. The expression level of a protein may be measured by quantitative real-time polymerase chain reaction, nucleic acid sequencing, fluorescent hybridization, or a combination thereof of the gene product encoding the protein (e.g., mRNA). The expression level of a protein may be measured by western blot, flowcytometry, mass-spectrometry, immunofluorescence, fluorescent tag / label, or a combination thereof of the protein.

[0166] The alteration of the expression of a protein described herein may be measured at the protein level. For example, western blot, eastern blot, immunofluorescence, antibody staining, mass spectrometry, gel electrophoresis, uses of a reporter that reflects the expression of the proteins, or any combination thereof may be used to measure the expression of the protein. The alteration of the expression of a protein described herein may be measured at the cell population level. For example, flow cytometry, counting of cells that express a protein (reporter or immunofluorescence), or a combination thereof may be used to measure the expression of the protein. The alteration of the expression of a protein described herein may be measured at the transcript level. For example, quantitative real-time PCR (QRT-PCR), reverse-transcription PCR, northern blot, RNA-sequencing, fluorescence hybridization, or a combination thereof may be used to measure the expression of the protein.

[0167] In some instances, promoting or facilitating differentiation of a cell may comprise an increase of the number of cells in population of cells that may undergo differentiation. The increase of the number of cells that undergoes differentiation, with the aid of the compositions or methods of the present disclosure, may be at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times greater than the number of cells that undergoes differentiation in the absence of the compositions or methods of the present disclosure.

[0168] In some instances, a cell that has taken up the nucleic acid molecule may have a change in intracellular processes. In some cases, a change in intracellular process may comprise changes in expression (increase or decrease) of one or more proteins in the cell (e.g., with the aid of or due to the presence of the nucleic acid molecules ). In some instances, two or more nucleic acid molecules may be used collectively to promote protein expression or facilitate differentiation of a cell. Each of the two or more nucleic acid molecules may be configured to affect an expression (e.g., enhancing or suppressing expression) of a given protein or gene in the cell. The two or more nucleic acid molecules may affect expression of the same protein or gene or different proteins or genes. The two or more nucleic acid molecules may collectively affect expression of the same protein, gene or different proteins or genes. When at least two nucleic molecules collectively facilitate protein expression or promote the differentiation of a cell, the overlap of the genes (or proteins or nucleicacids) with the expression affected by each nucleic acid molecule may comprise about 0 %, 1 %, 5%, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 100 % of the genes with altered gene expression.Cells / organisms being transfected

[0169] In some cases, the organism being contacted with the composition described herein may comprise an animal. The animal may comprise a human. The animal may comprise a non-human animal. The non-human animal may comprise a rodent, fish, dog, cat, cow, pig, or a combination thereof. In some cases, the organism being contacted with the composition described herein may comprise a plant. In some cases, the animal or the cell being contacted with the composition as described herein may have a disease or condition. The disease or condition may comprise cancer / neoplastic disorder, neurological disorder, or a combination thereof. In some cases, the animal or the cell being contacted with the composition as described herein may not have a disease or condition.

[0170] When contacting the organism, the composition may be taken by the cells of a liver, lung, spleen, skin, brain, heart, kidney, pancreas, thyroidjoints, or a combination thereof. When contacting the organism, the composition may be preferably taken up by (or localized to) the cells of a first tissue over the cells of a different tissue. For example, the composition may be taken up by (or localized to) at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 50, 100, 200, 300, 400, 500, 600, 700, 800, 900, lxl0A3, lxlOA4, lxl0A5, lxlOA6, 1 xlOA7, lxl0A8 more cells in the first tissue than the cells of a different tissue. In some cases, the composition may be taken up by (or localized to) at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 50, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1x10A3, 1x10A4, 1x10A5, 1x10A6, 1 xlOA7, 1x10A8 more cells in the first tissue than the cells of a different tissue. In some cases, subsequent to contacting the composition, a first tissue may have at least about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times of the composition, relative to a different tissue. In some cases, subsequent to contacting the composition, a first tissue may have at most about 1 %, 5 %, 10 %, 20 %, 30 %, 40 %, 50 %, 60 %, 70 %, 80 %, 90 %, 1 time, 1.5 times, 2 times, 2.5 times, 3 times, 4 times, 5 times, 6 times, 7 times, 8 times, 9 times, 10 times, 15 times, 20 times, 25 times, 30 times, 35 times, 40 times, 45 times, 50 times, 100 times, 1000 times of the composition, relative to adifferent tissue. The percentage or amount of the composition taken up or localized to a tissue may be measured using the methods as described herein, such as those described in Example 4.

[0171] In some cases, a cell type may comprise a cell that expresses a specific profile or group of genes, nucleic acid molecules, and / or proteins. In some cases, cells of a cell type may have a common expression profile or group of genes, nucleic acid molecules, and / or proteins. In some cases, cells of a cell type may express at least about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 50, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 5000, 10000 or more identical genes, nucleic acid molecules, and / or proteins. In some cases, cells of a cell type may express at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 50, 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 2000, 5000, 10000, 20000, or 30000 identical genes, nucleic acid molecules, and / or proteins. Cells of a cell type may have shared structural or functional characteristics.

[0172] A cell may comprise cells or a cell population. A cell may comprise progenies of a cell or cell population. Cells may comprise progenies of a single cell.

[0173] In some instances, a cell may comprise a fat cell, a blood vessel cell, a cardiac cell, a chondrocyte, an endothelia cell, an epithelial cell, a hematopoietic cell, a hepatocyte, a muscle cell, a neuron, or an osteogenic cell. In some cases, a cell may comprise a fat cell. In some cases, a cell may comprise a blood vessel cell. In some cases, a cell may comprise a cardiac ell. In some cases, a cell may comprise a chondrocyte. In some cases, a cell may comprise an endothelia cell. In some cases, a cell may comprise an epithelial cell. In some cases, a cell may comprise a hematopoietic cell. In some cases, a cell may comprise a hepatocyte. In some cases, a cell may comprise a muscle cell. In some cases, a cell may comprise a neuron. In some cases, a cell may comprise an osteogenic cell. In some instances, a cell may comprise at least 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of a fat cell, a blood vessel cell, a cardiac ell, a chondrocyte, an endothelia cell, an epithelial cell, a hematopoietic cell, a hepatocyte, a muscle cell, a neuron, and an osteogenic cell. In some cases, a cementogenic cell comprises a cell that can form or be a part of a tooth. In some cases, an endothelia cell comprises a cell that can form or be a part of the endothelium. In some cases, an epithelial cell comprises a cell that can form or be a part of the epithelium. In some cases, a fat cell may comprise a cell that can form or be a part of the adipose tissues. In some cases, a hematopoietic cell may comprise a cell that can form or be a part of the blood or hematopoietic tissues. In some cases, a hepatocyte may comprise a cell that can form or be a part of the hepatic or liver tissue. In some cases, a muscle cell may comprise a cell that can form or be a part of the muscle tissues. In some cases, a neuron maycomprise a cell that can form or be a part of the nervous systems. In some cases, a renal cell may comprise a cell that can form or be a part of the kidney tissues. In some cases, a retinal cell may comprise a cell that can form or be a part of the ocular or eye tissues. In some cases, an osteogenic cell may comprise a cell that can form or be a part of the bone tissues. In some cases, a blood vessel cell or an angiogenic cell may comprise a cell that can form or be a part of the blood vessel or vascular tissue. In some cases, a cardiac cell may comprise a cell that can form or be a part of the heart or cardiac tissue. In some cases, a chondrocyte cell may comprise a cell that can form or be a part of the cartilage tissue.

[0174] In some cases, a fat cell may be an adipocyte. In some cases, a fat cell may contain various sizes of fat droplets or granules. In some cases, a fat cell may comprise a white adipose cell or a brown adipose cell. In some cases, a white adipose cell may contain large fat droplets or granules and a small amount of cytoplasm. In some cases, a brown adipose cell may contain a large amount of cytoplasm and numerous mitochondria. In some cases, an adipocyte may be a cell primarily composed of adipose tissue, specialized in synthesizing and storing energy as fat. Adipocytes may be derived from induced pluripotent stem cells and / or mesenchymal stem cells through adipogenesis. Adipocytes may be white adipocytes, which store energy as a single large lipid droplet and have important endocrine functions, and brown adipocytes which store energy in multiple small lipid droplets but specifically for use as fuel to generate body heat.

[0175] In some cases, a muscle cell may develop sarcoplasm, sarcoplasmic reticulum, sarcosome, or sarcolemma that are specialized for muscle contraction and energy metabolism. In some cases, a muscle cell may contain myofibrils and myoglobins. In some cases, a muscle cell may contain a high amount of glycogen. In some cases, a muscle cell may also comprise a myocyte. In some cases, a muscle cell may develop from a myoblast. In some cases, a muscle cell may be a cardiac muscle cell, a smooth muscle cell, or a skeletal muscle cell. In some instances, a muscle cell may comprise a myofiber, a myotube, a myocyte, a myoblast, a spheroid, or a muscle cell progenitor.

[0176] In some instances, a cell may comprise an animal cell. An animal cell may comprise a cell isolated or derived from an organism from the kingdom Animalia. An animal cell may be isolated from an animal. A cell may also be an animal cell if the closest counterpart of its genome is from an animal or an animal cell.

[0177] In some instances, an animal cell may comprise a mammalian cell, a bird cell, or a fish cell, a mollusk cell, or an amphibian cell. In some instances, an animal cell may comprise a mollusk cell. Insome instances, an animal cell may comprise an amphibian cell. In some instances, an animal cell may comprise a mollusk cell. In some instances, a cell may comprise a plant cell.

[0178] In some instances, an animal cell may comprise a mammalian cell. In some instances, an animal cell may comprise a bird cell. In some instances, an animal cell may comprise a fish cell.

[0179] In some instances, a mammalian cell may comprise a porcine cell, a bovine cell, a bubaline cell, an ovine cell, a caprine cell, a cervine cell, a bisontine cell, a cameline cell, an elaphine cell, or a lapine cell. In some cases, a cell may comprise a porcine cell. In some cases, a cell may comprise a bovine cell. In some cases, a cell may comprise a bubaline cell. In some cases, a cell may comprise an ovine cell. In some cases, a cell may comprise a caprine cell. In some cases, a cell may comprise a cervine cell. In some cases, a cell may comprise a bisontine cell. In some cases, a cell may comprise a cameline cell. In some cases, a cell may comprise an elaphine cell. In some cases, a cell may comprise a lapine cell.

[0180] In some instances, a bird cell may comprise an anatine cell, a galline cell, an anserine cell, a meleagrine cell, a struthionine cell, or a phasianine cell. In some cases, a cell may comprise an anatine cell. In some cases, a cell may comprise a galline cell. In some cases, a cell may comprise an anserine cell. In some cases, a cell may comprise a meleagrine cell. In some cases, a cell may comprise a struthionine cell. In some cases, a cell may comprise a phasianine cell.

[0181] In some instance, a differentiated cell may undergo self-renewal. In other cases, a differentiated cell may not undergo self-renewal. In some cases, a terminally differentiated cell may not undergo self-renewal. In some cases, a differentiated cell may not be at a pluripotent cell state.

[0182] In some cases, a cell that can undergo differentiation may comprise a stem cell. In other cases, a stem cell may be at a totipotent cell state. In some cases, a stem cell maybe at a pluripotent cell state. In other cases, a stem cell may be at a multipotent cell state. In other cases, a stem cell may be at an omnipotent cell state. In other cases, a stem cell may be at a unipotent cell state. In some cases, a stem cell may comprise an iPSC. In some cases, a stem cell may comprise an embryonic stem cell, an immortalized stem cell, a mesenchymal stem cell, and / or a muscle progenitor cell. In some cases, a stem cell may comprise an embryonic stem cell, an immortalized stem cell, a mesenchymal stem cell, an iPSC, and / or a muscle progenitor cell. In some cases, a stem cell may comprise an embryonic stem cell. In some cases, a stem cell may comprise an immortalized stem cell. In some cases, a stem cell may comprise a mesenchymal stem cell. In some cases, a stem cell may comprise a muscle progenitor cell.

[0183] A stem cell may comprise a reprogrammed cell. Cellular reprogramming may be a process that reverses the developmental potential of a cell or population of cells (e.g., a somatic cell). Reprogramming may be a process of driving a cell to a state with higher developmental potential, such as driving a cell backwards to a less differentiated state. The cell to be reprogrammed can be either partially or terminally differentiated prior to reprogramming. Reprogramming may infer a complete or partial reversion of the differentiation state, such as an increase in the developmental potential of a cell, to that of a cell having a pluripotent state, driving a somatic cell to a pluripotent state, such that the cell has the developmental potential of an embryonic stem cell, such as an embryonic stem cell phenotype, or may encompass a partial reversion of the differentiation state or a partial increase of the developmental potential of a cell, such as a somatic cell or a unipotent cell, to a multipotent state. Reprogramming may also encompass a partial reversion of the differentiation state of a cell to a state that renders the cell more susceptible to complete reprogramming to a pluripotent state when subjected to additional manipulations.

[0184] In some instances, a stem cell may comprise an animal stem cell. In some instances, an animal stem cell may comprise a mammalian stem cell, a bird stem cell, or a fish stem cell, a mollusk stem cell, or an amphibian stem cell. In some instances, an animal stem cell may comprise a mollusk stem cell. In some instances, an animal stem cell may comprise an amphibian stem cell. In some instances, an animal stem cell may comprise a mollusk stem cell. In some cases, a mammalian stem cell may comprise a porcine stem cell, a bovine stem cell, a bubaline stem cell, an ovine stem cell, a caprine stem cell, a cervine stem cell, a bisontine stem cell, a cameline stem cell, an elaphine stem cell, or a lapine stem cell. In some cases, a mammalian stem cell may comprise a porcine stem cell. In some cases, a bird stem cell may comprise an anatine stem cell, a galline stem cell, an anserine stem cell, a meleagrine stem cell, a struthionine stem cell, or a phasianine stem cell. In some instances, an animal stem cell may comprise a mammalian stem cell. In some instances, an animal stem cell may comprise a bird stem cell. In some instances, an animal stem cell may comprise a fish stem cell. In some cases, a stem cell may comprise a porcine stem cell. In some cases, a stem cell may comprise a bovine stem cell. In some cases, a stem cell may comprise a bubaline stem cell. In some cases, a stem cell may comprise an ovine stem cell. In some cases, a stem cell may comprise a caprine stem cell. In some cases, a stem cell may comprise a cervine stem cell. In some cases, a stem cell may comprise a bisontine stem cell. In some cases, a stem cell may comprise a cameline stem cell. In some cases, a stem cell may comprise an elaphine stem cell. In some cases, a stem cell may comprise a lapine stem cell. In some cases, a stem cell may comprise ananatine stem cell. In some cases, a stem cell may comprise a galline stem cell. In some cases, a stem cell may comprise an anserine stem cell. In some cases, a stem cell may comprise a meleagrine stem cell. In some cases, a stem cell may comprise a struthionine stem cell. In some cases, a stem cell may comprise a phasianine stem cell.

[0185] In some instances, an iPSC may comprise an animal iPSC. In some instances, an animal iPSC may comprise a mammalian iPSC, a bird iPSC, or a fish iPSC, a mollusk iPSC, or an amphibian iPSC. In some instances, an animal iPSC may comprise a mollusk iPSC. In some instances, an animal iPSC may comprise an amphibian iPSC. In some instances, an animal iPSC may comprise a mollusk iPSC. In some cases, a mammalian iPSC may comprise a porcine iPSC, a bovine iPSC, a bubaline iPSC, an ovine iPSC, a caprine iPSC, a cervine iPSC, a bisontine iPSC, a cameline iPSC, an elaphine iPSC, or a lapine iPSC. In some cases, a mammalian iPSC may comprise a porcine iPSC. In some cases, a bird iPSC may comprise an anatine iPSC, a galline iPSC, an anserine iPSC, a meleagrine iPSC, a struthionine iPSC, or a phasianine iPSC. In some instances, an animal iPSC may comprise a mammalian iPSC. In some instances, an animal iPSC may comprise a bird iPSC. In some instances, an animal iPSC may comprise a fish iPSC. In some cases, an iPSC may comprise a porcine iPSC. In some cases, an iPSC may comprise a bovine iPSC. In some cases, an iPSC may comprise a bubaline iPSC. In some cases, an iPSC may comprise an ovine iPSC. In some cases, an iPSC may comprise a caprine iPSC. In some cases, an iPSC may comprise a cervine iPSC. In some cases, an iPSC may comprise a bisontine iPSC. In some cases, an iPSC may comprise a cameline iPSC. In some cases, an iPSC may comprise an elaphine iPSC. In some cases, an iPSC may comprise a lapine iPSC. In some cases, an iPSC may comprise an anatine iPSC. In some cases, an iPSC may comprise a galline iPSC. In some cases, an iPSC may comprise an anserine iPSC. In some cases, an iPSC may comprise a meleagrine iPSC. In some cases, an iPSC may comprise a struthionine iPSC. In some cases, an iPSC may comprise a phasianine iPSC.

[0186] In some cases, iPSCs may comprise any cells obtained by re-programming of adult somatic cells which are endowed with pluripotency, a cell being capable of differentiating into the three embryonic germ cell layers, the endoderm, ectoderm and mesoderm. Such adult cells may be obtained from any adult somatic tissue (e.g. a skin fibroblast or blood cells) and undergo reprogramming by integrative genetic manipulation or non-integrative protein expression methods, which reset the cell to acquire stem cell-like characteristics. iPSCs may be formed through such processes that reverses the development of the cell or population of cells (e.g., a somatic cell) thus resulting in a naive cell type. An iPSC may be a cell that has undergone a process of driving a cell toa naive state with higher developmental and proliferation potential, such as a cell that is reset into a less differentiated state. The somatic cell, prior to induction to an iPSC, can be either partially or terminally differentiated. There may be a complete or partial reversion of the differentiation state, i.e., an increase in the developmental potential of a cell, to that of a cell having a pluripotent state. A somatic cell may be driven to a pluripotent state, such that the cell has the developmental potential of an embryonic stem cell, similar to an embryonic stem cell phenotype. Induction of a somatic cell may also encompass a partial reversion of the differentiation state or a partial increase of the developmental potential of a cell, such as a somatic cell or a unipotent cell, to a multipotent state. Induction may also encompass partial reversion of the differentiation state of a cell to a state that renders the cell more susceptible to complete induction to a pluripotent state when subjected to additional manipulations.Therapeutic or edible products

[0187] In some instances, a cell that has contacted the composition or has taken up the nucleic acid molecule may be cultured. Some or all of the cells thereof may subsequently be cultured to generate cultured cells, which cultured cells may be differentiated to generate terminally differentiated cells. In some cases, the cell comprising the nucleic acid molecules may be used to produce a therapeutic product. In some cases, the cell comprising the nucleic acid molecules may be used to produce a tissue. In some cases, the cell comprising the nucleic acid molecules may be used to produce an edible meat product. The terminally differentiated cells can be used to produce an edible meat product.

[0188] In some cases, a therapeutic product may have a therapeutic effect when administered. A therapeutic effect may comprise an inhibition, amelioration, mitigation, treatment, and / or prevention of a disease condition. In some cases, a therapeutic product may comprise a cell used in cell therapy. In some cases, a therapeutic product may comprise a cell, a vaccine, an organ, a product produced by the cell. The product produced by the cell may comprise a protein, a nucleic acid molecule, a chemical compound, or a combination thereof. In some cases, a protein may comprise an antibody, an enzyme, a signaling molecule, an enzyme inhibitor, a hormone, a cytokine, a growth factor or a combination thereof. A nucleic acid molecule used as a therapeutic product may comprise any nucleic molecules described elsewhere in this disclosure. In some cases, a chemical compound may comprise an organic or inorganic compound. In some cases, a compound may comprise a nutrient. In some cases, a cell used as a therapeutic product may comprise any cell described elsewhere in thisdisclosure. In some cases, the therapeutic product may comprise a human or a non-human cell, protein, nucleic acid molecule, chemical compound, or a combination thereof. In some cases, a product produced by the cell may comprise an antimicrobial molecule. The antimicrobial molecule may be an antibacterial, an antifungal, or an antiparasitic molecule.

[0189] A therapeutic product may comprise a medicine. A medicine may comprise a chemical or a collection and / or mixture of chemicals that has a therapeutic effect when administered. In some cases, a therapeutic product may comprise a drug product. In some cases, a drug product may be a therapeutic product in a dosage form. In some cases, a therapeutic product may comprise a pharmaceutically active ingredient. A pharmaceutically active ingredient may comprise a chemical and / or cell that has a direct pharmaceutical activity contributes to and / or is responsible for the inhibition, amelioration, mitigation, treatment, and / or prevention of a disease condition. In some cases, a therapeutic product may comprise a drug substance. In some cases, a drug substance may comprise a pure form of the pharmaceutically active ingredient. In some cases, a sample comprising a drug substance may comprise at least 50 %, 60 %, 70 %, 80 %, 90 %, or 100 %, by volume, weight, and / or number of molecules, of the drug substance in the sample.

[0190] In some cases, a composition as described herein may be used in a method of treating a disease. For example, the composition may comprise a nucleic acid molecule which is a therapeutic nucleic acid molecule. The compositions described herein may enhance delivery of such therapeutic nucleic acid molecules to a cell, in order to exert a therapeutic effect. Accordingly, described herein is a composition for use in a method of treating a disease, the composition comprising (i) a nucleic acid molecule, wherein the nucleic acid molecule is a therapeutic nucleic acid molecule, and (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2). The composition may further comprise a cationic lipid molecule. The cationic lipid molecule may be selected from the group consisting of DOTAP and any compound of Formula (I)-(IX). The composition may further comprise a structural lipid, such as DOPE.

[0191] A tissue may comprise a collection of cells. The cells of a tissue may be from one cell type. The cells of a tissue may be from more than one cell type. In some cases, a tissue or group of cells may form an organ. In some cases, the group of tissues or cells of an organ may be found in an animal. In some cases, the tissues or cells from an organ may collectively perform a physiological or cellular function. In some cases, the tissues or cells from an organ may share structural and / or functional characteristics. In some cases, an organ may comprise a bladder, a blood vessel, a bone, abrain, a bronchi, a cartilage, a diaphragm, a fallopian tube, a gill, a hair, a heart, a hypothalamus, an intestine, a kidney, a larynx, a ligament, a liver, a lung, a lymph node, a muscle, a nail, a nerve, an ovary, a pancreas, a parathyroid, a penis, a pharynx, a pineal body, a pituitary gland, a prostate, a scale, a skin, a spinal cord, a spleen, a stomach, a tendon, a testis, a thymus, a thyroid, a tonsil, a tooth, a trachea, a ureters, a urethra, a vagina, a vas deferen, a vulva, or a combination thereof. In some cases, a tissue or group of cells may form an organoid. In some cases, the organoid may be formed in an in vitro or ex vivo culture. In some cases, the organoid may be 3D. In some cases, the tissues or cells from an organoid may collectively perform a physiological or cellular function. In some cases, the tissues or cells from an organoid may share structural and / or functional characteristics. In some cases, a model may comprise a cell or a collection of cells. In some cases, a model comprising a cell or a collection of cells may be a model of the organ, the organoid, the tissue, or a combination thereof. In some cases, a model comprising a cell or a collection of cells may be a model of the organ. In some cases, a model comprising a cell or a collection of cells may be a model of the organoid. In some cases, a model comprising a cell or a collection of cells may be a model of the tissue. In some cases, a model may have at least one functional and / or structural characteristic of the cell, the collection of cells, the tissue, the organ, the organoid, or a combination thereof. In some cases, a model may comprise organic or inorganic materials. In some cases, a model may comprise a cell, tissue, organ, or organoid; or a component derived thereof. In some cases, a model may be a biomimetic model.

[0192] An edible product may also comprise a group of cells. An edible product may also comprise a group of cells having a population of cell types of a tissue from an animal. An edible product may be edible when it is consumed by any species, and it causes no harm. Such harm may comprise poisoning, contamination, or infection. An edible product may comprise an edible meat product. The edibility of a meat may depend on the types or properties of the meat. For example, the meats may be processed or cooked to be edible. In some cases, meats may be baked, steamed, poached, boiled, grilled, dried, smoked, fried, heated, pickled, fermented, aged, or any combination thereof. Some meats, such as those from fish, may be edible without cooking. The edibility of a meat may depend on the level of toxins and contaminating organisms. In some cases, the edibility of a meat may also depend on the appetite of a person.

[0193] In some cases, cultured cells may receive some degrees of structural integrity from a scaffold on which the cells may be attached during culturing. A cell may also be cultured in cell suspensions. In some cases, a cell may be adherent. In other cases, a cell may not be adherent. In some cases, non-adherent cells may or may not require a substrate or surface for attachment. In some cases, cells may have been selected, evolved, modified or engineered to not require an adherence substrate. In some cases, cultured cells may be grown or configured to form cultured tissues that may be attached to a support structure such as a two-dimensional (2D) or three-dimensional (3D) scaffold or support structure. In some cases, cultured cells may be configured to form various shapes or forms by a three-dimension printer. In some cases, cultured cells may be grown on a two-dimensional support structure such as a tissue-culture plate where they may form several layers of cells that may be peeled and processed for consumption. In some cases, two-dimensional support structures may include porous membranes that allow for diffusion of nutrients from culture media on one side of the membrane to the other side where the cells are attached. In such a composition, additional layers of cells may be achieved provided media perfusion is sufficient e.g. by exposing the cells to culture media from both sides of the membrane. In some cases, cells may receive nutrients through diffusion from one side of the membrane and also from the culture media covering the cells growing on the membrane. Culture media may be replenished in the two-dimensional tissue culture environment to prevent the buildup of waste metabolites.

[0194] In some cases, cultured cells may be grown on, around, or inside a three-dimensional support structure. In some cases, the support structure may be sculpted into different sizes, shapes, and / or forms to provide the shape and form for the cultured cells to grow and resemble different types of tissues such as steak, tenderloin, shank, chicken breast, drumstick, lamb chops, fish fillet, or lobster tail. The support structure may be a natural or synthetic biomaterial. In some cases, a biomaterial may comprise any substance intended to interface with biological systems to evaluate, treat, augment, or replace any tissue, organ, or function in a biocompatible manner, such as with a level of acceptable biological response. In some cases, a biomaterial may interact passively with cells and tissues or may comprise a bioactive material which induces a specific and intended biological response. In some cases, a biomaterial may comprise a substrate that has been engineered to take a form which alone or as part of a complex system, is used to direct, by control of interactions with components of living systems. In some cases, a biomaterial may be natural, synthetic, or some combination thereof. A scaffold may be composed of one material or one or more different materials. In some cases, a scaffold may be non-toxic and edible so that they may not be harmful if ingested and may provide additional nutrition, texture, flavor, or form to the final food product. In some cases, a scaffold may comprise a hydrogel, a biomaterial such as an extracellular matrix molecule (ECM), cellulose, or biocompatible synthetic materials. ECM molecules may compriseproteoglycans, non-proteoglycan polysaccharides, or proteins. In some cases, a micro-scaffold may be smaller than a traditional tissue culture scaffold which may provide a macroscopic structure and / or shape for the cell population. In some cases, a micro-scaffold may provide a surface for adherent cells to attach to even while the micro-scaffold itself is in suspension. In some cases, a micro-scaffold may provide a seed or core structure for adherent cells to attach while remaining small enough to remain in suspension with stirring. The use of micro-scaffolds enables the culturing of adherent cells in a suspension culture which may enable the large-scale production of adherent cells. In some cases, cultured cells may be allowed to grow in aggregates, spheroids or embryoid bodies. ECM molecules will form between the cells providing a naturally occurring tissue culture scaffold to which cells continue to grow and adhere to.

[0195] A degradable scaffold may comprise a polymeric material. A polymeric material may comprise a natural polymeric material or a synthetic polymeric material. Biomaterials may comprise collagen, gelatin, fibrin, alginate, agar, cassava, maize, chitosan, gellan gum, corn-starch, chitin, cellulose, chia (Salvia hispanica), recombinant silk, decellularized tissue (plant or animal), hyaluronic acid, fibronectin, laminin, hemicellulose, glucomannan, textured vegetable protein, heparan sulfate, chondroitin sulfate, tempeh, keratan sulfate, or any combination thereof. A plantbased scaffold may be used for 3D culturing. A plant-based scaffold may comprise scaffolds obtained from plants such as apples, seaweed, or jackfruit. A plant-based scaffold may comprise at least about one plant-based material such as cellulose, hemicellulose, pectin, lignin, alginate, or any combination thereof. A textured vegetable protein (TVP), such as textured soy protein (TSP) may comprise a high percentage of soy protein, soy flour, or soy concentrate. TVP and TSP can be used to provide a meat-like texture and consistency to a meat product. Synthetic biomaterials may comprise hydroxyapatite, polyethylene terephthalate, acrylates, polyethylene glycol, polyglycolic acid, poly caprolactone, polylactic acid, their copolymers, or any combination thereof.

[0196] In some instances, a method may comprise expanding or culturing a cell. In some instances, an expanding or a culturing may comprise maintenance media, differentiation media, steatotic media, or proliferation media. In some cases, a media may be configured to promote cell culturing or expansion. In some cases, gas balance in the media may comprise a mixture of oxygen from about 21% to about 95% air saturation, Carbon dioxide partial pressure (mm Hg) from about 0% to about 10%. For example, a mixture of media of about 80% Oxygen about 5% carbon dioxide and held at 37°C with controlled pH may be provided. In some cases, complete transfection media are equilibrated overnight at about 37-39° C and 5% CO2 and pH adjustment is performed with. Incertain cases, terminal differentiation may comprise the sequential steps of culturing or maintaining a plurality of substantially undifferentiated pluripotent cells in a first defined media comprising at least about one growth factor, and incubating the cells in a second defined media which is sufficient to promote differentiation in a plurality of cells. In some aspects, a plurality of the pluripotent cells is differentiated into endodermal, ectodermal or mesodermal cells HPCs. In certain aspects, the second defined media may comprise FGF2, IGF-1. In certain aspects, the second defined media may comprise a GSK-3 inhibitor. In some cases, the first defined media further may comprise IGF-1, NRG-1, TGF-B, LIF and FGF2. In some cases, the second defined media further may comprise FGF2. In some cases, the method may comprise culturing the cells at an atmospheric pressure of less than 25% oxygen, such as less than 24%, 23%, 22%, 21%, 20%, 19%, 18%, 17%, 16%, 15%, 10%, 5%, or 0% oxygen.

[0197] In some cases, a maintenance media may comprise pluripotency media. In some cases, cells are treated with high dose (50 ng / mL) porcine- FGF2 for 48 hours prior to transfection. In some instances, a media is changed to second defined media after 72 hours. In some instances, at least about a portion of an expanding or a culturing is performed using a growth mode other than simple batch culture. As used herein for culturing cells, culturing a cell in vitro may also comprise culturing a cell, a group of cells, or a tissue ex vivo or outside of an organism or a host. A cell culture, in some cases, may also comprise the maintenance or induction of the differentiation (or de-differentiation) of a cell. In other cases, a cell culture may be maintained in growth media. A cell culture, in some cases, may be 2-dimensional (2D) or 3 -dimensional (3D). In some cases, growth media may comprise nutrients or other components required for the growth of a cell. In some cases, the types of media and the nutrients for a cell culture may depend on the cell being cultured or the purpose of the cultured cell.

[0198] The term “about” or “approximately” as used herein when referring to a measurable value such as an amount or concentration and the like, is meant to encompass variations of 20 %, 10 %, 5 %, 1 %, 0.5 %, or even 0.1 % of the specified amount. For example, “about” can mean plus or minus 10 %, per the practice in the art. Alternatively, “about” can mean a range of plus or minus 20 %, plus or minus 10 %, plus or minus 5 %, or plus or minus 1 % of a given value. Alternatively, particularly with respect to biological systems or processes, the term can mean within an order of magnitude, up to 5 -fold, or up to 2-fold, of a value. Where particular values can be described in the application and claims, unless otherwise stated the term “about” may be assumed to encompass the acceptable error range for the particular value. Also, where ranges, subranges, or both, of values can be provided, theranges or subranges can include the endpoints of the ranges or subranges. The terms "substantially", "substantially no", "substantially free", and "approximately" can be used when describing a magnitude, a position or both to indicate that the value described can be up to a reasonable expected range of values. For example, a numeric value can have a value that can be + / - 0.1 % of the stated value (or range of values), + / -1 % of the stated value (or range of values), + / - 2 % of the stated value (or range of values), + / - 5 % of the stated value (or range of values), + / - 10 % of the stated value (or range of values), etc. Any numerical range recited herein can be intended to include all sub-ranges subsumed therein.

[0199] Where values are described as ranges, it may be understood that such disclosure includes the disclosure of all possible sub-ranges within such ranges, as well as specific numerical values that fall within such ranges irrespective of whether a specific numerical value or specific sub-range is expressly stated.

[0200] The terms “comprise,” “have,” and “include” are open-ended linking verbs. Any forms or tenses of one or more of these verbs, such as “comprises,” “comprising,” “has,” “having,” “includes,” and “including,” are also open-ended. For example, any method that “comprises,” “has,” or “includes” one or more steps is not limited to possessing only those one or more steps and also covers other unlisted steps.EXAMPLES

[0201] These examples are provided for illustrative purposes only and not to limit the scope of the claims provided herein.

[0202] Example 1. Transfection of RNAs into cells using transfection complexes described herein

[0203] Provided here are methods for using transfection complexes described herein for transfecting cells.

[0204] As described herein, DOTAP is cost prohibitive to obtain. It is desirable to identify the lowest RNA / DOTAP ratio to minimize the cost of delivery.

[0205] Delivery of saRNA by DOTAP only without any other components could be achieved when saRNA / DOTAP mass ratio was about 1 to 4. However, as shown in FIG. 1A, transfection efficiency as demonstrated by GFP expression was very low.

[0206] Transfection complex was then generated and tested using various cationic lipids and glycolipids as described herein. Cationic lipids were selected from lipids containing quaternary-n-ammonium groups such as DOTAP or tertiary, secondary amino group such as

[0207] Glycolipids are food-grade degradable surfactants and may be selected from, but is not limited to, sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate. Transfection complexes can contain small amounts of cationic DOTAP (saRNA / DOTAP mass ratio does not exceed 1 to 4, optionally 1 to 2). Cationic lipid / gly colipid mass ratio was used to optimize the formulation for delivery of different nucleotides to different cell types (2D- or 3D-cell culture). For example, for saRNA delivery, cationic lipid / glycolipid mass ratio may include, but is not limited to, from 1 part of DOTAP to 5 parts of SMO, to 1 part of DOTAP to 100 parts of SMO to deliver 0.25 parts of saRNA.

[0208] FIGs. 1B-1E show the transfection efficiency of transfection complexes having DOTAP and various esterified saccharides (and amounts) or in different solvents in different types of cell cultures. FIG. IB shows that transfection efficiency was significantly higher than that of FIG. 1A when using saRNA transfection with a complex comprising DOTAP / STO (saRNA / DOTAP / STO mass ratio 1 / 4 / 10). FIG. 1C shows that transfection efficiency was significantly higher than that of FIG. 1A when using saRNA transfection with a complex comprising DOTAP / SMO (saRNA / DOTAP / SMO mass ratio 1 / 4 / 10). FIG. ID shows that transfection efficiency of 3D cultured cells was high when using saRNA transfection with a complex comprising DOTAP / SMO (saRNA / DOTAP / SMO mass ratio 1 / 2 / 40). FIG. IE shows that alcohol-based solvent (ethanol) could increase the transfection efficiency of the transfection complex in 3-D culture.

[0209] Further experiments show that transfection efficiency of cationic lipid / esterified saccharide can be improved with the addition of an additional lipid l,2-di-(9Z-octadecenoyl)-sn-glycero-3- phosphoethanolamine (DOPE; as a structural lipid). In three-component transfection complex cationic lipid / glycolipid / DOPE mass ratio can be used to determine the formulation delivery of different nucleotides to different cell types (2D- or 3D-cell culture). For example, for saRNA delivery, cationic lipid / glycolipid / DOPE mass ratio may include, from 1 part of DOTAP to 5 parts of SMO and 0.5 parts of DOPE, to 1 part of DOTAP to 100 parts of SMO and 4 parts of DOPE to deliver 0.25 parts of saRNA.

[0210] FIGs 2A-2B show that the transfection efficiency of the three-component transfection complex was significantly higher than that of the two-component transfection complex (with vs.without structural lipids). FIG. 2A shows that GFP expression was significantly higher in cultures transfected with saRNA transfection complex with three-component DOTAP / DOPE / MMO (saRNA / DOTAP / DOPE / MMO mass ratio 1 / 4 / 16 / 24; right) system compared with a two-component system. FIG. 2B shows that GFP expression was significantly higher in cultures transfected with saRNA transfection with DOTAP / SMO (saRNA / DOTAP / SMO mass ratio 1 / 2 / 40; left) and DOTAP / DOPE / SMO (saRNA / DOTAP / SMO / DOPE mass ratio 1 / 2 / 40 / 2; right). Additionally, as depicted in FIG. 2C, alcohol-based solvent significantly increased the transfection efficiency of the transfection complex.

[0211] The transfection complexes described herein can also be used to transfect various sequences and types of nucleic acids.

[0212] As shown in FIG. 3A, MyHC expression was observed in 3D cultured cells after MyoD- saRNA transfection using DOTAP / DOPE / SMO (saRNA / DOTAP / SMO / DOPE mass ratio 1 / 2 / 40 / 2). As shown in FIG. 3B, transfection efficiency of GFP carried on a plasmid DNA (pDNA) with a transfection complex described herein (DOTAP / DOPE / SMO; saRNA / DOTAP / SMO / DOPE mass ratio 1 / 2 / 40 / 2) in 2D-culture was significantly higher than that of using lipofectamine

[0213] To determine the cationic lipids that could increase the transfection efficiency of the transfection complex, a series of cationic lipids were generated (the methods for synthesizing the lipids are described elsewhere in this disclosure, such as those described in Example 2).

[0214] A library of lipids was developed based on carnitine derivatization, such as one described in formula (I), in which R1, R2, R3are hydrogen atom, saturated hydrocarbon chain, mono- and multiunsaturated hydrocarbon chains.

[0215] For example, using the cationic lipid of formula (IV) to generate various transfection complex (saRNA / lipid / SMO / DOPE mass ratio 1 / 2 / 40 / 2), significant GFP expression was observed when it was used to transfect 3D culture cells, as shown in FIG. 4.

[0216] saRNA transfection complex based on the same lipid with composition (saRNA / lipid / DOPE / SMO mass ratio 1 / 4 / 6 / 24) demonstrated, significant GFP expression was observed when it was used to transfect 2D culture cells, as shown in FIG. 5A.

[0217] Using the cationic lipids of formula (IV), (V), (VI), and (VII) to generate various transfection complex (saRNA / lipid / DOPE / SMO mass ratio 1 / 4 / 6 / 24, 1 / 5.3 / 6.7 / 16, and 1 / 5.3 / 6.7 / 16, respectively), significant GFP expression was observed when it was used to transfect 3D culture cells, as shown in FIGs. 5A-5D.

[0218] Thus, the data in this example shows that various components and dosages (ratio between each components) can be modified to significantly increase the transfection efficiency of various sequences or types of nucleic acids, using the methods and compositions as described herein. The methods described herein can also be used to identify the individual components of the transfection complexes (nucleic acids, saccharide, lipids, or solvents) for transfecting cells.

[0219] Example 2. Synthesis of lipids based on ethanolamine / carnitine.

[0220] Provided herein are methods for synthesizing the cationic lipids as described herein. Lipids of formula (I) or (II) were synthesized by alkylation of ethanolamine with alkyl bromides using different ethanolamine / alkyl bromide mol ratio between 1 / 1 to 1 / 5. Specifically, 0.12 g of ethanolamine was mixed (1.95 mmol) with 2.0 g of 1 -bromohexadecane (6.25 mmol) and 10 ml of acetonitrile. Mixture was heated at 80 °C under stirring for 72 hr. The reaction mixture was cooled down to room temperature, the white solid precipitated was filtered, and washed with acetonitrile. FIG. 5 A shows the Ih NMR spectra of the synthesized lipid.

[0221] Lipids of formula (III) were synthesized by esterification of carnitine with alkyl bromides in alkaline media. Specifically, 1 g of L-carnitine (6.20 mmol) was dissolved in 7 mL of acetonitrile. 2.36 g of 1 -bromohexadecane (7.75 mmol) were added to the solution. Reaction mixture was left under stirring for 118 hours (hr) at 80 °C. The L-carnitine did not dissolve initially but dissolved with time (after 72 hr the reaction mixture was a colorless solution). After 118 hr, the solution was cooled and white precipitate was formed. 80 mL of diethyl ether were added to the suspension and left under stirring for 30 mins. Precipitate was isolated by filtration and washed with diethyl ether (20 mL). FIG. 6 shows the IH NMR spectra of the synthesized lipid.

[0222] Example 3. Induction of Protein Expression, Cell Differentiation, or Cell Conversion Using Saccharides, Lipids, and Nucleic Acid Molecules

[0223] Provided herein are methods for facilitating protein expression, cell differentiation, and / cell conversion using the compositions / methods described in this disclosure.

[0224] FIG. 7 depicts a workflow of the method described herein. Step 701 comprises mixing a saccharide molecule and / or a lipid molecule as described herein with nucleic acids (e.g., RNA, mRNA, or saRNA) to generate a composition. Step 702 comprises contacting a cell population (e.g., stem cells, embryonic stem cells, immortalized stem cells, mesenchymal stem cells, muscle progenitor cells, or iPSCs) with the composition. Step 703 comprises subjecting the cells of the cell population in a sufficient condition so that at least a subset of the cells takes up the nucleic acidmolecules of the nucleic acid molecule. Step 704 comprises facilitating the transfected cells to alter their protein expression. Optionally, step 705 comprises facilitating the differentiation of the transfected cells. Optionally, step 706 comprises converting the transfected cells into a pharmaceutical active ingredient, a tissue, or an edible product.

[0225] A few milligrams of modified saccharide, 2 mg of thiamine pyrophosphate, and 0.5 mg of protamine sulfate are independently dissolved in 1 mL of sterile distilled water. Each solution is filtered through a 0.22 pm filter. Targeting nucleic acid is dissolved to a final concentration of 1 mg / mL in sterile distilled water. PEG-lipid is dissolved in 1 mL of ethanol with the total PEG-lipid weight amounting to 30 mg, followed by filtration through a 0.22 pm filter. The nucleic acid solution is mixed with the protamine solution to form pre-complexes and then mixed with the water- soluble saccharide solution, the thiamine pyrophosphate solution, and the PEG-lipid solution to prepare compositions containing lipoplexes.

[0226] Cells (listed in Table 1) are transfected daily with various saccharides and nucleic acid molecules (listed in Table 2 and 3, respectively) between 1-7 days. GFP / RFP / YFP mRNA, or scrambled siRNA are used as a transfection control. Transfection is carried out using either traditional chemical based methods (e.g. Lipofection), or non-chemical methods (e.g. electroporation or nucleofection) as controls for the colloidal nanoparticle method of choice. The diverse nature of nucleotides affects the delivery method chosen as can be seen in the difference of nucleotide lengths, double vs single stranded nucleic acids, and the dose range of nucleotides: Silencing RNA (siRNA): 20-40bps, double-stranded RNA molecule, messenger RNA (mRNA): range of 500bp-2-4kbp, single stranded RNA molecule. Dose range of nucleotides: 0.5 pg / mL- 50 pg / mL per nucleotide (DNA, RNA, mRNA, siRNA, saRNA). For example, mRNA and siRNA may be encapsulated together using a nanoparticle transfection option.Table 1. Example cell typesTable 2. Examples of modified saccharidesTable 3. Examples of nucleic acids

[0227] Example 4. Delivery of Nucleic Acids Into Cells Within an Organism in vivo

[0228] Provided herein are methods for delivery nucleic acids into the cells of an organism in vivo.

[0229] FIG. 8 depicts a workflow of the method described herein. Step 801 comprises mixing a saccharide molecule and / or a lipid molecule as described herein with nucleic acids to generate a composition. Step 802 comprises injection of the composition into a vein. Step 803 comprises subjecting the organism in a sufficient condition so that at least a subset of the organs takes up the nucleic acid molecules.

[0230] The capacity of the different formulations to delivery an mRNA cargo in vivo was studied in a biodistribution study. Two different test compositions were complexed with Firefly Luciferase mRNA and were given to Balb / c mice through an intravenous (IV) injection. 6 hours and 24 hours post injection, luciferase mRNA expression was visualized using in vivo imaging system (IVIS) (IV injection of the transfection complex with the mice; with 0.5 mg of RNA per 1 kg of mouse mass (0.5 mg / kg)). Test compositions and positive / negative controls were generated using the methods described herein and dosed with the mice only one time. At the end of the study, animals were sacrificed, and lungs, liver and spleen were collected. Collected tissues were imaged again with IVIS. In total, 4 groups of experiments were carried out (Group 1, 2, 3, 4 correspond to negative control, positive control, test composition 1, and test composition 2, respectively). For transfection testcomposition 1 , Flue mRNA was mixed with a cationised polysaccharide, a cationic lipid and a neutral saccharide-based lipid. Similarly, test composition (2) was made by mixing Flue mRNA with a cationic lipid, helper lipid DOPE and a neutral saccharide-based surfactant. Body weights of the mice from different experiment groups were measured for at least 3 days post-transfection. No statistically significant difference were detected between the groups.

[0231] In vivo IVIS imaging was performed 6 hours and 24 hours post dosing. In imaging, animal was weighed and approximately 150 mg / kg, 5 ml / kg Luciferin in Saline was injected, as i.p 10 minutes prior to IVIS scan. Then animal was anesthetized with isoflurane and placed into a small animal IVIS scanner (Lumina II, Caliper LifeScience) lying on their back (only one animal per scan). Thereafter, a 1 minute whole body IVIS scan was be performed. Image analysis was performed with Living Image software using 2D software. Rectangle ROI was drawn over the whole animal. Same size ROIs was used for all animals. All IVIS images were background corrected and same logarithmic scale was used in all images (data not shown). IVIS images were analyzed using the background corrected data. Data were transformed into logarithmic form so that the statistical analysis could be performed (the right panel of FIG. 9A, wherein the3 left panel of FIG. 9A shows the comparison of the experimental groups in untransformed fluorescence levels. Statistically significant differences were observed between Groups 1 and 2, Groups 2 and 3 and Group 2 and 4 in all time points. There was also a statistically significant difference between Groups 1 and 3 at timepoint 6 hour post dose.

[0232] Ex vivo IVIS was also performed to determine the biodistribution of the nucleic acids / transfection complexes in the 4 experimental groups. Lung, liver, and spleen tissues were collected and placed individually into wells of a 12- well plate immediately after collection. Luciferin solution was added to well until tissue was covered with solution and then well plate was imaged with small animal IVIS scanner. IVIS images were background corrected and same logarithmic scale was used in all images (data not shown). IVIS images were analyzed using the background corrected data and the logarithmic transformed data (see left and right panels of FIG. 9B, respectively). As depicted in FIG. 9B, only low levels of signals in spleen, liver, and lungs were detected in negative control animals (Groupl). Positive control animals (Group 2) had significantly higher signal in liver compared to other groups. Compositions of Group 3 and 4 as well as Positive control animals (Group 2) had higher uptake in spleen compared to negative control animals (Groupl). Test item 1 (Group 3) showed the highest uptake to lungs compared to other groups.

[0233] The data show the in vivo biodistribution of two compositions as described herein, and positive and negative controls in Balb / c mice using IVIS 6 hours and 24 hours post dosing. Signal intensitydifference was observed between the groups. The highest signal was observed the in positive control animals, whereas no signal was observed in the negative control animals. In ex vivo IVIS, no signal was observed in the negative control animals. The highest level of signal was detected in the positive control animals, especially in liver and spleen. The composition of Group 1 was most detected in the lung and spleen, whereas the composition of Group 2 was detected specifically in spleen.

[0234] Thus, the compositions described herein could efficiently deliver the nucleic acids into specific cells / tissues in an organism in vivo.

[0235] Example 5. Culturing an edible meat product

[0236] Cells are maintained and expanded in the first growth media supplemented with the required growth factors for the cell line in question. Cells are grown either on a 2D adherent surface or in 3D as aggregates. To promote protein expression, cells are treated for 24 hours with the lipoplex containing the nucleic acid self-assembled with the chosen delivery agent. Transfections are carried out in 2D (with or without biomaterial) or 3D (including but not limited to: spheroid, embryoid bodies, suspension or adherent, with or without biomaterial) culture conditions. Media may be changed after 24 hours. Cells are maintained in the second growth media for 7-50+ (min-max or anywhere in between) days depending on the desired outcome required. Maturation of cultures are carried out in the described 2D or 3D conditions, with or without biomaterial, or with or without electrical stimulation or contractile tension forces e.g. to promote maturation of myogenic fibers. The necessary controls are carried out for experimental consistency. Analysis may be conducted at any stage using qPCR, immunohistochemistry or flow cytometry. Any nucleic acid may be used in permutations to this methodology. Experimental changes may use the same materials and methods, but different compounds may be introduced. Following the required growth of cells (e.g. muscle cells expressing relevant myogenic markers, fat cells expressing relevant adipogenic markers), the cells are harvested and enter into food processing. Briefly, the cells are blended with plant based ingredients to the required quantities dependent on the end product in question. Between 1-99% (min-max) cell mass (or anywhere in between) may be used in the final 100% formulation. The formulation is formed into the desired shape and structure , sliced, cooked and frozen.

[0237] Example 6: in vitro delivery of siRNA

[0238] HEK cells were seeded at 2e4 / cm2and treated after 24 hours with (+) or without (-) 50pM siRNA [GAPDH], in the presence (+) or absence (-) of saRNA [GFP] which was used as a transfection control. Lipofectamine 2000 was used as a transfection control. The data in Fig. 10 shows that thedelivery composition, comprising a cationic lipid and a sorbitan polysaccharide, is able to deliver siRNA that is functional, and able to reduce GAPDH levels comparable to control.

[0239] As represented in Fig. 11, three cell lines (A549, MRC-5 and HEK-293) were seeded at densities 2e4 / cm2, 1.25e4 / cm2, and 2e4 / cm2respectively. Control cells were not transfected, and condition cells were transfected with 50pM siRNA [GAPDH], Lipofectamine 2000 was used as a transfection control. The data in Fig. 11 shows that the delivery composition, comprising a cationic lipid and a sorbitan polysaccharide, is able to deliver siRNA that is functional, and able to reduce GAPDH levels across all cell lines.

[0240] While preferred embodiments of the present disclosure have been shown and described herein, it may be obvious to those skilled in the art that such embodiments are provided by way of example only. It is not intended that the disclosure be limited by the specific examples provided within the specification. While the disclosure has been described with reference to the aforementioned specification, the descriptions and illustrations of the embodiments herein are not meant to be construed in a limiting sense. Numerous variations, changes, and substitutions may now occur to those skilled in the art without departing from the disclosure. Furthermore, it shall be understood that all aspects of the disclosure are not limited to the specific depictions, configurations or relative proportions set forth herein which depend upon a variety of conditions and variables. It may be understood that various alternatives to the embodiments of the disclosure described herein may be employed in practicing the disclosure. It is therefore contemplated that the disclosure shall also cover any such alternatives, modifications, variations or equivalents. It is intended that the following claims define the scope of the disclosure and that methods and structures within the scope of these claims and their equivalents be covered thereby.

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A method, comprising: a. providing a cell with a composition comprising: i. a nucleic acid molecule, and ii. a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2); and b. delivering said composition into said cell.

2. The method of claim 1, wherein said saccharide molecule comprises said sorbitan, said mannide, said trehalose, said functional variant thereof, said derivative thereof, or any combination thereof.

3. The method of claim 2, wherein said saccharide molecule comprises a second ester group.

4. The method of claim 1, wherein said saccharide molecule comprises said lactose comprising said ester group.

5. The method of any one of claims 1-4, wherein said composition further comprises a lipid molecule.

6. A method, comprising: a. providing a cell with a composition comprising: i. a nucleic acid molecule, ii. a saccharide molecule and / or a lipid molecule; and iii. a polar organic solvent miscible with water; and b. delivering said composition into said cell.

7. The method of claim 6, wherein said polar organic solvent is at most about 16 %, by volume, of said composition.

8. The method of claim 7, wherein said polar organic solvent is at most about 8 %, by volume, of said composition.

9. The method of any one of claims 6-8, wherein said polar organic solvent is DMSO, DMF, ethanol, propanol, 2-propanol, ethylene glycol, a functional variant thereof, a derivative thereof, or any combination thereof.

10. The method of claim 9, wherein said polar organic solvent is DMSO.

11. The method of claim 9, wherein said polar organic solvent is ethanol.

12. The method of any one of claims 6-11, wherein said composition comprises an alcohol-based solvent.

13. The method of any one of claims 6-12, wherein said saccharide molecule comprises a monosaccharide.

14. The method of any one of claims 6-12, wherein said saccharide molecule comprises a disaccharide.

15. The method of any one of claims 6-12, wherein said saccharide molecule comprises an oligosaccharide.

16. The method of any one of claims 6-12, wherein said saccharide molecule comprises a polysaccharide.

17. The method of any one of claims 6-16, wherein said composition comprises said saccharide molecule and said lipid molecule.

18. The method of any one of claims 5-17, wherein said lipid molecule is cationic in an aqueous neutral solution.

19. The method of claim 18, wherein said lipid molecule comprises DOTAP.

20. The method of claim 18 or 19, wherein said lipid molecule comprises a lipid molecule having a structure of formula (I), (II), or (III).

21. The method of claim 20, wherein said lipid molecule is selected from the group consisting of a compound of formula (IV)-(IX).

22. The method of any one of claims 1-21, wherein said saccharide molecule is neutral in an aqueous neutral solution.

23. The method of any one of claims 1-22, wherein said saccharide molecule comprises sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate, a derivative thereof, or any combination thereof.

24. The method of any one of claims 1-23, wherein said saccharide molecule comprises a lipid moiety.

25. The method of claim 24, wherein said lipid moiety comprises a fatty acyl.

26. The method of claim 25, wherein said fatty acyl is monounsaturated.

27. The method of claim 25, wherein said fatty acyl is polyunsaturated.

28. The method of claim 24, wherein said lipid moiety comprises a fatty hydrocarbon chain.

29. The method of claim 28, wherein said fatty hydrocarbon chain is saturated.

30. The method of claim 28, wherein said fatty hydrocarbon chain is monounsaturated.

31. The method of claim 28, wherein said fatty hydrocarbon chain is polyunsaturated.

32. The method of any one of claims 1-31, wherein said nucleic acid molecule comprises a ribonucleic acid (RNA); optionally wherein said RNA comprises a messenger RNA (mRNA), a self-amplifying RNA (saRNA), a small interfering RNA (siRNA), a transfer RNA (tRNA), a small activating RNA (RNAa), an endl ess / circular RNA (eRNA), a complementary DNA (cDNA), a plasmid DNA (pDNA), or any combination thereof.

33. A composition, comprising: a lipid molecule having a structure of formula (I), formula (II), or formula (II):wherein:R1is independently C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl; each of R2and R3is independently H, C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl;R4 is C6-C22 alkyl, C6-C22 alkenyl, or C6-C22 alkynyl;R5 is independently C1-C4 alkyl; andX is independently an anionic radical selected from the group consisting of: hydroxide, halide, sulfate, methylsulfate, ethylsulfate, and phosphate ions.

34. The composition of claim 33, wherein said lipid molecule is selected from the group consisting of:

35. The composition of claim 33 or claim 34, wherein said composition further comprises a nucleic acid molecule.

36. The composition of any one of claims 33-35, wherein said composition further comprises a saccharide molecule.

37. The composition of claim 36, wherein said saccharide molecule comprises a monosaccharide.

38. The composition of claim 36, wherein said saccharide molecule comprises a disaccharide.

39. The composition of claim 36, wherein said saccharide molecule comprises an oligosaccharide.

40. The composition of claim 36, wherein said saccharide molecule comprises a polysaccharide.

41. The composition of any one of claims 36-40, wherein said saccharide molecule is neutral in an aqueous neutral solution.

42. The composition of any one of claims 36-41, wherein said saccharide molecule comprises a sorbitan, a mannide, a trehalose, a lactose, a functional variant thereof, a derivative thereof, or any combination thereof; optionally wherein said saccharide molecule comprises sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate, a derivative thereof, or any combination thereof.

43. The composition of any one of claims 36-42, wherein said saccharide molecule comprises a lipid moiety.

44. The composition of claim 43, wherein said lipid moiety comprises a fatty acyl.

45. The composition of claim 44, wherein said fatty acyl is monounsaturated.

46. The composition of claim 44, wherein said fatty acyl is polyunsaturated.

47. The composition of claim 43, wherein said lipid moiety comprises a fatty hydrocarbon chain.

48. The composition of claim 47, wherein said fatty hydrocarbon chain is saturated.

49. The composition of claim 47, wherein said fatty hydrocarbon chain is monounsaturated.

50. The composition of claim 47, wherein said fatty hydrocarbon chain is polyunsaturated.

51. The composition of any one of claims 35-50, wherein said nucleic acid molecule comprises a ribonucleic acid (RNA); or wherein said nucleic acid molecule comprises a messenger RNA (mRNA), a self-amplifying RNA (saRNA), a small interfering RNA (siRNA), a transfer RNA (tRNA), a small activating RNA (RNA), an endless / circular RNA (eRNA), a complementary DNA (cDNA), a plasmid DNA (pDNA), or any combination thereof.

52. A method, comprising: a. providing a cell with said composition of any one of claims 33-51 ; and b. delivering said composition into said cell.

53. The method of any one of claims 1-32 and 52, wherein said providing in (a) comprises contacting said composition with an organism comprising said cell.

54. The method of claim 53, wherein said organism comprises an animal.

55. The method of claim 54, wherein said animal comprises a human.

56. The method of claim 54, wherein said animal comprises a non-human animal.

57. The method of claim 56, wherein said non-human animal comprises a rodent.

58. The method of any one of claims 1-32 and 52, wherein said providing in (a) comprises contacting said cell that is cultured in a culture medium.

59. The method of claim 58, further comprising converting said cell or a progeny thereof into a meat product.

60. The method of claim 58, further comprising generating a therapeutic product using said cell or a progeny thereof.

61. A composition for use in a method of treating a disease, said composition comprising (i) a nucleic acid molecule, wherein said nucleic acid molecule is a therapeutic nucleic acid molecule, and (ii) a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2).

62. The composition for use of claim 61, further comprising (iii) a cationic lipid molecule.

63. The composition for use of claim 61, wherein said cationic lipid molecule is selected from the group consisting of DOTAP and any compound of Formula (I)-(IX).

64. The composition for use of any of claims 61-63, further comprising a structural lipid.

65. The composition for use of claim 64, wherein said structural lipid comprises DOPE.

66. The method of any one of claims 5-32 and 52-65, wherein said composition further comprises a second lipid molecule different from said lipid molecule.

67. The method of claim 66, wherein said second lipid molecule is a structural lipid molecule.

68. The method of claim 66 or claim 67, wherein said second lipid molecule comprises a 1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE); optionally wherein a ratio of said nucleic acid molecule and said second lipid molecule is at least about 0.1, at least about 0.5, at least about 1, at least about 2, at least about 4, at least about 6, at least about 8, at least about 10, or at least about 20.

69. The method of any one of claims 1-32 and 52-68, further comprising, generating a nanoparticle molecule comprising said nucleic acid molecule, said saccharide molecule, said lipid molecule, and said second lipid molecule.

70. The method of any one of claims 1-32 and 52-69, wherein a ratio of said nucleic acid molecule and said saccharide molecule, by weight, is at least about 0.1, at least about 0.5, at least about 1, at least about 2, at least about 4, at least about 6, at least about 8, at least about 10, at least about 12, at least about 14, at least about 16, at least about 20, at least about 30, at least about 40, at least about 50, or at least about 100.

71. The method of any one of claims 5-32 and 52-70, wherein a ratio of said nucleic acid molecule and said lipid molecule is, by weight, is at least about 0.1, at least about 0.5, at least about 1, at least about 2, at least about 4, at least about 6, at least about 8, at least about 10, at least about 12, at least about 14, at least about 16, at least about 20, at least about 30, at least about 40, at least about 50, or at least about 100.

72. A composition comprising: i. a nucleic acid molecule, and ii. a saccharide molecule comprising (1) a sorbitan, a mannide, a trehalose, a functional variant thereof, a derivative thereof, or any combination thereof, (2) a lactose comprising an ester group, or (3) a combination of (1) and (2).

73. The composition of claim 72, wherein said saccharide molecule comprises said sorbitan, said mannide, said trehalose, said functional variant thereof, said derivative thereof, or any combination thereof.

74. The composition of claim 73, wherein said saccharide molecule comprises a second ester group.

75. The composition of any of claims 72, wherein said saccharide molecule comprises said lactose comprising said ester group.

76. The composition of any one of claims 72-75, wherein said composition further comprises a lipid molecule.

77. A composition comprising: a. a nucleic acid molecule, b. a saccharide molecule and / or a lipid molecule; and c. a polar organic solvent miscible with water.

78. The composition of claim 77, wherein said polar organic solvent is at most about 16 %, by volume, of said composition.

79. The composition of claim 78, wherein said polar organic solvent is at most about 8 %, by volume, of said composition.

80. The composition of any one of claims 77-79, wherein said polar organic solvent is DMSO, DMF, ethanol, propanol, 2-propanol, ethylene glycol, a functional variant thereof, a derivative thereof, or any combination thereof.

81. The composition of claim 80, wherein said polar organic solvent is DMSO.

82. The composition of claim 80, wherein said polar organic solvent is ethanol.

83. The composition of any one of claims 77-82, wherein said composition comprises an alcohol-based solvent.

84. The composition of any one of claims 77-83, wherein said saccharide molecule comprises a monosaccharide.

85. The composition of any one of claims 77-83, wherein said saccharide molecule comprises a disaccharide.

86. The composition of any one of claims 77-83, wherein said saccharide molecule comprises an oligosaccharide.

87. The composition of any one of claims 77-83, wherein said saccharide molecule comprises a polysaccharide.

88. The composition of any one of claims 77-87, wherein said composition comprises said saccharide molecule and said lipid molecule.

89. The composition of any one of claims 76-88, wherein said lipid molecule is cationic in an aqueous neutral solution.

90. The composition of any one of claims 76-88, wherein said lipid molecule comprises DOTAP.

91. The composition of any one of claims 76-88, wherein said lipid molecule comprises a lipid molecule having a structure of formula (I), (II), or (III).

92. The composition of any one of claims 76-88, wherein said lipid molecule is selected from the group consisting of a compound of formula (IV)-(IX).

93. The composition of any one of claims 72-92, wherein said saccharide molecule is neutral in an aqueous neutral solution.

94. The composition of any one of claims 72-93, wherein said saccharide molecule comprises sorbitan monooleate (SMO), sorbitan dioleate, sorbitan trioleate (STO), sorbitan monostearate, sorbitan monolaurate, mannide monooleate (MMO), lactose oleate, trehalose monooleate, a derivative thereof, or any combination thereof.

95. The composition of claim 94, wherein the saccharide molecule comprises a sorbitan.

96. The composition of any one of claims 72-95, wherein said saccharide molecule comprises a lipid moiety.

97. The composition of claim 96, wherein said lipid moiety comprises a fatty acyl.

98. The composition of claim 97, wherein said fatty acyl is monounsaturated.

99. The composition of claim 98, wherein said fatty acyl is polyunsaturated.

100. The composition of claim 97, wherein said lipid moiety comprises a fatty hydrocarbon chain.

101. The composition of claim 100, wherein said fatty hydrocarbon chain is saturated.

102. The composition of claim 100, wherein said fatty hydrocarbon chain is monounsaturated.

103. The composition of claim 100, wherein said fatty hydrocarbon chain is polyunsaturated.

104. The composition of any one of claims 72-103, wherein said nucleic acid molecule comprises a ribonucleic acid (RNA); optionally wherein said RNA comprises a messenger RNA (mRNA), a self-amplifying RNA (saRNA), a small interfering RNA (siRNA), a transfer RNA (tRNA), a small activating RNA (RNAa), an endl ess / circular RNA (eRNA), a complementary DNA (cDNA), a plasmid DNA (pDNA), or any combination thereof.

105. The composition of claim 104, wherein the nucleic acid molecule is a mRNA.

106. The composition of any one of claims 72-105, wherein the nucleic acid molecule is a mRNA, and the saccharide comprises a sorbitan.

107. The composition of claim 104, wherein the nucleic acid molecule is a siRNA.

108. The composition for use of any of claims 61-65, wherein the therapeutic nucleic acid molecule is a mRNA.

109. The composition for use of any of claims 61-65, wherein the saccharide comprises a sorbitan.

110. The composition for use of any of claims 61-65, wherein the therapeutic nucleic acid molecule is a mRNA and the saccharide comprises a sorbitan.

111. The composition for use of any of claims 61-65, wherein the therapeutic nucleic acid molecule is a siRNA.