69 results about "Multigene expression" patented technology

A multigene expression vehicle (MGEV) consisting essentially of a polynucleotide comprising 2 to 8 domain segments, D, each domain encoding a functional protein, each domain being joined to the next in a linear sequence by a Linker (L) segment encoding a Linker peptide, the D and L segments all being in the same reading frame, and at least one of the domains is not a type two protease inhibitor.

Disclosed are useful constructs and methods for the expression of proteins using primary translation products that are processed within a recombinant host cell. Constructs comprising a single open reading frame (sORF) are described for protein expression including expression of multiple polypeptides. A primary translation product (a pro-protein or a polyprotein) contains polypeptides such as inteins or hedgehog family auto-processing domains, or variants thereof, inserted in frame between multiple protein subunits of interest. The primary product can also contain cleavage sequences such as other proteolytic cleavage or protease recognition sites, or signal peptides which contain recognition sequences for signal peptidases, separating at least two of the multiple protein subunits. The sequences of the inserted auto-processing polypeptides or cleavage sites can be manipulated to enhance the efficiency of expression of the separate multiple protein subunits. Also disclosed are independent aspects of conducting efficient expression, secretion, and / or multimeric assembly of proteins such as immunoglobulins. Where the polyprotein contains immunoglobulin heavy and light chain segments or fragments capable of antigen recognition, in an embodiment a selectable stoichiometric ratio is at least two copies of a light chain segment per heavy chain segment, with the result that the production of properly folded and assembled functional antibody is made. Modified signal peptides, including such from immunoglobulin light chains, are described.

Introducing blocks of foreign genes in a single operon would avoid complications such as position effect and gene silencing inherent in putting one gene at a time into random locations in the nuclear genome. Cloning several genes into a single T-DNA does not avoid the compounded variable expression problem encountered in nuclear transgenic plants. This disclosure shows that a bacterial operon can be expressed in a single integration event as opposed to multiple events requiring several years to accomplish. Expression of multiple genes via a single transformation event opens the possibility of expressing foreign pathways or pharmaceutical proteins involving multiple genes. Expressing the Cry2aA2 operon, including a putative chaperonin to aid in protein folding, in the chloroplast via a single transformation event leads to production of crystalized insecticidal proteins. Expressing the Mer operon via a single transformation event leads to a phytoremediation system.

The invention is an automated multiple-purpose, integrated laboratory system comprising interchangeable modular elements for the construction and measurement of biological assays. The functions of the modular elements may include multiwell platform handling, chemical reagent or cell management, volumetric transfer of liquids for assay construction or for recovery of reaction products for analysis, incubation under controlled environmental conditions, measurement of spectrometric signals originating from the assays, processing and analysis of the resulting spectrometric data, and other functions. The modular elements are arranged around a number of robotic elements that deliver plates to different modular elements, transfer plates to groups of modules served by a different robotic element, or other actions necessary in plate handling. Liquid transfer to and from multiwell platforms, necessary for assay construction or for the initiation of physiological events in cells, is partitioned among different modules specialized for transferring nanoliter or smaller volume quantities of chemical concentrates, or microliter quantities of assay reagents, cells, media and other assay constituents. Applications of this invention include the quantitation and analysis of the expression of multiple genes in cells, measurement of multi-gene expression kinetics, analysis of activation or suppression of multiple signal transduction pathways, screening chemical compounds for modulatory effects on multi-gene expression or on signal transduction pathways or on other biochemical networks of cells, or other analytical biological or biochemical assays.

Methods and constructs for the introduction of multiple genes into plants using a single transformation event are described. Constructs contain a single 5′ promoter operably linked to DNA encoding a modified intein splicing unit. The splicing unit is expressed as a polyprotein and consists of a first protein fused to an intein fused to a second protein. The splicing unit has been engineered to promote excision of all non-essential components in the polyprotein but prevent the ligation reactions normally associated with protein splicing. Additional genetic elements encoding inteins and additional proteins can be fused in frame to the 5′-terminus of the coding region for the second protein to form a construct for expression of more than two proteins. A single 3′ termination sequence, such as a polyadenylation sequence when the construct is to be expressed in eucaryotic cells, follows the last coding sequence. These methods and constructs are particularly useful for creating plants with stacked input traits, illustrated by glyphosate tolerant plants producing BT toxin, and / or value added products, illustrated by the production of polyhydroxyalkanoates in plants.

The invention is an automated multiple-purpose, integrated laboratory system comprising interchangeable modular elements for the construction and measurement of biological assays. The functions of the modular elements may include multiwell platform handling, chemical reagent or cell management, volumetric transfer of liquids for assay construction or for recovery of reaction products for analysis, incubation under controlled environmental conditions, measurement of spectrometric signals originating from the assays, processing and analysis of the resulting spectrometric data, and other functions. The modular elements are arranged around a number of robotic elements that deliver plates to different modular elements, transfer plates to groups of modules served by a different robotic element, or other actions necessary in plate handling. Liquid transfer to and from multiwell platforms, necessary for assay construction or for the initiation of physiological events in cells, is partitioned among different modules specialized for transferring nanoliter or smaller volume quantities of chemical concentrates, or microliter quantities of assay reagents, cells, media and other assay constituents. Applications of this invention include the quantitation and analysis of the expression of multiple genes in cells, measurement of multi-gene expression kinetics, analysis of activation or suppression of multiple signal transduction pathways, screening chemical compounds for modulatory effects on multi-gene expression or on signal transduction pathways or on other biochemical networks of cells, or other analytical biological or biochemical assays.

Disclosed herein is a system and method for providing remotely accessible gene expression image data. The system and method allows for increased accuracy and semi-quantitative or fully quantitative data from images by enabling the remote user to select regions of interest on a compressed image, and then conducting quantitative analysis on original images at a central location. The subject invention relates to, in one embodiment, an BVIGEM (Interactive Multiple Gene Expression Maps) system: which provides internet based software tools for the extraction of functional information from gene expression images and also to act as a repository for gene expression image data.

The present invention relates to polynucleotides for multigene applications comprising a novel functional arrangement, as well as vectors, host cells, and recombinant animals comprising said polynucleotides. In addition, the present invention is directed to methods for generating multigene expression cassettes, methods for producing multiprotein complexes in vitro and in vivo, and methods for producing a vaccine. Furthermore, the present invention encompasses methods for screening protein complex interactions or modifications of proteins, and methods for the in vitro or in vivo screening of candidate compounds capable of protein complex interactions or modifications of proteins or capable of inhibiting protein complex interactions or inhibiting modifications of proteins. Also, the present invention relates to the use of the polynucleotides, vectors, host cells or recombinant animals of the invention for (i) preparing a medicament for gene therapy, for (ii) the recombinant production of multiprotein complexes, (iii) for producing a vaccine, or (iv) for screening compounds of interest. Last but not least, the invention is directed to a kit of parts comprising at least a polynucleotide, a vector, and / or a host cell according to the invention.

Disclosed are useful constructs and methods for the expression of proteins using primary translation products that are processed within a recombinant host cell. Constructs comprising a single open reading frame (sORF) are described for protein expression including expression of multiple polypeptides. A primary translation product (a pro-protein or a polyprotein) contains polypeptides such as inteins or hedgehog family auto-processing domains, or variants thereof, inserted in frame between multiple protein subunits of interest. The primary product can also contain cleavage sequences such as other proteolytic cleavage or protease recognition sites, or signal peptides which contain recognition sequences for signal peptidases, separating at least two of the multiple protein subunits. The sequences of the inserted auto-processing polypeptides or cleavage sites can be manipulated to enhance the efficiency of expression of the separate multiple protein subunits. Also disclosed are independent aspects of conducting efficient expression, secretion, and / or multimeric assembly of proteins such as immunoglobulins. Where the polyprotein contains immunoglobulin heavy and light chain segments or fragments capable of antigen recognition, in an embodiment a selectable stoichiometric ratio is at least two copies of a light chain segment per heavy chain segment, with the result that the production of properly folded and assembled functional antibody is made. Modified signal peptides, including such from immunoglobulin light chains, are described.

Methods and constructs for the introduction of multiple genes into plants using a single transformation event are described. Constructs contain a single 5′ promoter operably linked to DNA encoding a modified intein splicing unit. The splicing unit is expressed as a polyprotein and consists of a first protein fused to an intein fused to a second protein. The splicing unit has been engineered to promote excision of all non-essential components in the polyprotein but prevent the ligation reactions normally associated with protein splicing. Additional genetic elements encoding inteins and additional proteins can be fused in frame to the 5′-terminus of the coding region for the second protein to form a construct for expression of more than two proteins. A single 3′ termination sequence, such as a polyadenylation sequence when the construct is to be expressed in eucaryotic cells, follows the last coding sequence. These methods and constructs are particularly useful for creating plants with stacked input traits, illustrated by glyphosate tolerant plants producing BT toxin, and / or value added products, illustrated by the production of polyhydroxyalkanoates in plants.

The invention discloses a lung adenocarcinoma individuation prognosis evaluation method based on a polygene expression characteristic spectrum. The method comprises the following steps that a lung adenocarcinoma prognosis risk gene list and weight are acquired; a prognosis evaluation model is established by using a tumor tissue transcriptome and survival data of a patient with lung adenocarcinoma;a risk score of the patient is calculated according to a gene expression spectrum of the tumor tissue of the patient with lung adenocarcinoma; the annual survival probability of the patient is calculated according to the risk score of the patient. According to the method, the annual survival probability of the patient with lung adenocarcinoma and the actual annual survival probability are highlyidentical, the linear correlation R2 is equal to 0.994, and the P value is equal to 2.86E-43. It is proved that the method has high prediction accuracy, and the state is highly identical to the actualsurvival state. Meanwhile, for each patient with lung adenocarcinoma, the method can provide a specific survival probability curve of the patient.

The invention relate to 'backward-forward double-promoter plant RNAi double expression vector', its characteristic is contain backward-forward eukaryon expression promoter. The vector realizes the construction process of plant expression vector that can express doubly link RNA through one coupled reaction, which the traditional method needs two or three coupled reaction, saving time and financial resources greatly. The vector can be applied in the study of plant functional genome, virus silence inhibition protein and plant antiviral gene engineering; especially, application in establishing plant functional gene RNA interference base that can silence many gene expression in genome range. Simultaneously, there are adequate single enzyme slice sites in both extremities of every element in the vector expression frame; we can use a suitable element instead of the old element in accordance with experiment request.

The present invention provides compositions and methods useful for treating disorders amenable to therapy via introduction of multigenic expression vectors. More particularly, the invention provides vectors and polynucleotides encoding polypeptides for treatment of cardiac disorders wherein said polypeptides may comprise a cytokine, a chemokine, and / or an angiogenic polypeptide, or functional derivatives thereof. Also provided, as compositions of the invention, are linkers useful for connecting and expressing functional (biologically active) polypeptides from single, multigenic-expression constructs.

The invention discloses a method for constructing a multi-gene expression vector through combination of isocaudarner with a Gateway clone technology. The method comprises steps as follows: (1), an M35S-8GWN vector, an MNOS-8GWN vector and an MOCS-8GWN vector are constructed; (2), target genes are cloned, the genes are assembled to the M35S-8GWN vector, the MNOS-8GWN vector and the MOCS-8GWN vector through Not I and Sbf I respectively, and multiple gene expression cassettes are formed; (3), the multiple gene expression cassettes are connected in series through the isocaudarner AsiS I and Pac I; (4), the multiple gene expression cassettes are connected to the target expression vector through a Gateway LR reaction. The invention further discloses the three entry vectors M35S-8GWN, MNOS-8GWN and MOCS-8GWN used for constructing the multi-gene plant expression vector.

The invention discloses a xylose isomerase producing method and belongs to the technical field of biology. The xylose isomerase producing method includes: firstly, cloning xylose isomerase genes of thermus, streptomyces and escherichia coli; secondly, constructing a pichia pastoris expression vector, a saccharomyces cerevisiae expression vector and a bacillus subtilis expression vector which include three xylose isomerase gene expression cassettes, and transforming the vectors to corresponding host bacteria respectively; thirdly, respectively screening recombinants of over-expression xylose isomerases as engineering bacteria; and finally, fermenting the pichia pastoris engineering bacteria, the saccharomyces cerevisiae engineering bacteria and the bacillus subtilis engineering bacteria to produce a recombinant mixed xylose isomerase. Different from a traditional single-gene-coded xylose isomerase, the recombinant mixed xylose isomerase is wide in suitable reaction temperature and pH (potential of hydrogen) range and suitable for various purposes, and yield of the xylose isomerase expressed by the engineering bacteria including the polygene expression cassettes is obviously higher than that of the xylose isomerase expressed by engineering bacteria including single genes, so that production cost is reduced.

The invention discloses a pancreas cancer personalized prognosis evaluation method based on multi-gene expression characteristic spectrum. The pancreas cancer personalized prognosis evaluation methodbased on multi-gene expression characteristic spectrum includes the following steps: acquiring a pancreas cancer prognosis risk gene list and gene weight; constructing a prognosis evaluation model bymeans of pancreas cancer patient tumour tissue transcriptome and survival data; according to the gene expression spectrum of the pancreas cancer patient tumour tissue, calculating the risk score of the patient; and according to the risk score of the patient, calculating the annual survival probability of the patient. The annual survival probability of the pancreatic cancer patient, obtained through the pancreas cancer personalized prognosis evaluation method based on multi-gene expression characteristic spectrum is highly consistent with the actual annual survival rate (linear correlation R2 =0.980, P value = 9.76E-17). Therefore, it is proved that the pancreas cancer personalized prognosis evaluation method based on multi-gene expression characteristic spectrum has very high prediction accuracy, and highly matches the practical survival state. At the same time, for each tumour patient, the pancreas cancer personalized prognosis evaluation method based on multi-gene expression characteristic spectrum can provide a special survival probability curve for the patient.

The invention discloses a polygenic expression characteristic spectrum based glioblastoma individual prognosis evaluation method. The method comprises the steps of obtaining a prognosis risk gene listand a gene weight of a glioblastoma; building a prognosis evaluation model by utilizing a tumor tissue transcriptome and survival data of a glioblastoma patient; calculating a risk score of the patient based on a gene expression spectrum of tumor tissues of the glioblastoma patient; and calculating the annual survival probability of the patient based on the risk score of the patient. A fact thatannual survival probability of the glioblastoma patient obtained by the method is highly consistent with the actual annual survival rate (the linear correlation R2=0.963, and P value=1.38 E-13) provesthat the method has very high prediction accuracy and is highly consistent with the actual survival state. At the same time, for each tumor patient, the polygenic expression characteristic spectrum based glioblastoma individual prognosis evaluation method can work out the special survival probability curve of the patient.

The invention discloses a method for producing alpha amylase, belonging to the biological technical field, comprising the steps of cloning alpha amylase genes of barley, bacillus licheniformis and aspergillus respectively; constructing Pichia expression vector, saccharomyces cerevisiae expression vector and bacillus subtilis expression vector containing three alpha amylase gene expression frameworks; transforming the vectors to be corresponding host strains; screening high-expression alpha amylase recombinants as engineering bacteria; fermenting Pichia engineering bacterium, saccharomyces cerevisiae engineering bacterium and aspergillus engineering bacterium to produce recombined and mixed alpha amylase. Compared with the traditional alpha amylase coded by single gene, the combined and mixed alpha amylase produced by the method can be suitable for wide reaction temperature and pH range, thus being suitable for various applications; the yield of the alpha amylase expressed by the engineering bacterium containing multi-gene expression framework is notably higher than that of the alpha amylase expressed by engineering bacterium containing single gene, and therefore, the production cost can be reduced.

The invention provides a tool for the easy interpretation of the changes occurring in a cell, being a three dimensional complex and control system, by analyzing a limited number of data obtained by quantifying the intensity of the signals present on spots distributed in a two dimensional surface. These signals intensities are related to the level of genes or gene products present in the cells and after processing and data analysis, they provide an absolute or relative quantification of these genes and gene products present in the analyzed cell or tissue or organisms. The invention also provides a list of cellular functions, which are essential in order to obtain an overview of the modifications occurring in the vital or specific cellular functions under specific biological conditions.

The invention discloses a personalized prognosis evaluation method for gastric cancer on the basis of amultigene expression characteristic spectrum. The method comprises the following steps that: obtaining a gastric cancer prognosis risk gene list and gene weight; utilizing a gastric cancer patient tumor tissue transcriptome and survival data to construct a prognosis evaluation model; according tothe gene expression spectrum of a gastric cancer patient tumor tissue, calculating the risk score of the patient; and according to the risk score of the patient, calculating the annual survival probability of the patient. The annual survival probability, which is calculated with the method of the invention, of the gastric cancer patient has consistent height with a practical annual survival ratio(linear correlation R2 is equal to 0.980, and a P value is equal to 1.62E-17). The method is proved to have high prediction accuracy which is highly matched with the practical survival state. Meanwhile, for each tumor patient, the method disclosed by the invention can give the specific survival probability curve of the patient.

The invention relates to a medium-throughput gene expression analysis method based on a second-generation test platform. The medium-throughput gene expression analysis method comprises steps as follows: corresponding PCR (polymerase chain reaction) primers are divided into two groups according to the relative expression quantity of multiple genes, and the concentration proportion of primers is adjusted in the groups; competitive templates of standard substances and a competitive template of a to-be-tested sample are established, and three rounds of multiple competitive PCRs (polymerase chain reactions) are performed; reaction products of the three rounds of reactions are mixed to serve as a second-generation testing platform library for computer sequencing, data are extracted and analyzed, and the difference of relative expression quantities of multiple genes in different samples can be obtained. On the basis of the high-speed-developed second-generation sequencing platform, a target template and an internal reference template are accurately and rapidly quantified, expression difference analysis of five target genes can be performed on multiple samples simultaneously, the procedure is simple and easy to implement, and the cost is low; when multiple gene expression differences of multiple samples are required to be analyzed, the method can be a high-accuracy, moderate-throughput and low-cost method.

The invention discloses a colon cancer individuation prognosis evaluation method based on a multigene expression characteristic spectrum. The method comprises the following steps: obtaining a colon cancer prognosis risk gene list and gene weight; using the tumor tissue transcriptome and survival data of colon cancer patients to construct a prognosis evaluation model; calculating the risk scores ofthe patients according to the gene expression profile of tumor tissues of the colon cancer patients; and calculating the annual survival probability of the patients according to the risk scores of the patients. The annual survival probability of the colon cancer patients obtained by the method disclosed by the invention is highly coincident with actual annual survival probability (linear dependence R2 being equal to 0.988, and P value being 7.35E-39). It is proved that the method has extremely high prediction accuracy, and the result is highly coincident with actual survival condition. Moreover, for each tumor patient, the specific survival probability curve of the patient can be given out.

The invention discloses a multi-gene expression characteristic spectrum-based individualized prognosis assessment method for clear cell carcinoma of kidney. The method comprises the following steps of: obtaining a clear cell carcinoma of kidney prognosis risk gene list and a gene weight; constructing a prognosis assessment model by utilizing tumor tissue transcriptomes and survival data of a patient with clear cell carcinoma of kidney; calculating a risk score of the patient according to gene expression profile of tumor tissues of the patient with the clear cell carcinoma of kidney; and calculating an annual survival probability of the patient according to the risk score of the patient. According to the method, the annual survival probabilities of patients with the clear cell carcinoma ofkidney are highly consistent with the practical annual survival rates (linear dependence R2=0.999, P=8.21E-74), so that the method has high prediction correctness and is highly identical with practical survival states; and meanwhile, for each tumor patient, the method is capable of giving s peculiar survival probability curve of the patient.

The invention discloses an individual cervical cancer prognosis evaluation method based on a poly-gene expression characteristic spectrum. The method comprises the following steps: acquiring a cervical cancer prognosis risk gene table and a gene weight; establishing a prognosis evaluation model by using cervical cancer patient tumor tissue transcriptome and survival data; according to a gene expression spectrum of tumor tissue of a patient suffering from cervical cancer, calculating a risk score of the patient; according to the risk score of the patient, calculating a survival probability of the patient each year. By adopting the method disclosed by the invention, the survivals probability of the patient suffering from cervical cancer each year is highly accordant with an actual survival rate each year (linear relationship R2=0.988, a P value=5.04E-38). Tests show that the method has very high prediction accuracy, and is highly coincided with an actual survival state. Meanwhile, for each tumor patient, a unique survival probability of the patient can be drawn by using the method.

An efficient plant polygenic expression carrier pBtslm / aroM12 prepared from basic carrier pCAMBIA contains the antibiotic resistance gene NPTII, the glyphosate resistant gene aroAM12 and pest-resistant gene BtSlm. Said expression carrier can be used to transform colibacillus to obtain its colibacillus transformant, or to transform Agrobacterium to obtain its Agrobacterium transformant. Said transformat can be used to transform plant, obtaining the plant resisting glyphosate and pests.

The current invention relates to new tools and methods enabling neuroblastoma patient stratification into prognostic favorable or unfavorable groups. The invention is based on the re-analysis of published gene expression data-sets studying neuroblastoma tumors generating different prognostic gene lists. The overlapping gene lists were subsequently tested for their prognostic power on both the published tumor samples and on an unseen large set of unpublished samples, greatly increasing the statistical power of prognostic analyses. In addition, expression analysis of miRNAs in neuroblastoma tumors with different prognosis was performed. By doing this, the inventors could establish a neuroblastoma prognostic classifier with highly improved prognostic power, which is independent from the tumor sample set used to establish it. This classifier and its related prognostic tools and methods are thus perfectly suitable for routine clinical assessment of neuroblastoma prognosis.

A multi-gene carrier system is composed of a receptor carrier and at least two donor carriers, and is prepared by altermative gene assemblings with the said carriers. It can be used for conversion of multiple genes to obtain different genetically engineered products and multi-gene expression characteristics.