307 results about "Transposase" patented technology

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The present disclosure relates to materials and methods for spatially analyzing nucleic acids that have been fragmented with a transposase enzyme, alone or in combination with other types of analytes.

The invention discloses construction and optimization of a gene editing tool based on CRISPR (clustered regularly interspaced short palindromic repeats)-associated transsposases and application of the gene editing tool in escherichia coli, and belongs to the field of synthetic biology and gene engineering. According to the invention, an efficient gene editing system is established in Escherichia coli, and the burden of a MUSCULAR-CAST system on strains is reduced through element optimization; by eliminating the self-targeting effect, large-fragment (> 10k bp) target gene expression cassettes can be efficiently integrated, and the editing degree of the gene editing system is improved. According to the invention, a MUSCULAR-CAST gene editing system is used for constructing the chassis strains for producing 3-fucosyllactose, L-tryptophan, 2 '-fucosyllactose and neotetraose, so that the chassis strains have equivalent or higher expression yield compared with plasmids, and the growth condition is better or the conversion rate is higher.

The invention discloses a multi-omics analysis method and device, electronic equipment and a storage medium, and relates to the technical field of biological information, and the method comprises the following steps: obtaining a spatial single cell function map set based on multi-omics data, the multiple omics data at least comprise single cell transcriptome data, spatial transcriptome data, single cell chromatin transposase accessibility sequencing ATAC data and whole genome association research GWAS data, and the spatial single cell function map set comprises a plurality of maps used for indicating spatial distribution of different omics data; and establishing a mapping relationship from the spatial transcriptome data to the single cell transcriptome data, wherein the mapping relationship indicates spatial position information of each cell in the single cell transcriptome data. By utilizing the method, the flexibility during multi-omics analysis is improved, and the data integration capability is improved.

Provided herein are transposon / transposase systems for transposing and selecting of transposed cells where the system includes a transposase or polynucleotide encoding a transposase; and an exogenous nucleic acid sequence comprising: a transposon comprising: a 5' inverted terminal repeat (ITR), a first polynucleotide encoding a gene of interest; and a 3' inverted terminal repeat (ITR); and (ii) a polynucleotide encoding an at least first selection marker located outside of the 5' ITR and 3' ITR, where upon introducing (a) and (b) into a cell the transposon is transposed into the cell's genome and the polynucleotide encoding the selection marker is episomal.

Provided herein are methods, compositions, and kits for detecting analytes of interest from biological samples using tagmentation. The method comprises: (a) hybridising a nucleic acid analyte to a capture probe on an array, wherein the capture probe comprises: (i) a spatial barcode and (ii) a capture domain; (b) extending the capture probe using the nucleic acid analyte as a template, thereby generating a barcoded nucleic acid molecule on the array, wherein the barcoded nucleic acid molecule comprises a complement of the sequence of the nucleic acid analyte; and (c) tagmenting the barcoded nucleic acid molecule or a derivative thereof using a transposome comprising a transposase to insert a transposon end sequence into the barcoded nucleic acid molecule or the derivative thereof, thereby generating a barcoded nucleic acid fragment, wherein the barcoded nucleic acid fragment comprises: (i) the barcode, (ii) the transposon end sequence, and (iii) at least a portion of a sequence of the nucleic acid analyte or a complement thereof.

The invention provides a method for obtaining engineered recombinant mammalian cells and application of the engineered recombinant mammalian cells, and relates to the technical field of biology. The method comprises introducing into a mammalian cell a minicircle DNA comprising a transposon and a polynucleotide encoding a transposase, the transposon comprising a fragment encoding an antigen binding domain. The method has the advantages of shortening the preparation period of recombinant mammalian cells, simplifying the process and reducing the cost.

The present invention provides engineered transposase polypeptides useful in preparation of next generation sequencing libraries, as well as compositions, methods of utilizing these engineered polypeptides, and polynucleotides encoding the engineered transposases. The present application further relates to methods for using engineered transposases, e.g., in the preparation of nucleic acid libraries for nucleic acid sequencing. More specifically, the invention pertains to methods that use engineered transposases having improved performance to improve the transposition reactions during the preparation of nucleic acid libraries.

The invention provides a transposase library construction method for reducing linker dimers in a library and a related product, and belongs to the technical field of biology. According to the method, library amplification primers and polymerase are optimized in the transposase library building process, so that the proportion of linker dimers in the library under the condition of low input can be effectively reduced, and the sequencing quality is improved.

The invention discloses a high-flux and rapid chromatin immunoprecipitation sequencing method based on a HiBiT tag and a Tn5 enzyme, and belongs to the technical field of biology. According to the method, the high affinity characteristic of a HiBiT tag system is combined with a Tn5 enzyme in-situ library building technology, firstly, a DNA binding protein recombinant expression strain containing a HiBiT tag is constructed and subjected to induced expression, after formaldehyde crosslinking and cell lysis, an Anti-HiBiT antibody is used for rapid immunoprecipitation, and a Tn5 enzyme library is constructed; tn5 transposase is directly loaded on the magnetic beads combined with the protein and DNA fragment compound to complete fragmentation and linker connection, so that the construction of a next-generation sequencing library is quickly carried out, and the total consumed time from immunoprecipitation to the completion of library construction is less than or equal to 5 hours. Meanwhile, automatic treatment is realized through a micro experiment board (the reaction system is less than or equal to 200 microliters) and a liquid workstation, the experiment cost and the sample dosage are reduced, 48 transcription factors and 48 control samples can be synchronously treated, the efficiency is greatly improved, and an efficient tool is provided for microbial transcription regulation and control research.

The invention provides a library building method for direct sequencing of single-stranded nucleic acid, the library building method comprises providing a to-be-detected sequence fragment, and a preparation method of the to-be-detected sequence fragment comprises cutting a double-stranded molecular sequence formed by complementary pairing of internal bases of the single-stranded nucleic acid by using transposase so as to form the to-be-detected sequence fragment. Compared with an existing library building method of RNA direct sequencing, the library building method has the advantages that a pairing structure is formed by complementary bases of RNA or single-stranded DNA, and a transposase compound can be used for shearing and connecting the pairing structure. This is an attempt which has not been made before, and the library building efficiency is greatly improved through the library building mode.

The disclosure relates to nanoparticles comprising a surface-exposed immune cell binding moiety, a transposable element comprising a gene sequence flanked by inverted terminal repeats (ITRs), a nucleic acid encoding a transposase with specificity for the ITRs, and an mRNA encoding a first chimeric antigen receptor (CAR) or T-cell receptor (TCR). Methods are also described for treating a disease or disorder by administering such nanoparticles to a subject in need thereof.

The invention discloses a method for constructing an engineered bacterium of Clostridium aeruginosa, comprising the following steps: preparing a desired gene in a biosynthetic pathway of a target product, namely a pathway gene, to form a pathway gene cluster; embedding the pathway gene cluster into a transposase recognition site of a mariner transposon to form a pathway gene expression cassette; selecting a constitutive strong promoter commonly used in Clostridium jundal for driving transposase expression, loading the pathway gene expression cassette into a multiple cloning site of a plasmid, and inserting an inducible repressor element upstream of the replicon to obtain a transposition plasmid; transforming the transposition plasmid into Clostridium aeruginosa to obtain an engineered bacterium of Clostridium aeruginosa. The yield of 3-hydroxybutyric acid expressed by the engineered bacterium constructed by the above method is as high as 3g / L.

The invention discloses a recombinant PB transposase mRNA (messenger ribonucleic acid) and PiggyBac vector system and application of the recombinant PB transposase mRNA and PiggyBac vector system. The invention provides a recombinant PB transposase mRNA (messenger ribonucleic acid), the nucleotide sequence of the recombinant PB transposase mRNA is shown as SEQ ID NO: 11, and the 5'end of the recombinant PB transposase mRNA contains a cap structure. The invention provides an mRNA type recombinant PB transposase: PBase-Cap-mRNA, so that the risk that a PB transposase plasmid is inserted into a host cell genome is avoided, and the efficiency of inserting a donor plasmid into the host cell genome is improved. The recombinant PB transposase mRNA is capped, so that the stability of the mRNA is improved, the translation of the mRNA is promoted, and the transposon efficiency is improved. The CHO cells are co-transfected by recombining PB transposase mRNA and donor plasmids, so that the efficiency of transfecting host cells by the donor plasmids is improved, and the construction period of stable cell strains is remarkably shortened.

The invention relates to the technical field of gene engineering, in particular to a transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry based on a Tol2 transposition subsystem and a construction method of the transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry. The preparation method comprises the following steps: S1, constructing a Tol2 transposon expression vector pTol2-ef1a-tjp1a-P2A-mCherry which contains an ef1a promoter, a tjp1a gene, a P2A peptide sequence and an mCherry reporter gene; s2, carrying out in-vitro transcription to prepare Tol2 transposase mRNA; s3, mixing the expression vector with Tol2 transposase mRNA, and microinjecting the mixture into the single-cell stage embryo of the zebra fish; s4, performing fluorescence screening on the F0 generation embryos surviving after injection to obtain the Founder fish with positive transgenosis; s5, after the F0-generation positive fish is bred to be sexually mature, an F1 generation is obtained through mating, transgenic positive individuals with stable inheritance are screened out after identification, and a transgenic zebrafish strain is established. According to the transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry based on the Tol2 transposon system and the construction method of the transgenic zebrafish system pTo12-efla-tjp1a-P2A-mCherry based on the Tol2 transposon system, efficient integration of exogenous genes is achieved by utilizing the Tol2 transposon system, and the transgenic zebrafish line capable of being stably inherited to the F1 generation is successfully obtained.

The invention relates to the technical field of sequencing, in particular to a Tn5 transposase-based next-generation sequencing method and a DNA library rapid construction kit, and particularly provides a Tn5 transposase-based next-generation sequencing DNA library rapid construction kit, a next-generation sequencing method and a next-generation sequencing DNA library construction method, and the kit comprises a reagent for constructing a next-generation sequencing DNA library. The Tn5 transposase is adopted for double-end Index library building sequencing, the library building process can be simplified, high efficiency and accuracy of high-throughput sequencing can be ensured, DNA fragmentation and linker addition are achieved in a single step by means of the unique characteristics of the Tn5 transposase, the sequencing efficiency and data management flexibility are improved by introducing the double-end Index, and the method is suitable for large-scale popularization and application. Each Index sequence ensures high specificity and traceability, meanwhile, the sequencing method can effectively work under the condition of low initial template quantity, and the application range is expanded.

The present invention relates to a composition comprising-a first single-stranded nucleic acid molecule comprising a region complementary to a target and a transposase recognition site,-a second single-stranded nucleic acid molecule comprising a region complementary to the target and a transposase recognition site, and-a third single-stranded molecule comprising a region complementary to the target and a transposase recognition site, the third single chain molecule comprises a sequence encoding a portion of the interleukin 2 [gamma] receptor (IL2-R [gamma]) or encoding a complete IL2-R [gamma] receptor and a transposase binding site.

The invention provides a retrotransposase, which comprises a target DNA binding structural domain containing a zinc finger binding motif, a reverse transcriptase structural domain and an endonuclease structural domain, and can be used for reversely transcribing RNA (Ribonucleic Acid) into DNA. The amino acid sequence of the retrotransposase is shown as any one of SEQ ID No.1 to 6 or SEQ ID No.32 to 43 or SEQ ID No.68 to 71 or has at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98% and 99% identity with the amino acid sequence of any one of SEQ ID No.1 to 6 or SEQ ID No.32 to 43 or SEQ ID No.68 to 71. The invention also relates to a system for modifying DNA. The system comprises the retrotransposase provided by the invention or a nucleic acid encoding the retrotransposase provided by the invention; and a donor RNA or a nucleic acid encoding the donor RNA, the donor RNA comprising: a sequence that binds to the retrotransposase and a heterologous sequence.

This disclosure provides improvised reagents and methods for inserting a transgene into the genome of a living host cell. Sleeping beauty (SB) transposase is used in protein form rather than as a vector. This has been made possible by using rational mutagenesis in a particular region of the SB crystal structure to develop variant SB transposase protein that is more soluble. The changes increase solubility while maintaining transposase activity, thereby adapting the variant transposase for the purpose of promoting recombinant integration of a transgene into a target cell when used in protein form rather than as a polynucleotide vector. The modified transposase is highly soluble in electroporation buffer and thermostable during storage. When introduced into a host cell, it promotes integration of a transgene into the genome of the cell in a dose-dependent manner. It is degraded within 48 hours, thereby rapidly clearing transposase activity from the host cell.

This application relates to the field of molecular biology, and more specifically to isolated transposases and their use. More specifically to nucleic acids and nucleic acid constructs encoding transposases, nucleic acid sets and nucleic acid set constructs, and compositions, recombinant vectors, recombinant host cells and kits comprising transposases. More specifically to methods for introducing exogenous nucleic acid fragments into the genome of host cells, methods for editing the genome of host cells, and methods for obtaining host cells containing exogenous nucleic acid fragments in their genomes. More specifically to the use of transposases, nucleic acids and nucleic acid constructs, nucleic acid sets and nucleic acid set constructs, compositions, recombinant vectors, or recombinant host cells for introducing exogenous nucleic acid fragment genes into the genome of host cells, or for preparing drugs or formulations for gene therapy, cell therapy, genome research, or stem cell induction and post-induction differentiation.

The invention discloses a method for constructing a transgenic zebrafish model of hypoxia response. The method comprises the following steps: firstly assembling an sv40mp promoter after a hypoxia response element HRE sequence and cloning the promoter into a pT2AL200R150G plasmid carrying green fluorescent protein (GFP), then microinjecting the recombinant plasmid and transposase mRNA into zebrafish embryos, and inserting the recombinant sequence into the genome of F0 generation zebrafish by Tol2 transposon technology; raising the F0 generation fish to adult fish, hybridizing with wild-type zebrafish to produce F1 generation, hybridizing the F1 generation with the wild-type fish again to produce single-copy transgenic zebrafish F2 generation, self-pollinating male and female sibling zebrafish of the F2 generation, and screening out transgenic zebrafish Tg (HRE-sv40mp:GFP) of hypoxia response of F3 generation that can be stably inherited. The transgenic zebrafish Tg(HRE‑sv40mp:GFP) constructed in the present invention can be used as a simple and efficient hypoxia response model for biological research in hypoxia-related fields, as well as for screening molecular targeted drugs for HIF‑1α, providing a living animal screening model for the development of anti-tumor drugs.

Single-cell multi-omics by co-encapsulating a single cell with two beads, the first an RNA barcoding bead having barcoded mRNA capture primer oligonucleotides attached on the bead surface; and the second a DNA barcoding bead having two types of oligonucleotides releasably attached to the surface: (1) barcoded adapter oligonucleotides that are complementary to oligonucleotides bound to the transposase that are eventually incorporated into gDNA fragments and (2) polyadenylated barcoded oligonucleotides containing the same barcode sequence as the adapters. In addition, integrated analysis of RNA and protein, including intracellular protein, from individual cells using similar co-encapsulation of a single cell, an RNA barcoding bead, and with / without a specific or non-specific protein binding bead in a microwell, to avoid protein fixation by first lysing the cell to liberate intracellular contents, and then capturing protein either on a solid surface or in solution with barcoded affinity reagents.

The present disclosure relates generally to transposase domains, in particular transposase domains comprising an amino terminal deletion, as well as transposase domains that form obligate heterodimers and transposase domains comprising a DNA targeting domain.

The invention discloses a method and system for synchronously detecting host chromatin openness and in-vivo microbiome based on transposase, and belongs to the technical field of biological sequencing data analysis. According to the method, transposase is used for selectively fragmenting an open chromatin region of a host, and a microbial genome is almost randomly cut, so that synchronous enrichment of host and microbial DNA is realized; after high-throughput sequencing library construction and double-end sequencing, sequencing data is split into host source and non-host source reads through bioinformatics analysis, host chromatin state and microorganism composition are analyzed respectively, and a microorganism-host epigenetic regulation network is constructed; the invention further provides a matched DNA sequencing library and an analysis system, multi-scene research of infectious diseases, intestinal microecology, tumor microenvironment and the like is supported, a public database can be reanalyzed, and potential microbial interaction signals are mined. According to the method, the host-microorganism interaction research efficiency is remarkably improved, and a high-sensitivity and integrated technical scheme is provided for epigenetic regulation mechanism analysis.