17 results about "CDNA library" patented technology

The invention discloses a sequencing method of an immune repertoire, which comprises the following steps: 1) constructing a cDNA library of a sample, the cDNA in the cDNA library being connected with the position information of the cDNA on the sample; 2) enriching cDNA of TCR and / or BCR genes in the cDNA library in a targeted manner to obtain an immune repertoire of the sample; and (3) sequencing the immune repertoire. According to the method disclosed by the invention, targeted enrichment and high-throughput sequencing of TCR and / or BCR sequences are innovatively realized, and gene expression information and spatial position information of each cell in a space are detected at the same time.

A high-throughput screening system and method for small molecule-protein interactions includes first immobilizing selected small molecules on a solid matrix. Immobilization is preferably performed via photoaffinity labeling, where a linker is used to bind the small molecule to the matrix. cDNA libraries are constructed based on various human tissues. Tissue mRNA is transcribed into cDNA and transferred to a phage genome to construct a phage display library, which is then mixed with small molecules conjugated to microspheres and incubated. The incubated mixture is washed to remove unbound phages, including non-specific and weakly bound phages. Bound phages are amplified and used for the next round of biopanning. The biopanning cycle is repeated, and then PCR is performed on each plaque obtained from the final round of biopanning. The PCR products from each small molecule experiment are barcoded, merged, and then subjected to next-generation sequencing.

The application provides a prostate cancer biochemical recurrence prognosis risk prediction model based on fatty acid metabolism and cancer cell stemness genes and a construction method thereof, wherein the construction method comprises the following steps: S1: data collection: obtaining prostate cancer sample transcriptome data with biochemical recurrence information from a database, and dividing the data into a model training set and a model test set; S2: stemness score analysis; S3: fatty acid metabolism score analysis; S4: based on the analysis results of S2 and S3, identifying a co-expression gene module related to fatty acid metabolism and stemness characteristics in prostate cancer by a co-expression similarity algorithm and a hierarchical clustering algorithm, and obtaining a fatty acid metabolism and stemness-related gene set; S5: constructing a prostate cancer BCR prognosis risk prediction model; S6: constructing a nomogram model; S7: extracting RNA of a to-be-tested sample, constructing a cDNA library, quantifying the expression of the above genes, and calculating the prognosis risk level of prostate cancer through the expression level.

Synthetic polynucleotide libraries may include a plurality of polynucleotides. The polynucleotides may comprise DNA and may be configured to hybridize with one or more regions of target nucleic acids. The target nucleic acids may comprise a cDNA library. The cDNA library may comprise at least one exon-exon boundary between a first exon and a second exon.

The application discloses a high-throughput general sample protein interaction screening method. The method comprises the following steps: firstly, constructing a general sample open reading frame library, extracting mRNA of a target sample, and then obtaining a cDNA library by using a modified random primer or an oligo-dT primer; then, uniformly processing the general sample cDNA library; modifying plasmids of a double-hybrid system to obtain modified plasmids; homologously recombining the general sample library and the modified plasmids of the double-hybrid system, and introducing them into organisms to screen and obtain organisms with an interaction library; identifying single-cell PPI pairs of the screened organisms, and constructing a general sample protein interaction network. The application is suitable for screening and identifying the same Barcode cDNA combination pairs by using single-cell sequencing after obtaining tens of thousands of positive PPI combination clones by means of a "library-library" bacterial or yeast double-hybrid system, and realizing PPI network identification in different intra-species, inter-species and hybridization technology systems.

The invention belongs to the technical field of biology, and particularly discloses a TCR / BCR space transcriptome sequencing method. The method is characterized in that a unique space transcriptome chip is adopted, and a capture probe of the chip sequentially comprises a universal sequence 1, a space address code, a unique molecular tag, a universal sequence 2 and a Poly dT sequence; after a cDNA library is constructed by using the chip, cyclizing the cDNA library, so that a spatial address code is physically adjacent to a variable region; and then taking the cyclized DNA as a template, and carrying out multiple PCR amplification by using a primer complementary with the universal sequence 2 and a specific primer aiming at the constant region, so as to enrich a fragment simultaneously containing a complete variable region sequence and a space address code. Through cooperation of chip design and cyclization enrichment, a variable region primer pool is not needed, and the space variable region sequence of the immune receptor can be captured in an unbiased and efficient manner.

The application discloses a method for detecting viruses and virus replication in media based on Small-RNA high-throughput sequencing. The application belongs to the technical field of virus detection, and the method for identifying virus types in a to-be-detected sample by Small-RNA sequencing developed by the application comprises the following steps: 1) extracting total RNA from a virus-infected to-be-detected sample, constructing a cDNA library, and separating small RNA sequence fragments from the cDNA library; 2) downloading virus sequences and classification databases from GenBank, and comparing the small RNA sequence fragments with virus sequence files; 3) comparing a contigs3 file with a virus nucleic acid database, selecting sequences with the minimum e value in the comparison results in the contigs3 file, calculating the coverage of the contigs3 sequences in corresponding virus genomes, distinguishing virus information with a coverage threshold, and deducing the virus source of the small RNA sequence.

The application discloses a method for drawing a human full-coverage protein interaction network based on digital PCR and second-generation sequencing. First, mRNA is obtained from a human 293T cell line, and then a cDNA library is obtained by using a modified random primer or an oligo-dT primer; then, the human cDNA library is uniformly processed; the plasmid of a BACTH bacterial double-hybrid system is modified to obtain a modified plasmid; the sample library and the modified plasmid of the double-hybrid system are homologously recombined and introduced into E. coli to screen positive PPI clones with interaction; and the screened clones are subjected to second-generation sequencing based on digital PCR to draw a human full-coverage protein interaction network. The application is suitable for high-throughput screening after obtaining tens of thousands of positive PPI combination clones by using a "library vs. library" bacterial or yeast double-hybrid system, and is suitable for identifying the same cell barcode cDNA combination to realize PPI network identification in different intra-species, inter-species and hybridization technology systems.

A method for preparing interaction plasmid library and a method for testing interactions between cDNA libraries are provided. The cDNA libraries could be inserted into the first multiple cloning site and the second multiple cloning site of the dual expression vector for testing the interactions between the different cDNA libraries.

The present invention relates to the field of biotechnology and a method for creating a cDNA library. More specifically, the invention refers to a method of forming complementary DNA (cDNA) sequencing libraries from RNA in situ comprising the steps of: (a) fixating cells, immobilized on a solid surface; (b) performing an in situ reverse transcription (RT) inside cells, using RT primers comprising a PCR handle 1, and partially biotinylated dNTPs; (c) releasing single stranded cDNA from the cells using a release mix, wherein the release mix comprises an RNAse, such that the single stranded cDNA is released from intact cells; (d) collecting a supernatant comprising released cDNA into a single larger volume or in separate volumes; and (e) introducing an adapter molecule comprising a PCR handle 2 that will bind 3′ of the extended cDNA.

The invention provides a transcription factor CsNF-YA and application thereof in positive regulation of CsCBCAS gene expression of China-hemp, and belongs to the technical field of gene engineering. The invention provides a transcription factor CsNF-YA, and the nucleotide sequence of the transcription factor CsNF-YA is as shown in SEQ ID No.2. The transcription factor CsNF-YA is screened from a China-hemp yeast cDNA library, the transcription factor CsNF-YA can be specifically combined with a China-hemp CsCBCAS gene promoter, expression of the CsCBCAS gene can be remarkably promoted by overexpression of the transcription factor CsNF-YA, and the transcription factor CsNF-YA can be applied to synthesis genetic improvement of China-hemp CBC.

Disclosed herein are compositions and methods related to the fast and efficient generation of cDNA library from RNA. The method allows integration of the method in an automation adaptable system.

This invention provides a method for targeted enrichment of target sequences in a spatial transcriptome cDNA library. The method includes: using a hybridization capture probe set to hybridize and extend a target nucleic acid sequence or its complementary sequence on the target sequence in the spatial transcriptome cDNA library to obtain an extension product; wherein the hybridization capture probe set includes two or more probes, and any two probes in the hybridization capture probe set do not overlap in the hybridization region of the target sequence; the extension direction of any probe in the hybridization capture probe set is from the target nucleic acid sequence or its complementary sequence to the spatial sequence or its complementary sequence; the extension product includes: the spatial sequence or its complementary sequence, and part or all of the target nucleic acid sequence or its complementary sequence; the extension product is isolated to obtain an enriched library. This method solves the problem in existing technologies that only unbiased sequencing can be performed on spatial transcriptome libraries, and targeted sequencing cannot be achieved, making it applicable to the field of spatial transcriptome library enrichment.

The application discloses a method for drawing a mouse full-coverage protein interaction network by bacterial single-cell transcriptome sequencing. The method comprises the following steps: extracting mRNA of a mouse general tissue, obtaining a cDNA library by using a modified random primer or an oligo-dT primer, uniformly processing a mouse protein library, modifying a plasmid of a BACTH bacterial double-hybrid system to obtain a modified plasmid, homologous recombining the sample library and the modified plasmid of the double-hybrid system, introducing into DHM1 E. coli to screen positive PPI clones with an interaction library, and performing single-cell transcriptome sequencing based on the random primer on the screened clones to construct a mouse protein interaction network. The application is used for high-throughput screening of tens of thousands of positive PPI combination clones obtained by a 'library versus library' bacterial or yeast double-hybrid method, and identification of the same Barcode cDNA combination to realize PPI network identification in different intra-species, inter-species and hybridization technology systems.

The present invention provides a hydrogel bead-based full-length single-cell RNA sequencing method. The method comprises the following steps: performing single cell RT-PCR for cDNA synthesis on hydrogel oligo (dT) beads in droplets, assembling barcode Tn5 transposons on hydrogel beads, co-encapsulating cDNA attached oligo (dT) beads and barcode Tn5 beads in droplets for tagmentation, amplifying the double-stranded cDNAs to obtain a cDNA library, and sequencing the cDNA library.

The present application relates to the technical field of yeast two-hybridization, and in particular, to a method for constructing an interaction plasmid library, a method for identifying interactions using a cDNA library, and use. A yeast double expression vector is used to construct two cDNA libraries in the same vector and express the two cDNA libraries separately. Moreover, the expression of the two cDNA libraries does not interfere with each other, and interaction identification can be accurately performed in the same yeast strain. This reduces the workload associated with constructing and extracting the interaction plasmid library as well as constructing and culturing strains, thereby improving the identification efficiency.

This invention relates to the field of biotechnology, and more particularly to a transcription factor CmMYB44L that positively regulates sucrose content in melon fruits and its applications. This invention uses the CmSWEET10 promoter as bait to screen melon fruit cDNA libraries, and obtains a MYB transcription factor gene CmMYB44L that interacts with it using yeast one-hybrid technology. The amino acid sequence encoded by this gene is shown in SEQ ID NO:1. This transcription factor can directly bind to the CmSWEET10 promoter and activate its expression, thereby significantly increasing sucrose accumulation in melon fruits, confirming that CmMYB44L is a key factor positively regulating sucrose accumulation in melon fruits.