562 results about "CDNA library" patented technology

Nucleic acid from cells and viruses sampled from a variety of environments may purified and expressed utilizing microfluidic techniques. In accordance with one embodiment of the present invention, individual or small groups of cells or viruses may be isolated in microfluidic chambers by dilution, sorting, and/or segmentation. The isolated cells or viruses may be lysed directly in the microfluidic chamber, and the resulting nucleic acid purified by exposure to affinity beads. Subsequent elution of the purified nucleic acid may be followed by ligation and cell transformation, all within the same microfluidic chip. In one specific application, cell isolation, lysis, and nucleic acid purification may be performed utilizing a highly parallelized microfluidic architecture to construct gDNA and cDNA libraries.

The present invention provides methods and compositions for the analysis of gene expression in single cells or in a plurality of single cells. The invention provides methods for preparing a cDNA library from individual cells by releasing mRNA from each single cell to provide a plurality of individual mRNA samples, synthesizing cDNA from the individual mRNA samples, tagging the individual cDNA, pooling the tagged cDNA samples and amplifying the pooled cDNA samples to generate a cDNA library. The invention also provides a cDNA library produced by the methods described herein. The invention farther provides methods for analyzing gene expression in a plurality of cells by preparing a cDNA library as described herein and sequencing the library.

The invention relates to methods of depleting RNA from a nucleic acid sample. The RNA may be any RNA, including, but not limited to, rRNA, tRNA, and mRNA. The method is useful for depleting RNA from a nucleic acid sample obtained from a fixed paraffin-embedded tissue (FPET) sample. The method may also be used to prepare cDNA, in particular, a cDNA library for further analysis or manipulation.

The present invention provides a method and/or means for collecting and analyzing an individual cell in a tissue, and at the same time, quantitatively monitoring the expression levels of various genes while keeping two-dimensional information in the tissue. Specifically, the present invention provides a method comprising preparing a cDNA library from mRNA while keeping two-dimensional cellular distribution information and obtaining the gene expression levels at any site or all sites at a level of single cell. More specifically, the present invention provides a method comprising preparing a cDNA library in a sheet-form from mRNA while keeping two-dimensional cellular distribution information and repeatedly using the cDNA library in the detection of the gene expression, thereby allowing measurement of the expression distribution for a number of genes at a high accuracy.

New biochemical protocols for high throughput processing of mRNA samples into cDNA libraries with adaptor sequences compatible with automated sequencing systems are provided. The provided methods produces cDNA libraries which do not have 3′ bias associated with current cDNA library production methods. New methods for the production of DNA libraries from DNA are also provided.

It is an object to provide a method of suitably analyzing the amount of gene expression of a single-cell.

A method of detecting a nucleic acid comprising

a step of sampling a single-cell from a sample containing at least a single-cell,

a cell lysis step of lysing cell membrane of the sampled single-cell and extracting nucleic acids from the cell,

a DNase treatment step of degrading DNA of the extracted nucleic acids with DNase,

a step of hybridizing mRNA of the total RNA contained in the single-cell with oligo (dT) fixed onto a carrier,

a step of performing reverse transcription of the mRNA hybridized with the oligo (dT) to fix cDNA derived from the single-cell onto the carrier, thereby preparing a single-cell derived cDNA library fixed onto a carrier, and

a step of amplifying cDNA fixed onto the carrier and simultaneously detecting an amplification amount of the cDNA.

Disclosed are methods, compositions and kits related to making double-tagged DNA libraries from RNA/DNA samples. A double-tagged oligonucleotide (DTO) is employed to efficiently add two different tags to ends of DNAs to make a double-tagged DNA libraries. Also disclosed are methods to make mate pair libraries using the double-tagged oligonucleotide, and methods to make double-tagged single stranded DNA. The double-tagged DNA libraries of the invention are ready to be used on next generation sequencing machines.

The present invention relates to a filamentous phage display method wherein the polypeptides of interest displayed on the phage particle are cotranslationally translocated across the cytoplasmic membrane of Gram-negative bacteria based on the signal recognition particle pathway. This method is particularly suitable for polypeptides, which are known to be difficult to display on phages, and for proteins of cDNA libraries and other combinatorial libraries, in particular when derived from very fast folding, stable protein scaffolds. The invention further relates to phage or phagemid vectors useful in the method comprising a gene construct coding for a fusion polypeptide comprising the polypeptide to be displayed on the phage particle and an N-terminal signal sequence promoting cotranslational translocation.

The invention discloses methods of proliferation and differentiation of multipotent neural stem cells. Also provided are methods of making cDNA libraries and methods of screening biological agents which affect proliferation differentiation survival phenotype or function of CNS cells.

The invention provides novel polynucleotides and polypeptides encoded by such polynucleotides and mutants or variants thereof that correspond to a novel human secreted semaphorin-like polypeptide. These polynucleotides comprise nucleic acid sequences isolated from cDNA library from fetal liver-spleen (Hyseq clone identification numbers 5688868 (SEQ ID NO: 1)). Other aspects of the invention include vectors containing processes for producing novel human secreted semaphorin-like polypeptides, and antibodies specific for such polypeptides.

The invention relates to a construction method of a genome knockout library, and more specifically discloses a method used for construction of a CRISPR/Cas9 genome knockout library with an enzymatically digested genome. The method is used for solving problems in the prior art that species gene information is incomplete, artificial design coverage is low, and both time and labor are consumed. The method comprises following steps: 1, construction of a Mspl.f-library library; 2, obtaining of 20bp gRNA target sequence fragments; 3, construction of a PAM-f.library library; and 4, construction of a Lenti-gRNA-library library. The method used for obtaining gRNA libraries is independent of any known species genome sequence information, so that the problem in the prior art that species gene information is incomplete, artificial design coverage is low, and both time and labor are consumed are solved, and the knockout library coverage rate is increased greatly. The method is used for construction of a CRISPR/Cas9 genome knockout library.

Proteins having activity that promotes STAT6 activation, which are used for diagnosing, treating or preventing diseases associated with the excessive activation or inhibition of STAT6 are provided. Using a STAT6 response reporter plasmid, cDNA encoding a protein capable of promoting STAT6 activation was cloned from the cDNA library constructed from human lung fibroblasts, and the DNA sequence and the deduced amino acid sequence are determined. The protein, the DNA encoding the protein, a recombinant vector containing the DNA, and a transformant containing the recombinant vector are useful for screening a substance inhibiting or promoting STAT6 activation.

The invention provides a method for the expression of exogenous DNA libraries in filamentous fungi. The fungi are capable of processing intron-containing eukaryotic genes, and also can carry out post-translational processing steps such as glyclosylation and protein folding. The invention provides for the use of fungi with altered morphology, which permits high-throughput screening and directed molecular evolution of expressed proteins. The same transformed fungi may be used to produce larger quantities of protein for isolation, characterization, and application testing, and may be suitable for commercial production of the protein as well.

The invention provides a DNA library construction method for prenatal diagnosis. According to the DNA library construction method for the prenatal diagnosis, pregnant woman blood plasma is directly used for library construction, and a DNA extraction step is omitted, so that use amount of the pregnant woman blood plasma can be reduced, at the same time, possibility of deviation of fetal DNA content can be reduced, and cost and time for the DNA library construction are greatly reduced. Another aspect of the invention relates to DNA libraries prepared by any of above preparation methods. The third aspect of the invention relates to a DNA sequencing method. The DNA library construction method has good application prospect.

The invention discloses a preparing method of east Asia Tityus antibiotic peptide gene and appliance, which comprises the following steps: restructuring bacillus coli Escherichia coli DH5a/BmKAMP1, CCTCC NO: M207036; constructing east Asia Tityus ioterium cell cDNA library; choosing PCR method; sieving positive colony of scorpion antibiotic peptide gene from ioterium cDNA library; sequence-analyzing coding trait of antibiotic peptide gene; assuring amino acid sequence of the antibiotic peptide gene; adopting chemosynthesis antibiotic peptide; possessing inhibition of diverse density for Gram's bacterium. This antibiotic peptide possesses specificity and high active, which can be used as antibacterial drugs.

Methods for preparing cDNA libraries from single and low quantities of cells are disclosed. The methods are based on the principles of multi-strand displacement amplification or semi-random primed polymerase chain reaction. The methods typically include a step of reverse transcription and subsequent amplification of cDNA. The methods can be adapted for preparation of cDNA libraries that are representative of mRNA or whole RNA expressed by the cell or cells. The cDNA is suitable for sequencing or microarray analysis.