27 results about "Complementary deoxyribonucleic acid" patented technology

The invention belongs to the technical field of microbiological detection, and discloses a composite probe for detecting salmonella enteritidis and a preparation method. The composite probe for detecting the salmonella enteritidis comprises magnetic nanoparticles FeO, wherein the surface of the magnetic nanoparticles FeO is modified with an aptamer apt which is specifically combined with the salmonella enteritidis; the surface of the long afterglow nano particle PLNPs is modified with a DNA (Deoxyribose Nucleic Acid) sequence cDNA (Complementary Deoxyribonucleic Acid) complementary with the aptamer; the aptamer apt and the complementary DNA sequence cDNA are subjected to hybridization to form a composite probe structure FeO-SEapt (at) PLNPs-cDNA (complementary deoxyribonucleic acid). The long afterglow luminescence characteristic of PLNPs (ZnGaO: Cr) is utilized, and an excitation light source is stopped before detection, so that signal acquisition completely avoids autofluorescence of a sample matrix, background fluorescence interference is thoroughly eliminated, and the signal-to-noise ratio is remarkably improved.

The invention provides a glass nanopore probe based on an aptamer and application of the glass nanopore probe in dopamine detection, and belongs to the technical field of electrochemical sensing and scanning probe microscopy. The sensor comprises: a glass nanopore probe, the inner wall of which is modified with a gold layer; a cDNA (complementary deoxyribonucleic acid) single chain is fixed on the gold layer through a gold-sulfur bond; a part of the sequence of the dopamine aptamer is hybridized and combined with the cDNA single chain; wherein the combination of the dopamine and the dopamine aptamer causes the change of an ion current rectification signal of the glass nanopore probe, so that the detection of the dopamine is realized based on the change of the ion current rectification signal of the glass nanopore probe. The invention provides a valuable scheme for developing an aptamer nanopore scanning electrochemical sensor for single cell analysis and researching neurotransmitter-related diseases.

Disclosed herein are methods for processing a cell-free sample of a subject, comprising, providing said cell-free sample of said subject, wherein said cell-free sample comprises a plurality of nucleic acid molecules, subjecting said plurality of nucleic acid molecules to one or more amplification reactions to generate a plurality of complementary deoxyribonucleic acid (cDNA) molecules or derivatives thereof, and sequencing said plurality of cDNA molecules or derivatives thereof. Further disclosed herein are systems, comprising a processor, and a non-transitory computer readable storage medium encoded with a computer program that causes said processor to provide a cell-free sample of a subject, wherein said cell-free sample comprises a plurality of nucleic acid molecules, subject said plurality of nucleic acid molecules to one or more amplification reactions to generate a plurality of complementary deoxyribonucleic acid (cDNA) molecules or derivatives thereof, and sequence said plurality of cDNA molecules or derivatives thereof.

The invention discloses a rapid sterile detection method and kit based on a 16S / 18S rRNA-cDNA digital PCR technology, and the method comprises the following steps: S1, extracting total RNA in a to-be-detected sample to obtain bacteria 16S rRNA and fungi 18S rRNA; s2, carrying out reverse transcription on the bacterium 16S rRNA and the fungus 18S rRNA to obtain a stable cDNA (complementary deoxyribonucleic acid) fragment; and S3, carrying out digital PCR amplification by taking the cDNA fragment as a template, and judging whether bacterial contamination exists or not according to an amplification result so as to realize rapid sterile detection. According to the characteristics of high conservative and high copy of bacteria 16S rRNA / fungi 18S rRNA, 30 sets of universal primer probe combinations are verified, and gt can be covered by selecting the optimal two sets of primer probe combinations; 99.99% of common pollution bacteria / fungi in clinic and pharmacy, and no cross amplification with mitochondrial rRNA of mammals such as human, hamster, cattle and the like.

The invention discloses a kit and a method for preparing cDNA (complementary deoxyribonucleic acid) by reverse recording of a single B cell of a mouse. The kit comprises a B cell lysis solution and a reverse transcription buffer solution system, wherein the B cell lysis solution is prepared from a nonionic detergent, dNTP (deoxyribonucleoside triphosphate), dCTP (deoxycytidine triphosphate), a secondary structure stabilizer, a reducing agent, BioIS-mCH2-RT (reverse transcriptase), BioIS-CK-RT, a mouse RNA (Ribonucleic Acid) enzyme inhibitor and nuclease-free water; the reverse transcription buffer solution system comprises 5X RT Buffer, a secondary structure stabilizer, Mg < 2 + >, a reducing agent, a mouse RNA enzyme inhibitor, a Bio-Adaptor primer, reverse transcriptase and nuclease-free water. The kit can significantly improve the success rate, accuracy and sensitivity of reverse transcription of a single B cell, and simplify the operation process.

The invention discloses a polypeptide asparagine ligase VdiPAL1 and an application of the polypeptide asparagine ligase VdiPAL1. The polypeptide asparagine ligase VdiPAL1 is newly found from viola plants, 12 mg / L zymogen protein is obtained through expression of an escherichia coli prokaryotic system, and compared with homologous enzymes VyPAL2 and VdiPAL1 which are widely applied at present, the polypeptide asparagine ligase VdiPAL1 has the advantages of high expression, high catalytic efficiency and substrate specificity and has good development and application potential. The invention also shows the value of the VdiPAL1 for guiding homologous enzyme modification, and particularly discloses the mutant protein VyOPT of the polypeptide asparagine ligase VyPAL2 and a coding cDNA (complementary deoxyribonucleic acid) sequence of the mutant protein VyOPT. The invention also discloses a mutant VyOPT, which comprises five mutations obtained by modification guided by the VdiPAL1. Compared with the expression quantity of the natural VyPAL2, which is about 0.1 mg / L, the expression quantity of the VyOPT reaches 12 mg / L.

The invention discloses a tobacco cytochrome c1 subunit and application thereof in plant disease resistance. Belongs to the technical field of gene engineering. The cDNA (complementary deoxyribonucleic acid) nucleotide sequences of the tobacco NtCytc1-1a / 1-1b / 1-2a / 1-2b / 1-2c / 1-2d gene are as shown in SEQ ID NO: 1-SEQ ID NO: 6. The function loss of the protein coded by the tobacco NtCytc1-1a / 1-1b / 1-2a / 1-2b / 1-2c / 1-2d gene leads to mROS outbreak, disease resistance enhancement and GmMEKK1 response enhancement of the plant. The method has a guiding effect on cultivation of new disease-resistant varieties of crops.

The invention relates to the field of plant molecular biology, in particular to application of a CsERF113L gene or a biological material containing the CsERF113L gene in regulation and control of stress resistance and yield of cucumbers. The cDNA (complementary deoxyribonucleic acid) sequence of the CsERF113L gene is as shown in SEQ ID NO. 1. The CsERF113L gene is knocked out in cucumber by adopting a CRISPR-Cas9 gene editing technology, the antioxidant capacity and photosynthetic capacity of cucumber plants are remarkably enhanced, the saline-alkaline resistance is remarkably improved, the root growth is accelerated, the cucumber yield is remarkably increased, and knockout strains and over-expression strains have opposite trends. The result shows that the CsERF113L gene affects the regulation and control mechanism of cucumber stress resistance. The CsERF113L gene has important guiding significance and application value for guiding widening of cucumber cultivation conditions in actual production, improvement of cucumber high yield and stress resistance and cucumber seed selection and breeding.

The invention belongs to the technical field of insecticides, and particularly relates to a ds-ABC transporter gene for preventing and controlling resistant Frankliniella occidentalis as well as a screening method and application of the ds-ABC transporter gene. The ABC transport protein gene is selected from any one or more of ABCG20, ABCA1, ABCB8, ABCD2 and ABCA5, and the ABC transport protein gene is selected from any one or more of ABCG20, ABCA1, ABCB8, ABCD2 and ABCA5. The screening method comprises the following steps: (1) testing the resistance level of the Frankliniella occidentalis to spinetoram, imidacloprid and thiamethoxam; (2) extracting and purifying the total RNA of the Frankliniella occidentalis population by using a trace total RNA extraction kit, synthesizing cDNA (complementary deoxyribonucleic acid) by taking the RNA as a template, and then carrying out quantitative primer design and gene expression; and (3) carrying out significant analysis on difference analysis of all data through real-time fluorescent quantitative detection, and screening out 5 ds-ABC transporter genes which are subjected to significant up-regulation expression in a resistant population. According to the invention, the key ABC transporter gene for mediating the resistance of the Frankliniella occidentalis to the spinetoram is screened and determined, and a basis is provided for development of a novel anti-resistance insecticide taking the ABC transporter as a target.

The invention discloses an oligonucleotide pharmacobiological analysis system based on a GalNAc technology, and relates to the technical field of drug analysis, and the oligonucleotide pharmacobiological analysis system is technically characterized in that in a GalNAc conjugate coupling delivery module, a coupling unit covalently couples an oligonucleotide drug and a GalNAc-trimer through a click chemical reaction to form a liver-targeted delivery carrier; the targeting unit is specifically combined with an asialoglycoprotein receptor on the surface of a hepatocyte by utilizing the vector to realize endocytosis; in the RT-qPCR quantitative analysis module, a magnetic separation purification unit adsorbs oligonucleotides by using carboxylated magnetic beads; the reverse transcription unit uses a stem-loop primer to convert oligonucleotide into cDNA (complementary deoxyribonucleic acid) under the action of heat-resistant reverse transcriptase; the fluorescence quantification unit adopts an FAM-labeled TaqMan probe to monitor amplification, and the absolute concentration of the medicine is calculated in combination with a standard curve; according to the invention, the technical problems of poor targeting, serious off-target effect and poor stability are solved, and high-quality drug effect evaluation can be carried out.

The invention relates to the technical field of plant genetic engineering, in particular to a gene down-regulation method based on regulation of precursor mRNA (messenger Ribonucleic Acid) splicing efficiency, which comprises the following steps: firstly, identifying at least one intron 5'terminal splicing site containing a GT sequence in a target gene; aiming at the site, designing a guide RNA (Ribonucleic Acid) sequence capable of targeting the downstream fourth or fifth nucleotide, and verifying the cleavage activity of the guide RNA sequence; constructing a gene editing vector by using the guide RNA sequence which is verified to be effective and a Cas9 nuclease coding gene; introducing the vector into a plant cell, carrying out genetic transformation and regeneration to obtain a transformed plant, and screening out a mutant which is subjected to site-directed editing at a target site through DNA (Deoxyribose Nucleic Acid) sequencing; and finally, extracting RNA (Ribonucleic Acid) of the mutant, reversely transcribing into cDNA (Complementary Deoxyribonucleic Acid), and analyzing transcript change through PCR (Polymerase Chain Reaction) amplification and gel electrophoresis. According to the invention, stable, heritable and accurate knock-down of gene expression is realized, and a new tool is provided for essential gene function research and crop character improvement.

The invention discloses a cotton gene GhFBH1 and application thereof in identifying the height of a first fruit branch of cotton, and belongs to the field of biotechnology application. The cDNA (complementary deoxyribonucleic acid) sequence of the gene GhFBH1 associated with the first fruit branch height character of the cotton is as shown in SEQ ID NO. 1, and the genome sequence is as shown in SEQ ID NO. 2. The GhFBH1 is regulated and controlled by an SNP site on the gene, and after the SNP site is mutated, the height of a first fruit branch is obviously lower than that of a wild type. The gene has important research value and application prospect in efficient identification of cotton varieties with low first fruit branch height and improvement and breeding of excellent cotton varieties.

The invention discloses an identification method and application of a larimichthys crocea immortalized cell line. The identification method comprises the following steps: 1, providing a larimichthys crocea cell sample; step 2, screening the larimichthys crocea cell immortalized genes, namely extracting total RNA (Ribonucleic Acid) of each cell sample, reversely transcribing the total RNA into cDNA (Complementary Deoxyribonucleic Acid), measuring the expression quantity of immortalized related genes, and determining the genes with the expression quantity conforming to immortalized index characteristics, namely the larimichthys crocea cell immortalized genes; and step 3, based on the larimichthys crocea cell immortalized gene, identifying whether the to-be-identified larimichthys crocea cell is an immortalized cell. The method for identifying the immortalized cells of the pseudosciaena crocea greatly shortens the traditional time dependent on long-term passage, avoids the risk of noise pollution in long-term culture, improves the probability of successful identification, and has the characteristics of low cost and high efficiency.

The invention discloses a gibberellic disease resistance related receptor protein kinase gene as well as an expression vector and application thereof, and belongs to the field of gene engineering. The cDNA (complementary deoxyribonucleic acid) sequence of the TaRLK3-3B is SEQ ID NO. 1, and the amino acid sequence coded by the TaRLK3-3B is SEQ ID NO. 2. The gene is inserted into an expression vector pAHC25 to obtain an overexpression vector of the gene, and the overexpression vector of the gene is introduced into an infected wheat variety, so that the resistance of the wheat variety infected with the gibberellic disease to the gibberellic disease can be remarkably improved. The TaRLK3-3B is used for genetic engineering breeding, and when the TaRLK3-3B is introduced into wheat varieties susceptible to gibberellic disease, the gibberellic disease resistance of wheat can be improved.

The invention discloses a gene for increasing sorghum yield and application thereof, and belongs to the field of plant genetic engineering. The cDNA (complementary deoxyribonucleic acid) sequence of the SbWRKY50 (Sobic.003G037400.1) is equal to '1', and the amino acid sequence coded by the SbWRKY50 (Sobic.003G037400.1) is equal to '2'. The gene is from sorghum, and overexpression of SbWRKY50 in sorghum can delay leaf senescence and increase the yield of sorghum. The gene leaf is knocked out through a CRISPR / Cas9 technology to generate a phenotype which is aged in advance, so that the yield of sorghum is reduced. Therefore, the SbWRKY50 is a gene for increasing the yield of sorghum and is expected to be used for genetic engineering breeding.

The invention discloses a gene associated with cotton fiber length and application, and belongs to the field of biotechnology application. The cDNA (complementary deoxyribonucleic acid) sequence of the gene GhDTX48 associated with the cotton fiber length character is shown as SEQ ID NO. 1, and the genome sequence is shown as SEQ ID NO. 2. The gene GhDTX48 is regulated and controlled by an adjacent SNP site (D11: 24631774), the SNP basic group is mutated from T to C, and the length of genotype fiber after mutation is obviously greater than that before mutation. The invention further provides a method for identifying the variety of the long-fiber cotton, and the cotton with the basic group C corresponding to the SNP site is the variety of the long-fiber cotton. Meanwhile, a new variety of cotton long fibers is cultivated through a genetic engineering means, a gene containing a high-yield genotype GhDTX48 (C) is transferred into a cotton variety, or an SNP site in a low-yield genotype GhDTX48 (T) is subjected to site-specific mutagenesis to form (C), so that the new variety of cotton long fibers is cultivated. The gene has important research value and application prospect in efficient identification of long-fiber cotton varieties and improvement and breeding of excellent cotton varieties.

The invention provides a cDNA (complementary deoxyribonucleic acid) clone plasmid of an avian infectious bronchitis virus (IBV) for expressing secretory luciferase as well as a virus strain and a vaccine obtained by rescuing, and belongs to the technical field of biological medicines. The invention provides a construction method of cDNA (complementary deoxyribonucleic acid) clone plasmids of IBV (infectious bursal virus), which is completed by a one-step method based on yeast homologous recombination in a segmented amplification mode. The IBV infectious cDNA clone constructed by the method has good stability, an IBV genome can be transformed on a gene level by directly utilizing in-vitro homologous recombination, and the efficiency of constructing a recombinant virus is improved. According to the invention, a non-essential gene of the infectious cDNA clone is replaced by a Gluc reporter gene, and the obtained infectious cDNA clone directly transfects mammalian cells to rescue recombinant viruses. The recombinant virus is constructed through a virus rescue method, the growth characteristic of the recombinant virus is basically consistent with that of a parent virus, and genetic stability is kept.

The invention belongs to the field of plant molecular biology and functional genomics, and particularly relates to a method for identifying a barren-resistant index gene of a tea tree based on WGCNA analysis and application. The invention discloses a method for identifying a barren-resistant related gene of a tea tree, which comprises the following steps: firstly, selecting tea tree varieties with different growth vigor, and measuring the root biomass under the same condition; performing low-nitrogen treatment on the seedlings, extracting RNA (Ribonucleic Acid) from root tissues, establishing a cDNA (Complementary Deoxyribonucleic Acid) library, and performing high-throughput sequencing; screening a gene set with high expression and large variable coefficient based on sequencing data, constructing a gene co-expression network by using WGCNA, and identifying a green module highly related to root biomass; 14 core candidate genes which are stably and highly expressed in varieties with strong growth vigor are screened out as barren-resistant potential candidate molecular indexes, are used for germplasm resource evaluation, early screening of low-fertilizer-resistant varieties and molecular marker-assisted breeding, and have important theoretical values and application prospects.

The invention provides a gRNA sequence integrity and accuracy detection method, a template adapter and a kit, a specific adapter sequence is connected to the 3'terminal of a to-be-detected gRNA sequence, and magnetic bead purification is carried out to obtain a gRNA adapter connection product; carrying out reverse transcription on the gRNA adapter connection product, and adding a template conversion adapter to obtain cDNA (complementary deoxyribonucleic acid); carrying out PCR (Polymerase Chain Reaction) amplification and purification on the cDNA by using the primer to obtain a sequencing library; performing high-throughput sequencing on the sequencing library; and performing signal analysis on the sequencing result to determine the integrity and accuracy of the gRNA sequence. According to the high-sensitivity detection method based on high-throughput sequencing, single-base-resolution full-length sequence analysis can be carried out on gRNA, base variation and sequence integrity can be accurately recognized, clinical risks caused by gRNA sequence variation and low purity can be avoided from the source, and the sensitivity of gRNA detection is improved. And key data support can be provided for effectiveness and safety of gene editing drugs.

In an aspect, a method comprises obtaining cell-free ribonucleic acid (RNA) from a plasma sample of a subject; converting the cell-free RNA into complementary deoxyribonucleic acid (cDNA), thereby producing sample cDNA; and sequencing the sample eDNA to determine a level of the sample cDNA that corresponds to a set of RNA transcripts.

The invention discloses a spatial immune repertoire sequencing method based on a DNA cyclization strategy. The method comprises the following steps: (1) preparing an STB chip; (2) preparing a tissue sample; (3) obtaining cDNA (complementary deoxyribonucleic acid) with a space bar code; (4) cDNA amplification and quality control; (5) cyclizing the cDNA; (6) amplifying cDNA V (D) J; (7) library construction; and (8) sequencing and data analysis. The core problems of high-resolution in-situ capture, targeted enrichment and low-cost analysis are solved, and spatial immune repertoire analysis which is high in sensitivity, easy to operate, high in success rate and low in cost is achieved.

The invention discloses an application of a cotton gene GhYMA03G0104 in identification of cotton with a short growth period. A cDNA (complementary deoxyribonucleic acid) sequence of the cotton gene GhYMA03G0104 in a tetraploid YM8 is as shown in SEQ ID NO.1, and a genome sequence of the cotton gene GhYMA03G0104 is as shown in SEQ ID NO.2. The invention further discloses an application of the cotton gene GhYMA03G0104 in identification of cotton with a short growth period. The gene GhYMA03G0104 associated with the growth period is identified through YM8 variety population re-sequencing and whole genome association analysis, and the gene contains a VEFS-Box structural domain. Whole genome association analysis results show that the gene is closely associated with the cotton growth period.

The invention discloses an application of a NaNAC72 gene in increasing nicotine content, and relates to the technical field of plant genetic engineering, cDNA (complementary deoxyribonucleic acid) of gradually narrow-leaf tobacco is used as a template, amplification is performed to obtain a nucleotide coding sequence of the NaNAC72 gene, the nucleotide coding sequence is shown as SEQ ID NO.1, and the sequence of coded amino acid is shown as SEQ ID NO.2. The invention further discloses an application of the NaNAC72 gene in increasing nicotine content. The transcription factor NaNAC72 with the function of negatively regulating and controlling the nicotine content of the tobacco is obtained from the gradually narrow-leaf tobacco, and the nicotine content in the tobacco can be increased without influencing plant growth through gene editing; the NaNAC72 gene provides a target gene for creating a plant material with improved tobacco nicotine content through methods such as gene editing or mutation; the important target gene is provided for directional improvement and cultivation of new high-quality tobacco varieties, and the important application value is achieved.

The invention provides an RNA (Ribonucleic Acid) molecular marker for judging early gonad differentiation of young river snails in China and an identification method. Dmrt1 and Foxl2 genes are important genes related to gonad differentiation of river snails. The method comprises the following steps: designing and verifying primers SEQ ID NO: 1-2 according to conserved regions of Dmrt1 and Foxl2 genes; the method comprises the following steps: collecting gonad tissue samples in a young snail stage and a sexual maturity stage, wherein n is greater than or equal to 120, extracting total RNA (Ribonucleic Acid) of a field snail tissue, carrying out reverse transcription to obtain cDNA (Complementary Deoxyribonucleic Acid), detecting a double-gene expression quantity through qRT-PCR (Quantitative Reverse Transcription-Polymerase Chain Reaction), calculating a standardized expression value by taking beta-actin as an internal reference, and establishing a discrimination function D = log2 (Dmrt1 / Foxl2); 1, judging that the juvenile snails of the field snails start gonad differentiation; according to the invention, the problems of deficiency of specific molecular markers for sex differentiation of the young escargots in China and indefinite gonad differentiation time point are solved in a breakthrough manner, accurate locking of the gonad differentiation time of the young escargots is realized, the gonad differentiation time is 15-20 days earlier than morphological differentiation, and verification of more than one hundred of samples shows that the identification accuracy reaches 100%.

The invention discloses a marker combination, a detection reagent and a kit for tuberculosis diagnosis and application of the marker combination, the detection reagent and the kit, and belongs to the technical field of molecular medicine and infectious disease diagnosis. The marker combination comprises a gene signed by a PO5 gene, mRNA (messenger ribonucleic acid), cDNA (complementary deoxyribonucleic acid) or protein; the PO5 gene signature comprises a TGM2 (Transforming Growth M2), a RARRES3 (Radar Regulation Regulation S3), an HLA-DPB1 (Human Leukocyte Antigen-DPB1), a CD74 The marker combination has high efficiency and high specificity in distinguishing tuberculosis patients from healthy control, pneumonia and lung cancer patients, can be used as a marker for tuberculosis diagnosis, can also be used for monitoring the curative effect of tuberculosis treatment, provides a more accurate treatment strategy for clinic, and is helpful for improving the treatment effect.

The invention relates to a next-generation sequencing noise reduction method for simultaneously detecting BCR-ABL1 fusion and ABL1 mutation based on RNA and application, and belongs to the technical field of molecular diagnosis. The noise reduction method comprises the following steps: S1, carrying out reverse transcription on total RNA (Ribonucleic Acid) extracted from a leukemia sample to be detected to obtain a cDNA (Complementary Deoxyribonucleic Acid) product; s2, carrying out multiple PCR (Polymerase Chain Reaction) amplification to obtain DNA (Deoxyribonucleic Acid) subjected to PCR amplification; s3, performing linker connection, library construction and next-generation sequencing to obtain DNA library total quantity and sequencing data; and S4, carrying out raw signal filtering analysis, filtering false positive sites in a non-target region and background signals caused by reverse transcription, and synchronously judging a BCR-ABL1 fusion type, an ABL1 kinase region and point mutation and insertion deletion of a BCR gene. The noise reduction method can effectively reduce the interference of the sequencing background, not only expands the detection coverage range, but also has the technical advantages of simplicity and convenience in operation, excellent detection performance, low cost and high throughput.

The invention provides a rapid sequence alignment and annotation method for siRNA off-target analysis, and solves the problem of combinatorial explosion caused by generation of mismatched variation sequences in a traditional method. Comprising the following steps: carrying out sliding window interception on a cDNA (complementary deoxyribonucleic acid) sequence of a target species, and carrying out base sliding interception on a fragment according to the length L of siRNA (small interfering ribonucleic acid) and the step length 1; mapping basic groups in each fragment and a small interfering deoxyribonucleic acid siRNA sequence into 2-bit binary codes, and converting the 2-bit binary codes into 64-bit integers containing 2L effective bits for storage; executing bitwise exclusive or operation of the coding value S of the small interfering deoxyribonucleic acid siRNA and the coding value C of the complementary deoxyribonucleic acid cDNA window; performing logic or operation on every two adjacent bits of the execution result R, and compressing the execution result R into an L-bit result B; calculating the Hamming weight of the B as a mismatch number; and for the matching fragments meeting a preset mismatching threshold value, associating and outputting gene expression annotation information from the MySQL database through the GeneID. The method has the advantages that combinatorial explosion is avoided, the operation efficiency is improved, and resource allocation is optimized.