11 results about "Complementary deoxyribonucleic acid" patented technology

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.

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 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 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 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.

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 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 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.