18 results about "Degenerate primer" patented technology

This invention relates to a primer set and method for high-throughput assessment of drug resistance levels in rodent populations. The invention provides a set of universal degenerate primers for amplifying the Vkorc1 gene in multiple rodent species. The primer set includes primers for amplifying exon 1, exon 2, and exon 3 of the Vkorc1 gene, respectively. By designing universal degenerate primers and coupling them with next-generation sequencing technology, this invention can simultaneously amplify the Vkorc1 gene coding region of at least 15 rodent species across genera and families. This significantly improves the species universality and detection throughput of rodent drug resistance monitoring, reduces the cost of large-scale screening, overcomes the limitation of Sanger sequencing in accurately analyzing heterozygotes with insertions and deletions, and enhances the ability to discover new resistance mutations. It provides a standardized and low-cost technical solution for efficient assessment of rodent drug resistance.

The present invention discloses degenerate primers for identifying ethyl acetate-producing strains and their applications, belonging to the field of molecular biology. The present invention respectively designed 6 pairs of degenerate primer pairs based on the alcohol acyltransferases encoded by the functional genes ATF1 and EAT1 for ethyl acetate synthesis. By comparing the detection effectiveness, accuracy, and sensitivity of each primer pair, 2 pairs of degenerate primer pairs with significantly excellent effects were screened. Through multiplex PCR technology, it can quickly detect whether yeast and test samples have the potential to produce ethyl acetate, and can determine the gene categories responsible for ethyl acetate synthesis in the test samples.

The invention relates to the field of soy sauce quality detection, in particular to a degenerate primer, a composition, application and a method for rapidly detecting soy sauce aerogenic bacteria, the sequence of the degenerate primer is shown as SEQ ID NO.1-2, the composition further comprises a specific primer and a probe, the sequence of the specific primer and the sequence of the probe are shown as SEQ ID NO.3-6, and the degenerate primer and the composition are used for rapidly detecting the soy sauce aerogenic bacteria. The method comprises the steps of constructing a dual real-time fluorescent quantitative PCR reaction system and performing a dual real-time fluorescent quantitative PCR amplification reaction. The detection method has the advantages of rapidness, simplicity, convenience, high accuracy and high sensitivity.

The invention discloses a kit for rapidly and quantitatively detecting an integrated HIV (Human Immunodeficiency Virus) previrus genome and application of the kit. The kit comprises an anchor primer in an Alu region in an HIV virus, and degenerate primers in gag, nef and LTR regions of an HIV sequence. According to the kit disclosed by the invention, after pre-amplification, the integrated copy number AU-LTR detected by the QPCR is obviously improved compared with AU-gag and AU-nef, and the capture efficiency is higher. Meanwhile, the kit provided by the invention can be used for carrying out three-generation sequencing on a detected sample with a relatively high integration level on the basis of Alu-LTR capture and detection of an integrated copy number, and can be used for carrying out sequence composition and genome position analysis on a storage library of HIV (Human Immunodeficiency Virus).

The embodiment of the invention discloses a primer probe combination for detecting TTV:: RARA fusion genes by a real-time fluorescent PCR method, a kit and application. According to the invention, it is found for the first time that the UTR region of TTV is a necessary element for formation and effective detection of the TTV:: RARA fusion gene, and the UTR sequence has high conservative property among different TTV strains. Based on the key characteristic, a specific detection primer is specifically designed. Wherein the upstream primer TTV:: RARA-F adopts a degenerate primer design strategy, hypoxanthine (I basic group) is introduced, the design can cover TTV strains with different genome sequences in the body of an APL patient, broad-spectrum detection of multiple TTV:: RARA fusion genes is realized, expression products of the fusion genes can be specifically recognized, the advantages of detection efficiency and accuracy are achieved, and the kit has the advantages of high specificity, high sensitivity, high sensitivity and the like. The detection period is effectively shortened and the detection accuracy is improved.

The present disclosure relates to an integrated system for generating optimized primer probe pair design for one or more quantitative polymerase chain reaction (qPCR) assays using a sequence alignment free design approach. The system includes a processor; and a computer-readable medium storing instructions which, when executed by the processor, cause the processor to: receive data including a genomic dataset; generate one or more k-mers from the genomic dataset; cluster the one or more k-mers to identify one or more targeted genomic regions; generate a plurality of primer-probe pair candidates corresponding to the one or more targeted genomic regions, wherein the one or more primer-probe pairs include degenerate primers; and perform one or more in silico operations using the one or more primers, wherein the one or more in silico operations comprise at least one of primer-probe pair optimization, specificity testing, secondary structure analysis, and in silico PCR simulation.

The invention discloses a bacterial cadmium response element screening method, and belongs to the technical field of biology. According to the invention, a strain of Microbacterium oxydans SC-30 is screened, the tolerance to Cd < 2 + > is relatively high, and the minimum inhibitory concentration (MIC) value is 5.72 mmol / L, so that the cadmium resistance of microorganisms is greatly broken through. Degenerate primers of bacterium sources cadR and cadC are further designed to be used for obtaining a Cd < 2 + > response element in a microbacterium oxydans SC-30 strain, then cadR and cadC gene sequences in the strain SC-30 are obtained, the sizes of the cadR and cadC gene sequences are 438 bp and 2097 bp, and the cadR and cadC gene sequences have the highest homology with cadR4 and cadC1 gene sequences in Pseudomonas putida, and the cadR4 and cadC1 gene sequences have the highest homology with the cadR4 and cadC1 gene sequences in Pseudomonas putida, and the cadR4 and cadC1 gene sequences are 54.90% and 55.85% respectively.

The present invention belongs to the field of molecular biology, and specifically discloses a degenerate primer pair for detecting the tet(X) gene and its homologous genes and a method therefor. The degenerate primers are designed based on the multiple sequence alignment of the tet(X) gene and its homologous genes, and have good specificity and sensitivity. By optimizing the PCR parameters, a PCR method for rapidly detecting the tet(X) gene and its homologous genes is established. Repeated experiments have proven that this method is rapid, sensitive, and can detect multiple tet(X) homologous genes with a pair of primers, with low detection cost, short time consumption, and good universality. It provides an efficient and reliable method for the epidemiological investigation of drug-resistant genes.

The invention relates to a method for detecting DNA modification, in particular to a construction method of a DNA library with modified nucleotide, a detection method, a kit set and application. According to the scheme provided by the invention, the necessity of designing degenerate primers is avoided, the design difficulty of multiple primers is greatly reduced, the specificity of multiple PCR amplification is improved, and the formation of primer dimers is reduced.

The invention provides a kelp sex specific molecular marker and application thereof, wherein the nucleotide sequence of a male marker fragment is SEQ ID NO: 1, and the nucleotide sequence of a female marker fragment is SEQ ID NO: 2. A gene sequence fragment obtained by screening is from a conserved homologous gene in a sex determining region of brown algae. The screened sex identification marker primer sequence is from a region with the highest similarity in a homologous gene coding sequence, the sex of the kelp gametophyte can be rapidly identified only through a pair of degenerate primers, the efficiency is high, the accuracy is high, and the interspecific universality is high. The method is suitable for identification of male and female mixed samples, distinguishing of parthenogenesis source and sexual reproduction source kelp sporophytes and sex identification of gametophytes of other species of kelp, and important support is provided for efficient management and research and utilization of a kelp gametophyte germplasm bank.

The invention relates to a primer system for specifically detecting alpha-proteobacteria carotenoid synthesis key gene crtZ and an application method thereof, the primer system is based on a crtZ gene conserved region of alpha-proteobacteria, degenerate primers are designed by adopting a CODEHOP method, and the degenerate primers comprise a forward primer and a reverse primer, the primer system amplifies a target band of about 300bp in alpha-proteobacteria erythrobacterium aurantiacum, the homology of the target band and the known crtZ gene is as high as 94.8%, and no crtZ gene sequence is detected in an amplification product of non-alpha-proteobacteria muricaria aureus, so that the specific recognition capability of the primer system on the alpha-proteobacteria crtZ gene is proved, and the primer system can be used for identifying the crtZ gene of the alpha-proteobacteria. The primer system disclosed by the invention has remarkable advantages in the aspects of specificity, sensitivity and accuracy, provides an efficient and reliable tool for detecting the alpha-proteus crtZ gene, and is suitable for the fields of carotenoid synthesis mechanism research, metabolic engineering optimization, related biological function exploration and the like.

The invention discloses a detection method for distinguishing human genetic materials based on a nanopore sequencing technology and a DNA bar code technology. The detection method is characterized by comprising the following steps: (1) extracting total nucleic acid of a sample to be detected; (2) carrying out PCR (Polymerase Chain Reaction) amplification on a sample to be detected by adopting the degenerate primer; and (3) sequencing the PCR product based on a nanopore sequencing technology, and determining whether the to-be-detected sample contains the human genetic material based on a sequencing result. Primer design and optimization are carried out according to the COI gene of human mtDNA, compared with a general identification primer for mammals, the capture capacity of the human COI gene is improved, meanwhile, a sequencing experiment process suitable for on-site rapid detection is developed, enough data can be obtained by computer sequencing for 10 minutes, and then comparison with a known sequence is carried out.

The invention discloses a respiratory adenovirus nucleic acid detection kit, which comprises: an adenovirus specific primer pair containing a T7 promoter, the nucleotide sequence of which is as shown in SEQ ID NO.1-2; synchronous detection of 14 serotypes of B / C / E subgenus is realized through degenerate primers (SEQ ID NO: 1-2) of a target pIII protein gene conserved region; a double-probe bridging chromatography technology (a CES / LES probe is bridged with RNA and a color development system through a 4-5T joint) is adopted, and the sensitivity reaches 100 copies / mL; a three-line test strip (ADV-T line / internal reference-T line / C line coated specific probe) is matched, and visual interpretation is performed within 5 minutes. The total detection time is 40 min, RNA products are naturally degraded to prevent pollution, special instruments and equipment are not needed, and influenza virus nucleic acid detection can be easily popularized to basic-level and remote rural medical institutions.

The application discloses a primer set for detecting a whole genome of a pig HEV, a kit comprising the primer set and application. The primer set comprises nucleotide sequences as shown in SEQ ID NO. 1-62, the primer set is designed for amplification fragments of 450 bp of a whole gene of the pig HEV and 650 bp of a non-conserved region, is divided into two sets of shingle degenerate primers, comprises five primer groups of A, B, C, D and E, and five amplification primer pools are constructed by using the five primer groups, so that the primer set can cover the complete genome of different strain types of the pig HEV. The application also establishes a method for sequencing the whole genome of the pig hepatitis E virus by using multiplex targeted RT-PCR and nanopore sequencing based on the primer set. The method is rapid, sensitive and specific, can be used for rapidly obtaining the whole genome information of the pig hepatitis E virus in a sample, and provides a reliable technical means for tracing the pig hepatitis E virus and accurately determining the genotype.

This invention provides a sex-specific molecular marker for kelp and its application, wherein the nucleotide sequence of the male marker fragment is SEQ ID NO:1, and the nucleotide sequence of the female marker fragment is SEQ ID NO:2. The gene sequence fragments obtained by this invention are derived from conserved homologous genes in the sex-determining regions of brown algae. The primer sequences for the selected sex identification markers are derived from the regions with the highest similarity in the coding sequences of the homologous genes. This method enables rapid identification of the sex of kelp gametophytes using only a pair of degenerate primers, offering high efficiency, high accuracy, and high interspecies compatibility. This method is applicable to the identification of mixed male and female samples, the differentiation of kelp sporophytes from parthenogenesis and sexual reproduction, and the sex identification of gametophytes in other species of the Laminariales order, providing important support for the efficient management and research utilization of kelp gametophyte germplasm banks.

The present disclosure relates to an integrated system for generating optimized primer probe pair design for one or more quantitative polymerase chain reaction (qPCR) assays using a sequence alignment free design approach. The system includes a processor; and a computer- readable medium storing instructions which, when executed by the processor, cause the processor to: receive data including a genomic dataset; generate one or more k-mers from the genomic dataset; cluster the one or more k-mers to identify one or more targeted genomic regions; generate a plurality of primer-probe pair candidates corresponding to the one or more targeted genomic regions, wherein the one or more primer-probe pairs include degenerate primers; and perform one or more in silico operations using the one or more primers, wherein the one or more in silico operations comprise at least one of primer-probe pair optimization, specificity testing, secondary structure analysis, and in silico PCR simulation.

The invention discloses a primer group for detecting a porcine HEV whole genome, a kit comprising the primer group and application. The primer group comprises nucleotide sequences as shown in SEQ ID NO.1-62, the primer group is designed aiming at amplified fragments of 450bp of a whole gene and 650bp of a non-conserved region of the pig HEV, and is divided into two sets of imbalanobis degenerate primers, the two sets of imbalanobis degenerate primers comprise five primer groups A, B, C, D and E, five amplification primer pools are respectively constructed by the five primer groups, and the primer groups can be ensured to cover complete genomes of different plant types of viruses of the pig HEV. The invention also establishes a method for sequencing the whole genome of the porcine hepatitis E virus by multi-targeting RT-PCR (Reverse Transcription-Polymerase Chain Reaction) and nanopore sequencing on the basis of the primer group. The method is quick, sensitive and specific, can be used for quickly acquiring the whole-genome information of the porcine hepatitis E virus in a sample, and provides a reliable technical means for tracing the source of the porcine hepatitis E virus and accurately judging the genotype.

The present invention provides a nucleic acid combination, a kit, and a method for the simultaneous quantitative detection of HBV cccDNA and HBV rcDNA, relating to the field of molecular biology technology. The nucleic acid combination comprises: a degenerate primer that spans the gap on the negative strand of the HBV rcDNA, the degenerate primer being as shown in SEQ ID No. 1 or SEQ ID No. 2; and a qPCR primer for amplifying HBV DNA, the qPCR primer being used to amplify the HBV PreC / C gene or the S gene. The present invention utilizes the differences in the molecular structure of cccDNA and rcDNA to design primers that increase the copy number of the cccDNA positive strand template in proportion. qPCR is then performed simultaneously with a sample that has not undergone cccDNA pre-amplification to quantify HBV DNA and compare the results to calculate the initial copy number of cccDNA and rcDNA. The present invention can simply and efficiently achieve accurate and efficient quantification of HBV cccDNA and rcDNA simultaneously.