185 results about "Amplicon" patented technology

The invention discloses a 26 Multi-InDel genetic marker system for highly degraded sample typing, a detection primer and application of the 26 Multi-InDel genetic marker system, and belongs to the technical field of forensic medicine identification. According to the invention, through bioinformatics screening and experimental verification, a multi-InDel genetic marker system of which the lengths of 26 amplicons do not exceed 125 bp is constructed. The problem that degradation detection material detection and high system efficiency are difficult to consider at the same time is solved. The random matching probability of the system in Han population in Hunan reaches 2.25 * 10 <-17 >, the cumulative paternity exclusion rate is 0.999925, complete typing of highly degraded DNA can be achieved, triad paternity test can be completed, and a new technical scheme which is efficient and compatible with a capillary electrophoresis platform is provided for forensic practice.

The present invention relates to a method for the ex vivo molecular diagnosis of gastrointestinal stromal tumors, comprising the steps of: providing at least one isolated tissue sample comprising at least one nucleic acid and the subsequent extraction of the at least one nucleic acid. The method according to the present invention further comprises a step of quantifying the at least one previously isolated nucleic acid. Furthermore, the method according to the present invention comprises preparing at least one amplicon library of said previously isolated nucleic acid, which comprises sequences obtained from the FGFR1, NF1, SDHA, SDHB, SDHC and SDHD genes. This step is followed by the step of purifying the at least one amplicon library and the step of simultaneously sequencing the amplicons themselves. The present invention also relates to a multi-gene panel for ex vivo molecular diagnosis of gastrointestinal stromal tumors and a kit comprising such a panel.

The invention provides methods for simultaneously amplifying multiple nucleic acid regions of interest in one reaction volume as well as methods for selecting a library of primers for use in such amplification methods. The invention also provides library of primers with desirable characteristics, such as minimal formation of amplified primer dimers or other non-target amplicons.

The invention provides methods for simultaneously amplifying multiple nucleic acid regions of interest in one reaction volume as well as methods for selecting a library of primers for use in such amplification methods. The invention also provides library of primers with desirable characteristics, such as minimal formation of amplified primer dimers or other non-target amplicons.

The application provides a STR typing method applied to second-generation sequencing data, and relates to the technical field of biology.The method specifically comprises the following steps: customizing an STR locus configuration file; constructing a k-mer index library of a flanking sequence of the STR locus; extracting STR allele sequences from each READ of a sequence alignment file in combination with a CIGAR value and the k-mer index; constructing a PCR ghost peak model of the STR locus, and estimating parameters of the PCR ghost peak model by using a maximum likelihood estimation method; estimating candidate allele frequencies by using a second-generation sequencing sample data set; calculating posterior probabilities of all candidate alleles by using an allele set extracted from a given sequence alignment file, and inferring the most possible allele.The STR typing method has higher detection rate and typing accuracy, and has faster typing speed, and can be used for whole genome sequencing data, high-coverage targeted sequencing data or amplicon sequencing data.

Synthetic Southern Phaseoloviridae satellite RNA molecules and satellite particles containing the same are disclosed. Synthetic Southern Phaseoloviridae satellite RNA molecules containing an internal heterologous RNA virus (HRV) amplicon are also disclosed. Also disclosed are methods of using Southern Phaseoloviridae satellite RNA molecules and satellite particles containing the same to alter plant phenotypes, improve stress resistance in plants, and improve pest and pathogen resistance in plants.

The invention discloses a megalurothrips megalurothrips methylamino abamectin benzoate resistance molecule detection method which comprises the following steps: (1) collecting megalurothrips megalurothrips female adults in a plurality of regions, and extracting DNA (deoxyribonucleic acid) in a single region in a mixed manner; (2) designing glutamic acid gated chloride ion channel gene specific primers of the megalurothrips megalurothrips, respectively adding specific tags at the front ends of the primers in all regions, and carrying out PCR (Polymerase Chain Reaction) amplification; mixing the PCR products of each region to obtain a library; (3) performing second-generation amplicon sequencing on the library to obtain off-machine data; and (4) analyzing offline data, merging the data in the same region, calculating the mutation frequency of each region by taking the ratio of the mutation (G232S) of the target sequence as an index, and judging the resistance of the megalurothrips usitatus to the emamectin benzoate. The method can be used for rapidly and massively monitoring the emamectin benzoate resistance condition of the field megalurothrips megalurothrips, and an important basis is provided for high-level emamectin benzoate resistance early warning and megalurothrips megalurothrips chemical treatment.

The invention relates to a full-length sequence amplification primer, an amplification method and a three-generation sequencing method of an HLA-I antigen gene, and belongs to the field of gene detection. The HLA-I antigen gene is subjected to PCR (Polymerase Chain Reaction) amplification by using 8 pairs of characteristic amplification primers. And each gene can be subjected to high-yield specific amplification on a full-length sequence containing complete intron and exon regions only through one round of amplification and enrichment, and 7.8 Kb long-fragment genes can be captured once at most. The experimental deviation caused by simultaneous amplification of a single gene by using a plurality of pairs of segmented primers and the indirect error caused by gene splicing are effectively avoided. The method comprises the following steps: carrying out quality inspection and purification on amplicon PCR (Polymerase Chain Reaction) products, mixing samples, constructing an HLA-I type antigen gene library, carrying out accurate and complete sequencing on the full-length sequence of the HLA-I type antigen gene by adopting a three-generation PacBio Sequel II platform, and ensuring that the Hifi reads base accuracy can reach 99% or above. The reading of HLA-I type antigen gene full-length sequence variation information and the haplotype analysis at a high resolution level can be realized.

Disclosed are synthetic Marivirus eye satellite RNA molecules and satellite particles containing the same. Also disclosed are synthetic mateur virus satellite RNA molecules containing an internal heterologous RNA virus (HRV) amplicon. Also disclosed are methods of altering plant phenotypes, improving stress resistance in plants, and improving pest and pathogen resistance in plants using the Marivirus eye satellite RNA molecules and satellite particles containing the same.

The present disclosure generally relates to cartridge-based methods and devices for preparing a sequencing nucleic acid library from a sample. In embodiments, a cartridge-based method comprises placing a sample in a self-contained cartridge having a plurality of chambers, a reaction vessel, and a filter disposed in a fluidic path between the plurality of chambers and the reaction vessel, utilizing the filter to isolate the nucleic acid from the sample; forming a library prep reaction mixture; causing the library prep reaction mixture to flow into the reaction vessel and preparing the sequencing nucleic acid library comprising the tagged nucleic acid fragments, amplicons, and / or adapter labeled nucleic acid; isolating the tagged nucleic acid fragments, amplicons, and / or adapter labeled nucleic acid from the library prep reaction mixture on to the filter and eluting each from the filter; and purifying and optionally size selecting the sequencing nucleic acid library on the filter.

Methods and systems are provided that include measuring the size of a plurality of nucleic acid molecules using multiplex digital amplification reactions. In various embodiments, a digital amplification reaction uses multiple separate forward and reverse primer pairs to amplify targeted regions of the nucleic acid molecule, where the targeted regions are separated by a specified number of base pairs. In other embodiments, the digital amplification reaction uses a plurality of primer pairs that share a universal pair to amplify target regions that overlap each other. The methods and systems are particularly useful for determining the size distribution of nucleic acid molecules, or classifying the pathology of a subject from which nucleic acid molecules are sampled. Advantageously, the methods and systems can measure the size of long nucleic acid molecules without relying on inefficient amplification of very long amplicons.

An in vitro approach for detecting Enterovirus-D68 (EV-D68) is disclosed. In one aspect, two or more EV-D68 genomic regions are amplified with primers that each include a 5' universal tail and an EV-D68-specific sequence (SEQ ID NOs: 47-92). The tails enable indexing and sequencing of tiled amplicons to confirm EV-D68 presence and permit lineage or subclade assignment. In another aspect, a type-specific RT-qPCR assay employs a forward primer (SEQ ID NO: 95), a reverse primer (SEQ ID NO: 96), and a fluorescent probe (SEQ ID NO:97) for rapid detection. Primer sets and kits comprising the primers, probe, and optional reagents are provided for environmental, clinical, or research testing performed outside the body, including wastewater-based surveillance. The universal tails are selected to avoid sequence overlap with any target amplicon. The methods, uses, and kits support sensitive EV-D68 detection across circulating variants and facilitate scalable sequencing and public-health monitoring.

The application relates to the field of second-generation sequencing, in particular to an ultrahigh-throughput multiplex PCR amplicon capture method for second-generation sequencing. The NGS capture method comprises the following steps: 1) performing multiplex PCR amplification on a sample to be detected by using a digestible composite modified primer pair, to obtain a PCR product; 2) performing an enzyme cutting reaction on the PCR product by using a second enzyme mixture containing a primer digestion enzyme, a mismatch correction enzyme and a terminal repair enzyme, including removing the excess primer part on the amplification product by using the digestion enzyme, and removing the non-specific amplification product by using the mismatch correction enzyme; 3) performing a ligation reaction by using a full-length adapter containing an index, to directly obtain a sequencing library or obtain a sequencing library after amplification. The method significantly improves the amplification efficiency and specificity, and has the advantages of simple steps, low cost, short time, wide application value in the fields of genetic disease detection, tumor companion diagnosis and genetic detection and scientific research.

The invention discloses a specific amplification primer pair for human intestinal archaea flora 16S rRNA amplicon sequencing and application of the specific amplification primer pair, and belongs to the technical field of biology.The primer pair is designed for 16S rRNA high-throughput sequencing and can specifically amplify human intestinal archaea, the length of an amplification product is about 385bp, and the amplification product covers a V4-V5 variable region of a 16S rRNA gene; the invention also optimizes amplification conditions of the primer pair, and experimental results show that the primer pair is superior to a conventional universal primer for bacteria and archaea in the aspect of amplifying species diversity and species abundance of the intestinal archaea, and a standardized method is provided for efficient and accurate amplification of intestinal archaea florae.

The invention provides methods for simultaneously amplifying multiple nucleic acid regions of interest in one reaction volume as well as methods for selecting a library of primers for use in such amplification methods. The invention also provides library of primers with desirable characteristics, such as minimal formation of amplified primer dimers or other non-target amplicons.

The invention relates to a method, uses, a system, and a kit for detecting a human papillomavirus (HPV) genotype associated with the development of cervical, oral, or anogenital cancer in a subject and for determining the HPV genotype or genotypes thus detected in the same. Additionally, the present invention relates to uses of direct and / or reverse amplicons or primers generated by multiplex real-time PCR comprising a molecular tag for identifying a marker of human papillomavirus genotypes 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, and / or 68.

Synthetic Tombusviridae satellite RNA molecules and satellite particles containing the same are disclosed. Also disclosed are synthetic Tombusviridae satellite RNA molecules containing internal heterologous RNA virus (HRV) amplicons. Methods of using the Tombusviridae satellite RNA molecules and satellite particles containing the same to alter plant phenotype, improve stress resistance of plants, and improve pest and pathogen resistance of plants are also disclosed.

Disclosed herein includes a method of determining the RhD genotype of a fetus, comprising: (a) extracting cell-free DNA from a sample of a pregnant person, wherein the extracted DNA comprises a mixture of maternal cell-free DNA and fetal cell-free DNA; (b) performing targeted multiplex amplification on the extracted DNA to amplify a plurality of target loci together in the same reaction mixture using a plurality of primer pairs, wherein at least three of the primer pairs each targets a different exon or intron of the RhD gene and the RhCE homologue gene and is each designed to amplify a target locus that comprises a variant between the RhD gene and the RhCE homologue gene; and (c) sequencing the amplicons by high-throughput sequencing and determine the RhD genotype of the fetus using the sequencing reads of the amplicons targeting the RhD gene and the RhCE homologue gene.

The application discloses a single-cell mitochondrial whole-genome efficient and specific amplification method. The application provides a single-cell or multi-cell mitochondrial genome DNA high-fidelity amplification method, which comprises the following steps: taking total DNA obtained by lysing a test single cell as a template, using a chimeric primer composed of a 5' end non-template matching common sequence and a 3' end mitochondrial DNA (mtDNA) specific matching sequence, and a primer corresponding to the 5' end non-template matching common sequence of the chimeric primer, and using a high-fidelity DNA polymerase with strand displacement activity to simultaneously amplify several overlapping amplicons which completely cover the single-cell or multi-cell mitochondrial genome DNA. The application avoids various problems in the current single-cell mtDNA in-vitro amplification approach, and is a very reliable mtDNA whole-genome amplification method.

The invention belongs to the technical field of microbiological detection, and particularly relates to a Klebsiella pneumoniae MLST typing primer group and method based on a hospital localized microbiological mNGS platform. The primer group comprises primer pairs for amplifying an rpoB gene, an infB gene, a pgi gene, a phoE gene, a gapA gene, an mdh gene and a tonB gene, and corresponding nucleotide sequences are as shown in SEQ ID NO: 1-14. According to the technical characteristics of short sequencing read length (50 bp) and single-ended sequencing of a next-generation sequencing platform in a hospital, the primer group for Klebsiella pneumoniae MLST housekeeping gene amplification is redesigned, and amplicons are adaptive to the sequencing working principle of the existing next-generation sequencing platform in the hospital by optimizing the length and position of the amplicons. By utilizing the primer group, the kit or the method, the genetic relationship between hospital infection pathogenic bacteria can be quickly and accurately judged (within 24 hours), the timeliness and scientificity of infection prevention and control are remarkably improved, and the primer group, the kit or the method have a wide clinical application prospect.

This invention discloses a method for constructing a multi-target amplicon abundance standard for microorganisms and its application, belonging to the fields of molecular biology detection and microbiome analysis. This standard material screens sequences of common human gut microbiota strains, designs and adds universal primers, and obtains 14 DNA fragments of different lengths through gene synthesis, cloning, and purification, which are then mixed according to a preset abundance. It retains natural characteristics such as primer binding sites and GC content, and can systematically correct technical deviations in amplicon sequencing, solving the problem of species abundance distortion. This standard material functions as both a non-homologous internal reference and a homologous external reference, and can be used for laboratory proficiency testing, reagent kit performance evaluation, and cross-platform data calibration, promoting the leap from relative qualitative to absolute quantitative research in microbiome studies and providing metrological support for the standardization and precision of detection results.

An object of the present invention is to provide a design method, a manufacturing method, a design device, a design program, and a recording medium of a primer for amplicon methylation sequence analysis, which can improve a design success rate of the primer while suppressing the formation rate of a primer dimer extremely low.The present invention is a primer design method for amplicon methylation sequence analysis, including a primer sequence determination step of selecting one or more primer candidate sequence pairs related to a predetermined target site from one or more primer candidate sequences, calculating a local alignment score between predetermined primer sequences, and adopting and determining a primer candidate sequence pair having a score equal to or less than a predetermined threshold value as a forward primer sequence and a reverse primer sequence for amplifying a region including the predetermined target site.

The present invention provides compositions and methods for novel digital PCR (dPCR) assay designs (droplet digital or other systems) for the detection and quantification of multiple Hepatitis B virus (HBV) gene targets. The compositions and methods can further include a non-extendible blocker oligonucleotide to reduce nonspecific inter-amplicon extension.

The invention provides a method for biomimetic directional construction of crop growth-promoting synthetic bacteria and application, and belongs to the technical field of microbial screening. According to the method, optimized microbial communities are naturally selected based on wild sibling species, a species composition feature table is determined through high-throughput sequencing data analysis, core bacteria are calculated, and core bacteria composition feature tables of amplicons and metagenomes are obtained respectively; and then a hybrid correction strategy of a machine learning algorithm is adopted to obtain a high-quality core bacterium composition feature table of the wild related species of the crops, and growth-promoting synthetic bacteria are designed according to the core bacterium composition feature table, so that candidate microorganisms are directionally screened from the wild related species of the crops in a targeted manner, and the growth-promoting synthetic bacteria of the cultivated species of the crops are constructed. According to the method disclosed by the invention, interaction between crop species specificity and synthetic bacteria is considered, the method has the advantages of high efficiency, accuracy, orientation and the like, and construction of crop synthetic bacteria composition is converted from artificial random combination to bionic oriented construction.

Synthetic turnip yellow mosaic virus satellite RNA molecules and satellite particles containing the same are disclosed. Synthetic turnip yellow mosaic virus satellite RNA molecules containing an internal heterologous RNA virus (HRV) amplicon are also disclosed. Also disclosed are methods of altering plant phenotypes, improving stress resistance in plants, and improving pest and pathogen resistance in plants using turnip yellow mosaic virus satellite RNA molecules and satellite particles containing the same.

Sequencing methods for sequencing populations of nucleic acid molecules in which sequencing reads of amplicons are grouped into families according to the nucleic acid molecule of origin by partitioning, sample indexes and information from the sequencing reads, such as start and end points. The methods described herein provide many advantages, including the identification of sequencing reads deriving from the same nucleic acid in the original sample while minimizing the number of aliquots that are processed.

The application relates to a method, device, equipment and storage medium for simulating amplicons of primers on a genome. The application designs a novel method for simulating amplicons of primers on a genome, focuses on supporting high-degeneracy primers, can accurately cope with complex primer conditions, allows flexible setting of mismatch number parameters, thereby effectively improving the accuracy and reliability of simulated amplification, can significantly improve the running efficiency by means of multi-thread technology, can cope with large-scale genome set processing tasks, is developed based on the Go language, can stably run in mainstream systems such as Linux and Windows, realizes perfect balance between running speed and stability, fully compensates for the deficiencies of the prior art, and provides strong technical support and guarantee for primer simulation amplification work related to targeted sequencing in the field of gene detection.

Multiplex PCR amplification may be improved by using primers with exonuclease-resistant bonds or nucleotides to reduce the formation of primer dimers. Multiplex PCR-based assays may be improved by using a load-balanced set of primers with different proportions of primers in the set to provide balanced amplicon read depth.

The invention provides methods for simultaneously amplifying multiple nucleic acid regions of interest in one reaction volume as well as methods for selecting a library of primers for use in such amplification methods. The invention also provides library of primers with desirable characteristics, such as minimal formation of amplified primer dimers or other non-target amplicons.

The invention discloses application of bitter gourd exosomes in regulating intestinal flora. The method comprises the following steps: establishing an Alzheimer's disease mouse model, injecting the extracted balsam pear exosome into the mouse body in a gavage manner to serve as an experimental group drug, taking normal saline as a model negative control group, detecting the change of the intestinal flora of the AD mouse by applying a 16S amplicon sequencing principle, and detecting the change of the intestinal flora metabolite of the AD mouse by applying an LS-MS technology. An open field experiment, a nesting experiment and a water maze experiment are adopted to verify the treatment effect of the balsam pear exosome on AD mouse intestinal flora and metabolic level changes thereof. Results prove that the bitter gourd exosome extracted by the invention can effectively improve the intestinal flora micro-ecology of AD mice; the compound can be used for preparing a microecological preparation for regulating intestinal flora of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, different types of spinal cerebellar ataxia and the like and diseases such as cerebral apoplexy, cerebral injury, epilepsy, tumor and the like.