36 results about "Direct sequencing" patented technology

The invention belongs to the technical field of Hu sheep molecular marker-assisted breeding, and particularly relates to application of an AAGAB gene molecular marker g.23443T > C in Hu sheep molecular marker-assisted breeding. According to the invention, Hu sheep is taken as a research object, PCR amplification, direct sequencing of products and sequence analysis are adopted, AAGAB gene polymorphism is analyzed, and correlation between different genotypes of polymorphic sites and different growth traits is comprehensively analyzed. An analysis result shows that g.23443T > C is remarkably related to the chest circumference of Hu sheep and the daily gain of Hu sheep from six-month-old to one-year-old, and the birth weight, weaning weight and six-month-old body weight of CC genotype are remarkably higher than those of TT and TC genotype individuals.

The invention provides a library building method for direct sequencing of single-stranded nucleic acid, the library building method comprises providing a to-be-detected sequence fragment, and a preparation method of the to-be-detected sequence fragment comprises cutting a double-stranded molecular sequence formed by complementary pairing of internal bases of the single-stranded nucleic acid by using transposase so as to form the to-be-detected sequence fragment. Compared with an existing library building method of RNA direct sequencing, the library building method has the advantages that a pairing structure is formed by complementary bases of RNA or single-stranded DNA, and a transposase compound can be used for shearing and connecting the pairing structure. This is an attempt which has not been made before, and the library building efficiency is greatly improved through the library building mode.

Described herein are methods of generating a coupled sequencing read pair for a polynucleotide, and methods of analyzing the coupled sequencing read pair. The coupled sequencing read pair can be analyzed to detect polynucleotide variants, including at loci that are not directly sequenced within the coupled sequencing read pair. Other analytical methods can include using coupled sequencing read pairs to construct or validate a consensus sequence. The coupled sequencing read pair may be generated for a polynucleotide by generating sequencing data for a first region by extending a primer using labeled nucleotides; further extending the primer through a second region using nucleotides provided in a second region flow order, wherein primer extension through the second region is faster than primer extension through the first region; and generating sequencing data associated with a sequence of a third region of the polynucleotide by further extending the primer using labeled nucleotides.

Provided herein are methods of interrogating spatial gene expression in a sample using substrates having sequencing pores. The disclosed methods allow for spatial analysis of analytes from biological samples using a pore-based sequencing approach and without the need for a barcoded spatial array. In some embodiments, an analyte or intermediate agent thereof, is released from a biological sample and directly sequenced by traversing through pores of a sequencing array including a plurality of pores, e.g., nanopores. In some embodiments, light or other stimuli is used to release an analyte or intermediate agent thereof from a specific region of interest in the biological sample followed by sequencing using sequencing array including a plurality of pores.

The invention relates to the technical field of biology, and discloses a linker element and a method for constructing an oxford nanopore direct RNA sequencing library by the linker element, and the linker element comprises a partial double-strand dump structure formed by annealing a first nucleotide strand S and a second nucleotide strand AS; the first nucleotide chain S sequentially comprises a ligase recognition motif region, a sample tag sequence region, a first universal sequence region and a second universal sequence region in the 5 '-3' direction; the second nucleotide AS sequentially comprises a third universal sequence region, a first universal sequence reverse complementary region, a sample tag sequence reverse complementary region, a ligase recognition motif reverse complementary region and a polyadenylate tail recognition region according to the 5 '-3' direction. According to the method, RNA can be efficiently utilized, the high sequencing data utilization rate and high resolution uniformity are achieved, the technical problems that an existing Oxford nanopore direct RNA sequencing technology cannot effectively utilize initial RNA, and a plurality of library samples cannot be effectively split on the same chip are solved, and the economic and time cost of a practical application end is reduced.

The present invention relates to a single nucleotide polymorphism detection primer for the sheep BMPR1B gene and its application, belonging to the field of gene technology. The SNP molecular marker corresponding to the primer of the present invention is located on chromosome 6 of the BMPR1B gene, and the primer is used for sheep marker-assisted selection breeding. The present invention provides a plurality of SNP molecular marker sites related to sheep reproductive traits, and determines the genotype of each site by directly sequencing the PCR product using sheep blood genomic DNA as a template. The present invention can be used as an effective molecular marker to improve sheep reproductive traits. The detection method is simple, rapid, and highly accurate, which can improve the breeding efficiency of sheep. It has high application value and provides more ways for the screening research of sheep reproductive traits.

The invention discloses a target direct sequencing method based on the sequence and epigenetic modification of a full-length RNA (Ribonucleic Acid) molecule of a nanopore, and belongs to the technical field of RNA sequencing. Aiming at the problem that a nanopore cannot directly perform targeted sequencing on RNA, an extension joint is designed to prolong 5'and 3 'ends of an RNA molecule and compensate high sequencing errors of the tail ends, so that a middle RNA sequence is completely sequenced; the extension linker comprises a DNA probe, 5'extension RNA (ER5) and 3 'extension RNA (ER3), the DNA probe can specifically capture RNA molecules and enhance connection with the extension RNA, self-connection of RNA or the extension RNA is reduced to a great extent so as to ensure efficient and accurate RNA extension, target RNA is captured based on sequence complementation, connection deviation caused by different sequences is reduced, and the detection sensitivity is improved. And target sequencing can be carried out on interested RNA types. By constructing a targeted direct sequencing method of specific RNA based on nanopores, the problems that RNA cannot be detected and cannot be accurately detected in direct sequencing based on nanopores are solved, high-throughput and high-precision RNA sequencing is realized, full-length RNA direct sequencing is realized, information such as RNA abundance, sequence, epigenetic modification and the like is obtained at the same time through single sequencing, and the method is suitable for large-scale popularization and application. And a cancer diagnosis model can be constructed in combination with methods such as machine learning and the like, so that high-accuracy and high-specificity diagnosis of cancers is realized.

The invention belongs to the technical field of Hu sheep molecular marker-assisted breeding, and particularly relates to application of an FABP3 gene molecular marker g.2596A > C in Hu sheep molecular marker-assisted breeding. According to the invention, Hu sheep is taken as a research object, PCR amplification, direct sequencing of products and sequence analysis are adopted, FABP3 gene polymorphism is analyzed, and correlation between different genotypes of polymorphic sites and different growth traits is comprehensively analyzed. An analysis result shows that g.2596A > C is obviously related to the birth weight of Hu sheep, and the birth weight of AA genotype is obviously higher than that of CC genotype (P > 0.05).

Methods for generating paired sequencing reads for a polynucleotide, and methods for analyzing paired sequencing reads, are described. Paired sequencing reads can be analyzed to detect polynucleotide variants, including at loci that are not directly sequenced within the paired sequencing reads. Other methods of analysis include using paired sequencing reads to construct or validate a consensus sequence. Paired sequencing reads for a polynucleotide can be generated as follows: by extending a primer with a labeled nucleotide to generate sequencing data for a first region; further extending the primer through a second region using nucleotides provided in a second region flow order, wherein the primer extends through the second region faster than the primer extends through the first region; and generating sequencing data related to the sequence of a third region of the polynucleotide by further extending the primer with a labeled nucleotide.

The present invention belongs to the technical field of molecular marker-assisted breeding of Hu sheep, and specifically relates to the application of the AAGAB gene molecular marker g.23443T>C in molecular marker-assisted breeding of Hu sheep. The present invention uses Hu sheep as the research object, employs PCR amplification, direct product sequencing, and sequence analysis methods to analyze AAGAB gene polymorphisms, and comprehensively analyzes the correlation between different genotypes at the polymorphic sites and different growth traits. The analysis results show that g.23443T>C is significantly correlated with the chest circumference and daily weight gain of Hu sheep from six months of age to one year old. The birth weight, weaning weight, and six-month-old weight of individuals with the CC genotype are significantly higher than those with the TT and TC genotypes.

The invention provides an SNP (Single Nucleotide Polymorphism) molecular marker related to the number of hatches of pigeons, which is mainly characterized in that the nucleotide sequence of the SNP molecular marker is shown as SEQ ID NO.3 or SEQ ID NO.4. The invention also provides an application of the SNP molecular marker related to the number of hatches of pigeons in identification and breeding of the number of hatches of pigeons, a specific primer pair for detecting the SNP molecular marker related to the number of hatches of pigeons, and a method for identifying or breeding of the number of hatches of pigeons. Through the molecular marker and the primer, a rapid, efficient and accurate molecular marker assisted breeding technology can be established by utilizing a PCR technology and a direct sequencing method, early breeding of Shifig pigeons is carried out, the feeding cost is reduced, and the production benefit is improved.

This invention provides a simple, rapid, low-cost, high-throughput, and accurate integrated solution that eliminates the need for nucleic acid extraction and purification. It enables targeted capture, amplification, and first-generation sequencing analysis of all key regions of the ABO gene directly from various clinical samples, including whole blood, saliva, oral swabs, and dried blood spots. The operation is simple and low-cost, thus achieving accurate ABO blood typing, subtype analysis, and the discovery of new variants. The method described in this invention can be applied to the identification of difficult blood types, and can discover new ABO alleles, blood group gene chimeras, and rare blood types such as subtype-related intron mutations.

The application discloses a SNP molecular marker of a site rs338618851 related to pork production efficiency of pigs and application. The genetic marker is cloned from a 93991071 site of a chromosome 6 of pigs. American Landrace pigs are selected as test materials, whole genome DNA is extracted from blood of the pigs, and a primer is designed according to a pig genome sequence NC_010448.4 published in an NCBI database. A base substitution of T or C exists at a 172th position of a primer amplification sequence, the substitution causes polymorphism at the position. The SNP site is typed through a direct sequencing technology, and correlation analysis shows that, compared with other gene individuals, a TT genotype individual at the 172th position is large in birth weight, large in eye muscle area and short in time to reach 100 kg at a certain age. The application provides a new marker for molecular breeding of pork production traits of pigs.

Disclosed are methods of preparing a sequencing library to quantify tRNA abundance and tRNA modifications in an RNA sample that comprises contacting RNA with oligonucleotides in the presence of a ligating agent and performing nanopore direct sequencing. It also discloses a kit to perform said method.

Disclosed is a method of training a complete DNA basecaller using high complex direct RNA sequencing (DRS) training datasets and applications of the trained complete DNA basecaller. Including a method to sequence single-cell RNA using capture oligonucleotides and oligonucleotides-DNA barcodes. The training of the DNA basecaller use a linked DNA-RNA oligonucleotide, in which the DNA part is complementary to the RNA part.

The invention provides a digital micro-fluidic chip and an integrated sequencing system and method.The digital micro-fluidic chip comprises a chip body and at least one nucleic acid extraction and library construction unit, and the nucleic acid extraction and library construction unit is arranged in the chip body and comprises a nucleic acid extraction module and a library construction module which are communicated; the nucleic acid extraction process and the library construction process can be continuously completed on the same digital micro-fluidic chip, manual or external intervention is not needed, and the constructed library can be transferred to a sequencer for direct sequencing. The whole process of nucleic acid extraction, library construction and sequencing can be automated, the reagent dosage can be reduced, the library construction and sequencing cost can be reduced, and the library construction and sequencing time can be shortened. The integrated sequencing system disclosed by the invention can realize smaller volume and electricity consumption, needs less manual operation, has low requirements on use environments (such as temperature, humidity, ventilation and the like), can be used in environments outside a laboratory, and is suitable for various application scenes.

The application discloses a SNP molecular marker related to growth traits of Haimen goats and application thereof, and belongs to the field of biotechnology and livestock breeding. The SNP molecular marker comprises an EGFR g.776097C>T mutation site. The application adopts a method of directly sequencing a PCR product as a template of blood genomic DNA of the Haimen goats to determine the genotype. The CC genotype is significantly related to the body length of the Haimen goats at three months old and the body weight, body length, body height, chest depth and chest circumference traits of the Haimen goats at six months old, and can be used as an effective molecular marker for improving the growth traits of the Haimen goats. The detection method is simple, rapid, accurate and low in price, can improve the breeding efficiency of the Haimen goats, and has high application value.

Disclosed are methods of preparing sequencing libraries to quantify tRNA abundance and tRNA modifications in an RNA sample, the methods comprising contacting the RNA with an oligonucleotide in the presence of a ligation agent, and performing nanopore direct sequencing. Also disclosed are kits for performing the methods.

The invention relates to the technical field of livestock breeding, in particular to a detection method of a goat ASAP1 gene insertion or deletion marker and application of the detection method. Comprising the following operations: taking a whole genome of a goat to be detected as a template, marking the insertion / deletion of the goat ASAP1 gene, designing a primer according to the sequence of a regulation region of the goat ASAP1 gene, taking genome DNA of the Shaanbei white cashmere goat as the template, and detecting the insertion / deletion of the goat ASAP1 gene through PCR amplification, agarose gel electrophoresis and a direct sequencing technology. According to the present invention, the insertion / deletion polymorphism of the goat ASAP1 gene NC030809.1: rs65531471 site, the insertion / deletion polymorphism of the goat ASAP1 gene g.7140795171407952 site and the insertion / deletion polymorphism of the goat ASAP1 gene NC030809.1: rs652252293 site, the insertion / deletion polymorphism of the goat ASAP1 gene g.7144677071446796 site can be accurately and rapidly detected with low cost; the PCR amplification technology is used for detecting the insertion / deletion polymorphism of the goat ASAP1 gene, genotype and allele frequency analysis is carried out, it is found for the first time that the ASAP1 gene is remarkably related to the resistance of the Shaanbei white cashmere goat to the brucellosis, and the ASAP1 gene can serve as an effective DNA marker for goat brucellosis-resistant genetic breeding.

The application belongs to bioengineering technology and particularly relates to a haplotype breeding marker related to pig feed utilization rate and meat production traits and application. The genetic marker is cloned from 19 sites of a SYISL gene promoter of a pig chromosome 8. Three pairs of primers are designed according to a pig genome sequence (NC_010450.4) published by NCBI database. There are 19 base substitutions in the amplified sequences of the three pairs of primers. The SNPs sites are typed by a direct sequencing technology, and the linkage disequilibrium analysis result shows that the 19 sites are all in a complete linkage disequilibrium state, and two haplotypes are generated by analysis. Correlation analysis shows that compared with other haplotype individuals, the H2-H2 combined haplotype individuals have thinner backfat thickness, larger eye muscle thickness, less average feed intake of 30-100 kg body weight and smaller feed-meat ratio. The application provides a new marker for pig meat production trait molecular breeding.

Embodiments of the present application provide a direct RNA sequencing multi-omics analysis method, system, device and storage medium, relating to the technical field of biological information processing. The method comprises: obtaining sequencing data of direct RNA sequencing; aligning the sequencing data with a reference genome to obtain sequencing alignment data; performing full-length transcript identification according to the sequencing alignment data to obtain full-length transcript sequence data; performing transcript quantification processing on the sequencing data based on the full-length transcript sequence data to obtain transcript quantification data; processing the sequencing data according to a methylation modification prediction model to obtain methylation modification data; processing the full-length transcript sequence data according to a nascent mRNA prediction model to obtain nascent mRNA data; and performing correlation analysis according to the full-length transcript sequence data, the transcript quantification data, the methylation modification data and the nascent mRNA data to obtain direct RNA sequencing multi-dimensional information. The method can achieve the technical effect of sequencing accuracy.

The invention discloses a probe set and a kit for specifically enriching RSV whole genome in a respiratory tract sample and application, the probe set provided by the invention can perform high-sensitivity and high-specificity capture and enrichment on RSVA subtype and RSVB subtype free nucleic acid fragments in the respiratory tract sample so as to obtain a high-quality whole genome sequence. Compared with sequencing after purification culture, the method disclosed by the invention can be used for carrying out whole genome sequencing on RSV which is a pathogen difficult to culture, and is wider in application range. Compared with direct sequencing of a positive sample, the method has the advantages that the pathogen detection target is higher, the influence of other sequences is smaller, the interference of impurities can be effectively reduced, the sample pathogen abundance required by high-quality whole genome sequencing can be completed, and the high-quality RSV whole genome sequence acquisition of more samples can be completed.

Disclosed are nucleic acid analysis and sequencing technologies. The disclosure addresses the preparation of transfer RNA (tRNA) constructs suitable for direct RNA sequencing and structural applications by enzymatically ligating multiple tRNA molecules into a single, longer RNA strand using designed RNA oligonucleotides. Disclosed are methods of preparing a sequencing library for a plurality of tRNAs and kits.

Disclosed is a method of pre-processing and inferencing data for RNA chemical modification detection. The method employs direct RNA sequencing on synthetic RNA sequences, various outputs. Basecalled RNA reads are aligned to reference synthetic RNA sequences to obtain sequence alignment reads. Nucleotide assignments of the sequence alignment reads are then refined, data segmented and the segmented data aligned to the individual k-mers. Raw signal features corresponding to specific nucleotide positions in the reference synthetic RNA sequence are then identifiable. The features of the raw signals are subjected to a multiple-instance supervised learning model training framework for the detection of one RNA chemical modification or multiple RNA chemical modifications. The trained model infers from an unseen RNA sequence data set the likelihood of one RNA chemical modification or multiple RNA chemical modifications.

This invention discloses a method for evaluating the egg production performance of Leizhou Black Ducks. By directly sequencing the FRRS1L gene and detecting single nucleotide polymorphisms, this invention identified multiple molecular markers and haplotypes associated with the egg production performance of Leizhou Black Ducks (starting weight, starting egg weight, and 300-day egg production). These findings can provide a scientific basis for the conservation, breeding, and development of Leizhou Black Ducks, and lay the foundation for the subsequent construction of a high-yielding core population.

The present invention discloses a method to quantify tRNA abundance and tRNA modifications in an RNA sample that comprises contacting RNA with oligonucleotides in the presence of a ligating agent and performing nanopore direct sequencing. It also discloses a kit to perform said method.

Disclosed are nucleic acid analysis and sequencing technologies. More particularly, it concerns compositions and methods for capture, enrichment, adaptation, and direct sequencing of short RNA molecules, including microRNAs (miRNAs), using sequencing platforms. Disclosed are methods of preparing a sequencing library for a target short RNA and kits. The methods further comprising sequencing a cassette comprising the target short RNA using a sequencing platform.

The application discloses a base recognition method and system based on a full convolution network, a medium and a program, and belongs to the technical field of base recognition. The method comprises the following steps: constructing a base calling model based on a DenseNet convolutional neural network; constructing a data set to train the base calling model; collecting an electrical signal of a base to be recognized; inputting the electrical signal into the trained base calling model for recognition to obtain a sequence of the base to be recognized. The base calling model provided by the application significantly improves the base calling accuracy of direct RNA sequencing through an innovative full convolution network architecture and species-specific model training, overcomes many defects of the prior art, and has a wide application prospect and remarkable beneficial effects.

This invention discloses an InDel molecular marker associated with growth traits in Haimen goats and its applications, belonging to the fields of biotechnology and livestock breeding. The InDel molecular marker is a 58bp ins mutation located at position 67616252bp in the genomic sequence of the PPP1R13B gene. This invention uses Haimen goat blood genomic DNA as a template and a direct sequencing method of PCR products to determine genotype. Haimen goats with the ID genotype exhibit excellent growth traits and can serve as an effective molecular marker for improving growth traits in Haimen goats. The detection method of this invention is simple, rapid, accurate, and inexpensive, which can improve the breeding efficiency of Haimen goats and has high application value.

The invention belongs to the technical field of Hu sheep molecular marker-assisted breeding, and particularly relates to application of a CNGA3 gene molecular marker g.39045A > C in Hu sheep molecular marker-assisted breeding. According to the invention, Hu sheep is taken as a research object, PCR amplification, direct sequencing of products and sequence analysis are adopted, CNGA3 gene polymorphism is analyzed, and correlation between different genotypes of polymorphic sites and different growth traits is comprehensively analyzed. An analysis result shows that g.39045A > C is remarkably related to the body length of the Hu sheep, and the body length of the AA genotype and the body length of the CC genotype are remarkably higher than the body length of the AC genotype (P > 0.05).