12 results about "Amplification/Sequencing" patented technology

This invention provides a gene related to loquat flowering. SVP1 This invention relates to the field of molecular biology, specifically to a tightly linked SNP marker and its applications. The nucleotide sequence of this SNP marker is shown in SEQ ID NO.1. A genotype of C at the locus corresponds to a spring-flowering phenotype in loquat, while a genotype of G corresponds to an autumn-flowering phenotype. The invention also provides specific primers (SEQ ID NO.2 and SEQ ID NO.3) for detecting this SNP marker, a kit containing these primers, and a screening method for detecting loquat flowering time using this marker. By extracting loquat DNA and performing PCR amplification and sequencing analysis, the flowering period of loquat can be quickly and accurately determined. The SNP marker of this invention is highly linked to loquat flowering time, providing an effective tool for marker-assisted selection breeding of loquat at different flowering periods. This significantly improves breeding efficiency, shortens the breeding cycle, and is of great significance for extending the loquat fruit supply period and optimizing the industrial structure.

Provided herein are methods of targeted genomic sequencing requiring only a single amplification step. The Single Amplification Sequencing (SAS) assay provided herein amplies and sequences targeted genomic regions at reduced time and cost. The assay provides robust coverage and accurate drug resistance profiles for multiple first- and second-line Tuberculosis drugs, showcasing its clinical utility for rapid antimicrobial resistance (AMR) detection. SAS significantly simplifies targeted Next Generation Sequencing (tNGS) workflows and holds significant promise for revolutionizing precision diagnostics across a wide array of clinical and research settings.

The present invention relates to methods and compositions for preparing a sample enriched for single stranded nucleic acids. The single stranded nucleic acids find use as templates for synthesizing Xpandomer copies of the nucleic acids. The Xpandomer copies may be passed through a nanopore sensor to determine the nucleic acid sequence of the nucleic acids. The methods include asymmetric PCR, amplicon immobilization melt, exonuclease-mediated partial or complete digestion of one or both strand of an amplicon and linear amplification.

This invention provides a molecular marker associated with sheep weight traits and its application. Primers were designed based on the TNNT1 gene sequence to extract DNA from sheep blood. Through PCR amplification, DNA sequencing, and sequence analysis, a G / C polymorphism site was found at 176 bp of the amplified fragment. Further analysis using AQP primers detected the polymorphism site in 918 Hu sheep and established a least-squares model to analyze the association between genotype and weight traits. Ultimately, the TNNT1 gene fragment amplified by this invention was determined to serve as a molecular marker associated with sheep weight traits. By detecting the genotype of this molecular marker, this invention allows for the selection of GG homozygous sheep for breeding in the core herd, which can improve sheep weight and contribute to increased economic benefits.

The application belongs to the technical field of biology and relates to a phi29 DNA polymerase mutant with reduced polymerization rate and a preparation method and application thereof. The application designs and provides a mutant of phi29 DNA polymerase. The amino acid residues at at least one of positions 190, 324, 411, 49, 83, 375, 526, 85, 149, 284, 287, 35, 115, 178, 102, 246, 336, 506, 257, 502, 55, 113, 187, 289, 306, 517, 527, 140, 203, 368, 372, 89, 54, 277, 327, 163, 224 in the amino acid sequence of the phi29 DNA polymerase are modified, a new protein with reduced polymerization rate of the polymerase and good polymerase activity is generated, problems caused by the too fast polymerization rate of the phi29 DNA polymerase in practical application are successfully solved, the accuracy, stability and reliability of the phi29 DNA polymerase in DNA amplification, sequencing and other molecular biology experiments are improved, and the sequencing quality is greatly improved.

The application discloses a kind of extraction determination methods of soil nematode digestive tract microorganism, belong to the field of microbial detection technology. Including the following steps: (1) separation extraction nematode: shallow dish method is used to separate nematode in soil, in turn through washing and screening, shallow dish standing, starvation treatment, concentration and preservation operation;(2) digestive tract microorganism extraction: after species identification to nematode, select the nematode individual of specific nutrition group, after surface microorganism elution treatment, extract digestive tract microorganism DNA;(3) DNA amplification: using primer to the V3-V4 region of digestive tract bacteria 16S rRNA gene and the ITS region of fungal ITS gene is carried out PCR amplification;(4) sequencing and data analysis: after purification and quantification to PCR amplification product, using Illumina MiSeq platform carries out double-end sequencing, original data is split, quality filtering, sequence splicing, chimeric sequence removal, ASV clustering and species annotation processing.

The application provides a molecular marker related to an immune trait of sheep and application thereof. The molecular marker is designed according to a primer of a GRB10 gene sequence, DNA is extracted from sheep blood, PCR amplification, DNA sequencing and sequence analysis are carried out, a G / A polymorphic site is found at 119bp of the amplified fragment, further, the polymorphic site of 1086 Hu sheep is detected by using an AQP primer, a least square model is established, genotype and immune traits are associated, and finally it is determined that the G / A polymorphic site at 119bp of the amplified GRB10 gene fragment can be used as the molecular marker related to the immune trait of sheep. By detecting the genotype of the molecular marker, the application can select and keep GG homozygous sheep into a core group for breeding, can be used for improving the immune capacity of sheep, and is helpful to increasing economic benefits.

The present disclosure discloses a method for preparing a spatial chip, a spatial chip, and a use thereof. The method comprises: hybridizing to attach a single-stranded nucleic acid molecule to a surface of a solid-phase substrate, the single-stranded nucleic acid molecule comprising a first fixed sequence, a spatial barcode sequence, a sequencing primer binding sequence and a second fixed sequence, a first probe being attached to the surface of the solid-phase substrate of the chip, and the first probe comprising a complementary sequence of the first fixed sequence and a complementary sequence of the second fixed sequence; ring closing, causing the single-stranded nucleic acid molecule to form a circular nucleic acid molecule; rolling circle amplification; sequencing, to obtain sequence information of the spatial barcode sequence; synthesizing a single-stranded nucleic acid molecule by using the rolling circle amplification product as a template, to obtain a single-stranded nucleic acid molecule library; and fixing and clustering, fixing the single-stranded nucleic acid molecule library on the surface of the solid-phase substrate to form a single-stranded nucleic acid molecule cluster. According to the present disclosure, a chip having a nucleic acid probe carrying spatial position coordinate information can be prepared.

The present invention belongs to the field of biotechnology, and relates to a phi29 DNA polymerase mutant having a reduced polymerization rate, a preparation method therefore, and a use thereof. The present invention designs mutants of phi29 DNA polymerase, and by means of modifying amino acid residues at at least one of positions 190, 324, 411, 49, 83, 375, 526, 85, 149, 284, 287, 35, 115, 178, 102, 246, 336, 506, 257, 502, 55, 113, 187, 289, 306, 517, 527, 140, 203, 368, 372, 89, 54, 277, 327, 163, and 224 in the amino acid sequence of phi29 DNA polymerase, generates novel proteins that reduce the polymerization rate of the polymerase while maintaining good polymerase activity, successfully solving problems caused by the excessively fast polymerization rate of phi29 DNA polymerase in practical applications, improving the accuracy, stability, and reliability thereof in DNA amplification, sequencing, and other molecular biology experiments, and greatly enhancing sequencing quality.

The application provides a molecular marker related to sheep immunity traits and application thereof. The application uses a method for detecting a PLEKHH2 gene SNP site of Lake sheep by combining DNA pool sequencing with KASPar technology. A primer is designed according to a PLEKHH2 gene sequence, DNA is extracted from sheep blood, and PCR amplification, DNA sequencing and sequence analysis are carried out, so that a C / G polymorphic site at the 368th position of the amplified fragment is found. Further, KASPar primers are used to detect the polymorphic site of 889 Lake sheep and to establish a least square model, and correlation analysis is carried out on the genotype and immunity trait indexes, so that it is found that the site is significantly related to the platelet count (PLT) and the mean corpuscular hemoglobin concentration (MCHC) of sheep. Through detection of the molecular marker of the application, the application can be used in marker-assisted disease-resistant breeding of sheep and can accelerate the breeding process of sheep.

The application belongs to the technical field of molecular biology, and particularly relates to a detection method and a detection kit for a poly-A tail of RNA. The first aspect provides a detection method for a poly-A tail of RNA, comprising the following steps: incorporating first dNTPs into RNA by reverse transcriptase to synthesize a complementary DNA chain; amplifying and sequencing the complementary DNA chain by using DNA polymerase and taking second dNTPs as a substrate; and analyzing the length of the poly-A tail according to the sequencing result. The application develops a method for replacing raw materials based on deoxyinosine triphosphate (dITP) of hypoxanthine, in which, after template conversion of RNA to form cDNA, the pairing mechanism of dITP and other bases is used to replace the poly-A chain with a random chain, so that a relatively complex sequence can be obtained, the stability of the sequencing signal is improved, and the length of the poly-A tail can be more accurately obtained based on the complex sequence.