7 results about "Primer binding site" patented technology

This application discloses a kit and method for quantitative detection of double-stranded DNA libraries compatible with multiple sequencing platforms, relating to the field of sequencing technology. The kit includes standards and a mixed primer combination. The standards include a set of serially diluted double-stranded DNA fragments of known concentration, each fragment including a PCR primer binding site sequence of a sequencing adapter from a first sequencing platform or a PCR primer binding site sequence of a paired-end barcode sequencing adapter from a second sequencing platform. The mixed primer combination includes a first primer pair and a second primer pair. The first primer pair specifically binds to the PCR primer binding region in the sequencing adapter from the first sequencing platform, and the second primer pair specifically binds to the PCR primer binding region in the paired-end barcode sequencing adapter from the second sequencing platform. The molar ratio of the first primer pair to the second primer pair in the mixed primer combination is 1:1. This kit can perform quantitative detection of libraries from different sequencing platforms. All samples are calculated using the same standard curve run, reducing systematic errors.

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.

The present invention relates to the field of genomic engineering. In particular, a chimeric prime editing (cPE) system is disclosed comprising elements including, but not limited to a Cas9 nickase (nCas9) / high fidelity nucleotide polymerase (HFNTPol) RNA, one or more single guide RNAs (sgRNAs), and a chimeric prime editor template oligonucleotide (cpetODN) comprising a deoxyribonucleic acid nucleotide polymerase template (NPT) and a primer binding site. For example, the sgRNA and the cpetODN are ligated into a single oligonucleotide. Alternatively, the sgRNA and the cpetODN are free and independent molecules (e.g., modular). This cPE system results in precise and efficient genome editing in cells and in adult mouse liver which is advantageous over conventional sgRNA prime editor fusion constructs. This flexible and modular system is an improvement in the art to obtain precise genome editing.

Provided are methods and compositions used for paired-end sequencing. The method comprises providing a DNA array comprising a surface immobilized with DNA concatemers. Foreach of the plurality of DNA concatemers on the array, the method comprises annealing first read primers to primer binding sites on the DNA concatemer, extending at least some of the first read primers to incorporate dNTPs or dNTP analogs, thereby producing first read strands, each of the dNTPs or dNTP analogs being incorporated is identified to produce first reads, performing controlled MDA by extending at least some of the first read strands with a polymerase having strand-displacement activity to generate a plurallty of second strands, each second strand comprising a portion that is hybridized to the DNA concatemer, and an unhybridized, single-stranded branch; and annealing second read primers to the single-stranded branches of the plurality of second strands, and extending the second read primers to generate the second reads.

Provided are prime editing systems with modification-tolerant polymerases and petRNAs and / or pegRNA with chemically modified primer binding sites (PBS) and / or nucleotide polymerase template (NPT).

The present invention relates to the field of biotechnology, genetic engineering and medicine, in particular, to a highly efficient system for editing the F508del mutation in the cystic fibrosis transmembrane regulator (CFTR) gene to the wild type, comprising a polynucleotide which comprises a nucleotide sequence, encoding the SpCas9 nickase recognizing PAM NGG or NG, and a prime editing guide RNA (pegRNA) comprising in its structure a sequence complementary to the target locus for editing the F508del mutation in human CFTR gene, a reverse transcription template (RTT) and a primer binding site (PBS), wherein the prime editing guide RNA (pegRNA) has sequence SEQ ID NO: 1-6 or a sequence comprising one or more replacements in the RTT compared to SEQ ID NO: 1-6, selected from SEQ ID NO: 7-168. The system for editing the F508del mutation in the cystic fibrosis transmembrane regulator (CFTR) gene to the wild type provides more efficient editing of the F508del mutation in the cystic fibrosis transmembrane regulator (CFTR) gene to the wild type and is a highly efficient cystic fibrosis treatment. The present invention also relates to the method of editing the F508del mutation in the cystic fibrosis transmembrane regulator (CFTR) gene using the system for editing the F508del mutation in the cystic fibrosis transmembrane regulator (CFTR) gene according to the present invention and the use of the system for editing the F508del mutation in the cystic fibrosis transmembrane regulator (CFTR) gene for treating cystic fibrosis.

This invention discloses the development and application of a specific marker associated with the shell thickness trait of the Chinese mitten crab (Eriocheir sinensis). This molecular marker is located in Chr09 of the Chinese mitten crab genome. Esine02654 The gene region exhibits significant sequence length polymorphism: the wild type contains a 72 bp nucleotide sequence, while in the mutant, this sequence is mutated to a single base A or deleted. This invention utilizes the specific design of downstream primers located within this variant region to develop a primer pair for detecting this structural variation. Amplification using this primer pair shows a specific 109 bp band in thick-shelled individuals (wild-type), while thin-shelled individuals (homozygous deletion mutant) show no amplification band due to the absence of the primer binding site. This marker detection is simple, and the results are intuitively interpreted, showing either a band or no band, making it suitable for early-stage auxiliary screening and breeding of the thick-shell trait in Chinese mitten crabs.