22 results about "Human DNA sequencing" patented technology

Among other things, provided herein are systems that replace the natural random integration activity of a retrovirus with site-specific integration machinery. This approach allows for a more precise targeting of a gene of interest into a human genome, e.g., for therapeutic purposes. The system may include integration-deficient retrovirus (e.g., lentivirus) (IDLV), in which the natural integration activity has been reduced (e.g., by mutation to the viral integrase polypeptide). Instead, the system may comprise a site-specific recombinase (e.g., a serine recombinase, e.g., a serine integrase) capable of directing insertion of a template DNA, or portion thereof, into a desired site in the human genome.

Disclosed are methods of preparing a composition of non-naturally occurring DNA, comprising: (a) extracting DNA from a sample of a subject; (b) contacting the extracted DNA or its derivative with a panel of oligonucleotide probes designed to hybridize to a plurality of preselected genomic regions, thereby generating selected DNA, wherein at least 50% of the preselected genomic regions are CpG regions that each comprises at least 3 CpG sites and a CpG density of at least 0.02 CpG / bp, wherein the preselected genomic regions cover 10-500 Mb of sequence space in human genome, wherein the extracted DNA or its derivative or the selected DNA or its derivative is further treated with an agent or a combination of agents that discriminates between methylated and unmethylated cytosines; and (c) performing high-throughput sequencing on the selected DNA or its derivative and generating sequencing reads.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

A genomic data analyzer may be configured to detect and characterize, with a variant calling module, genomic variants from next generation sequencing reads out of a pool of enriched genomic patient samples without suffering from next generation sequencing workflow biases such as those introduced by sequencing errors in particular in repeat patterns regions of the human genome such as homopolymers or heteropolymers. The variant calling module may estimate the probability distribution of the length of the repeat pattern for each patient sample and cross-analyze it against other samples in a single experimental pool to identify best-fit variant models for each pair of samples. The variant calling module may further group samples according to their matching best-fit variant models and identify which group of patient samples carries the wild type reference without the need for control data in the pool. The variant calling module may subsequently characterize the homozygous or heterozygous repeat patterns variants for each patient sample with improved specificity and accuracy even in the presence of next generation sequencing biases.

The invention discloses a colorectal cancer diagnosis system and application thereof, and the colorectal cancer diagnosis system comprises human SDC2, SFRP2 and TFPI2 gene methylation PCR detection reagents, 20 ng / L of unmethylated human genome standard substances, 50 excrement sample nucleic acids with the concentration of 5-40 ng / L and clinically and clearly diagnosed as normal people, and a kit for detecting the colorectal cancer. The kit is characterized in that the kit comprises 20 ng / L of 100% methylated human genome standard substance and 30 cases of excrement sample nucleic acid of patients with colorectal cancer stage I clinically and clearly diagnosed, wherein the concentration of the excrement sample nucleic acid is 5-40 ng / L. A multi-layer composite interpretation system formed by a multi-component system or a calculation method is used for effectively interpreting whether methylation of the gene in the sample to be detected is positive or negative, the positive rate of colorectal cancer diagnosis can be effectively increased, and therefore the value of early screening of the colorectal cancer is greatly increased.

This invention relates to the field of biomedical technology and discloses a broad-spectrum antimicrobial peptide and its applications. The invention obtains a broad-spectrum antimicrobial peptide H by mining non-classical open reading frames (ncORFs) in the human genome. Experimental verification shows that the broad-spectrum antimicrobial peptide H has significant inhibitory effects on both Gram-positive bacteria (Staphylococcus aureus ATCC 29213) and Gram-negative bacteria (Escherichia coli K88). Furthermore, the broad-spectrum antimicrobial peptide H can form an α-helix. The α-helix structure projection diagram shows that one side is rich in positively charged hydrophilic amino acid residues, and the other side is rich in hydrophobic amino acid residues, exhibiting good amphiphilicity. In addition, the broad-spectrum antimicrobial peptide H provided by this invention has low hemolytic activity and no cytotoxicity, ensuring in vivo safety while achieving highly efficient bactericidal activity. It also features a short synthetic sequence, small molecular weight, and is easy to chemically synthesize.

The present application relates to the technical field of biological medicine, and in particular to a molecular marker of alveolar rhabdomyosarcoma and application thereof, wherein the molecular marker is circFOXO1R-loop site, and a DNA-RNA hybridization region thereof is located at chr13:41133915-41134263 of a human genome. In the present application, the circFOXO1R-loop site is verified by CUT&Tag-qPCR and DRIP-qPCR, and it is found that the circFOXO1R-loop can promote the combination of FOXO1 gene and RNAPol II and DNA damage, and it has important clinical application value for developing a reagent or kit for diagnosing alveolar rhabdomyosarcoma, a drug or a gene therapy strategy for preventing and / or treating alveolar rhabdomyosarcoma aiming at the target.

A method of altering a eukaryotic cell is provided including transfecting the eukaryotic cell with a nucleic acid encoding RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site specific manner, wherein the cell expresses the RNA and the enzyme, the RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site specific manner.

An enzyme capable of mediating the integration of large DNA fragments into the mammalian genome and its applications are provided. Specifically, novel enzymes capable of integrating large DNA fragments into the genome are provided, including integrases as shown in SEQ ID NO: 1-7. Further, polynucleotides containing nucleotide sequences encoding integrases and host cells containing these polynucleotides are disclosed. A method for recombining target nucleic acids into the human genome using said integrase is also disclosed.

This invention relates to the field of bioinformatics analysis of sequencing data, specifically providing a genotyping method for short tandem repeat sequences based on next-generation sequencing. The detection algorithm considers various positional relationships between sequencing read lengths and STR regions, models each case separately, eliminates the effects of partial sequence mismatches, insertions, and deletions, and integrates all read information to calculate the optimal STR genotyping result that best matches the observation. This method can be used for genotyping detection of short tandem repeat sequences in the human genome.

The invention relates to methods for indexing samples during the sequencing of polynucleotide templates, resulting in the attachment of tags specific to the source of each nucleic acid sample such that after a sequencing run, both the source and sequence of each polynucleotide can be determined. Thus, the present invention pertains to analysis of complex genomes (e.g., human genomes), as well as multiplexing less complex genomes, such as those of bacteria, viruses, mitochondria, and the like.

The application provides a human genome T5 series fragment for multi-cancer identification and use thereof. A set of human genome differential methylation regions for multi-cancer identification, referred to as T5 series fragments (T5-1, T5-2, T5-3, T5-4, T5-5, T5-6, T5-7, T5-8) is disclosed. On this basis, the application provides a multi-cancer methylation technology which can cover more than 20 high-incidence cancer types, has high sensitivity (≥90%) and high specificity (≤5% false positive rate), and can realize rapid and micro-sample detection.

The invention belongs to the technical field of bioinformatics, and discloses a common value evaluation method and device for tumor neoantigens, equipment and a storage medium, missense mutation analysis is performed by acquiring mutation data of a pan cancer sample, a plurality of missense mutation sites are obtained and compared with a human genome to obtain candidate mutation sites, and the candidate mutation sites are used for evaluating the common value of the tumor neoantigens. Extracting the newborn peptide fragment corresponding to each candidate mutation site, inputting the newborn peptide fragment into the prediction model, predicting the binding score of the newborn peptide fragment and the human leukocyte antigen, and calculating the common value score of each newborn peptide fragment according to the number of the newborn peptide fragments generated at the candidate mutation sites and the corresponding binding score of the newborn peptide fragment and the human leukocyte antigen; according to the method, the new peptide fragments are sorted according to the descending order of the scores, and the new antigen sequence is screened out from the sorting result, so that the new antigen value can be effectively evaluated based on the number of the new peptide fragments generated by the site and the binding scores of the corresponding new peptide fragments and the human leukocyte antigen, and the new antigen with higher adaptability is screened out.

This invention discloses a method for constructing sequencing libraries based on blocking primers to remove human nucleic acids, and its application. The blocking primers include blocking primers for binding human repetitive sequences, blocking primers for binding human rRNA sequences, and blocking primers for binding human housekeeping gene sequences. These blocking primers can specifically bind to widely distributed scattered repetitive sequences, rRNA sequences, and housekeeping gene sequences in the human genome. The construction method includes the following steps: constructing a DNA library or an RNA library, or a co-constructed RNA and DNA library; using a library with added sequencing adapters as a template; and performing PCR amplification using adapter primers and blocking primers. The blocking primers can specifically bind to human nucleic acids through annealing and prevent modification by DNA polymerase extension, thus preventing the enrichment of human nucleic acid libraries through PCR amplification, while allowing normal amplification of pathogenic microorganism nucleic acids. This achieves the goal of reducing the proportion of human nucleic acids and provides guidance for pathogen detection.

A genetic prime editing screening platform to identify functional variants related to human health and disease, is configured substantially to annotate genome with nucleotide resolution with actionable disease prediction and treatment for personalized medicine.

The invention relates to a method for constructing a CDKL5 deficiency humanized mutation animal model and application of the CDKL5 deficiency humanized mutation animal model. The method comprises the step of knocking human CDKL5 gene R550X mutation and upstream and downstream genome sequences thereof into experimental animal genome CDKL5 gene loci. The human CDKL5 gene R550X mutation and upstream and downstream genome sequences thereof are knocked into experimental animal genome CDKL5 gene loci, CDKL5 protein C-terminal structural domain disease mutation is simulated, and the model has humanized mutation loci and nearby genome sequences, namely, has a to-be-repaired gene sequence same as that of a clinical patient, and can be applied to the field of clinical diagnosis and treatment of CDKL5 protein C-terminal structural domain disease mutation. The constructed model shows behavioral abnormality of a CDKL5 deficiency symptom, can be used for disease simulation and pathogenesis analysis of the CDKL5 deficiency symptom, can be used for directly testing gene editing and repairing strategies and therapeutic drugs of human genome specificity in the model body, and has important significance in the field of CDKL5 deficiency symptom treatment.

The application discloses a single-cell mixed sample sequencing and sample splitting method based on HLA genes. The application first acquires HLA gene typing information of each sample to be mixed; secondly, single-cell mixed sample sequencing is performed on the mixed sample to obtain single-cell mixed sample sequencing results of the mixed sample; finally, based on the HLA gene typing information, the sample source of each cell is identified by calculating the expression characteristics of HLA genes in the single-cell mixed sample sequencing sequence, and sample splitting is completed. The application breaks through the technical obstacle that few-cell clinical samples cannot be subjected to single-cell sequencing, greatly improves the utilization rate of these precious samples, significantly reduces the calculation consumption of sample splitting through an efficient HLA gene sequence pseudo-alignment algorithm, improves the ability to obtain genetic information from low-coverage cells by focusing on the HLA region with the highest polymorphism in the human genome, and effectively reduces the sequencing cost of a single sample through mixed sample sequencing.

The invention provides a method for constructing simulated tumor standard sequencing data and application of the simulated tumor standard sequencing data. The method comprises the following steps: acquiring a first strain mutation site set from a first human genome standard; obtaining a second strain mutation site set from a second human genome standard; selecting a unique germline mutation site relative to the second germline mutation set in the first germline mutation set; obtaining sequencing data of the second human genome standard substance and the first human genome standard substance; and for a preset simulated somatic mutation site, replacing corresponding data in the sequencing data of the second human genome standard with the sequencing data of the first human genome standard according to a predetermined replacement ratio, and generating the sequencing data of the tumor standard containing simulated somatic mutation. Compared with a real tumor cell standard substance, the method has the advantages that the mutation number and frequency can be accurately controlled, positive and negative sites are defined, the cost is saved, and the research efficiency is improved; and meanwhile, a real sequencing result is reserved.

There is provided a method of altering a eukaryotic cell comprising transfecting the eukaryotic cell with a nucleic acid encoding an RNA complementary to genomic DNA of the eukaryotic cell, transfecting the eukaryotic cell with a nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site-specific manner wherein the cell expresses the RNA and the enzyme, and modifying the eukaryotic cell with the nucleic acid encoding an enzyme that interacts with the RNA and cleaves the genomic DNA in a site-specific manner. The RNA binds to complementary genomic DNA and the enzyme cleaves the genomic DNA in a site-specific manner.

The invention provides a transcriptional activator-like effector nicking enzyme for editing a B2M gene locus and application thereof, the transcriptional activator-like effector nicking enzyme comprises a TALE protein, a target DNA recognition site of the TALE protein is located at the B2M gene locus, and the TALE protein comprises 18 repetitive modules; the B2M gene locus is a coordinate NC000015.10: 44711391-44721145 of a human genome of a human reference genome 38 edition (GRCh38 / hg38), and the B2M gene locus is a coordinate NC000015.10: 44711391-44721145 of The repeating module is selected from any one of NK, NI, HD and NG. The TALENickase gene editing tool suitable for double copy sites (B2M gene loci) is prepared by optimizing the length of a recognition module and combining amino acid mutation of a Fok I catalytic domain, and the optimized TALENickase has the highest site-specific integration efficiency and low toxicity.

Cerebral small vessel disease (SVD) is a leading cause of stroke and a major contributor to cognitive decline and dementia in the population. Evidences indicate that blood brain barrier dysfunction may play a significant role in VD pathogenesis. Recently, an inverse association of TRIM47 expression in brain and vascular tissues with extensive-SVD severity was reported in a human genome wide association study combined with summary-based Mendelian randomization studies and profiling of human loss-of-function allele carriers. Now, the inventors demonstrate TRIM47 is a key regulator of actin cytoskeleton organization through KEAP1 / NRF2 signalling pathway and might be protective from oxidative stress in brain EC. In particular, the in vitro TRIM47 knockdown decreases directed EC migration and delays EC adhesion process with loss of actin cortical reorganization and focal adhesion contacts. Furthermore, RNA sequencing and BioID results indicate that TRIM47 knockdown in brain EC, represses the expression of genes associated with cytoskeleton and NRF2 antioxidant pathway through a potential interaction with KEAP1. Accordingly, the present invention relates to the use of Nrf2 activators for the treatment of SVD.

The present invention relates to the medical field of single-gene disorders caused by functional loss or gain of an allele. The innovative approach developed being based on editing the human genome at the level of the Kozak sequence by means of CRISPR-Cas programmable nucleases. Particularly, the present invention relates to variant Kozak sequences and related in vitro or in vivo methods for obtaining such variant Kozak sequences for therapeutic applications in the treatment of single-gene diseases caused by monoallelic losses or gains. These in vitro and in vivo methods include CRISPR-Cas homology-directed repair, CRISPR-Cas prime editing, CRISPR-Cas base editing or genome editing with other programmable RNA-guided nucleases, and the introduction of specific nucleotide conversions in the Kozak sequence of genes causative of diseases. These nucleotide conversions enhance or inhibit the translation of the mRNA produced by the gene, compensating for the functional loss or gain of one allele in the diseases.