35 results about "Rare mutations" patented technology

Many areas of biomedical research depend on the analysis of uncommon variations in individual genes or transcripts. Here we describe a method that can quantify such variation at a scale and ease heretofore unattainable. Each DNA molecule in a collection of such molecules is converted into a single particle to which thousands of copies of DNA identical in sequence to the original are bound. This population of beads then corresponds to a one-to-one representation of the starting DNA molecules. Variation within the original population of DNA molecules can then be simply assessed by counting fluorescently-labeled particles via flow cytometry. Millions of individual DNA molecules can be assessed in this fashion with standard laboratory equipment. Moreover, specific variants can be isolated by flow sorting and employed for further experimentation. This approach can be used for the identification and quantification of rare mutations as well as to study variations in gene sequences or transcripts in specific populations or tissues.

The invention is directed to methods for increasing the sensitivity of high throughput sequencing, particularly for distinguishing true rare mutations from amplification, sequencing and other sample processing errors that occur in sequencing techniques. In one aspect, methods of the invention includes steps of (a) preparing templates from nucleic acids in a sample; (b) labeling by sampling the templates to form tag-template conjugates, wherein substantially every template of a tag-template conjugate has a unique sequence tag; (c) linearly amplifying the tag-template conjugates; (d) generating a plurality of sequence reads from the linearly amplified tag-template conjugates; and (e) determining a nucleotide sequence of each of the nucleic acids based on the frequencies, or numbers, of each type of nucleotide at each nucleotide position of each plurality of sequence reads having identical sequence tags.

The present invention is directed to a method for detecting and quantifying rare mutations in a nucleic acid sample. The nucleic acid molecules under investigation can be either DNA or RNA. The rare mutation can be any type of functional or non-functional nucleic acid change or mutation, such as deletion, insertion, translocation, inversion, one or more base substitution or polymorphism. Therefore, the methods of the present invention are useful in detection of rare mutations in, for example, diagnostic, prognostic and follow-up applications, when the targets are rare known nucleic acid variants mixed in with the wildtype or the more common nucleic acid variant(s).

The invention provides primers, a method and a chip for detecting alpha and/or beta-thalassemia point mutation and deletion mutation based on whole-gene capture sequencing and application of such primers, such method and such chip. The primers, the method, the chip and application thereof have the advantages that through designing of capture probes, relevant genes involved in alpha-thalassemia and beta-thalassemia are enriched and all mutation information including SNP and indel in full-length sequences of genes is detected; through addition of autosome, X-chromosome and Y-chromosome regions as well as upstream and downstream regions of coded genes as references, structure variations such as SNV and CNV are detected; compared with existing various hotspot mutation site detection technologies, the method is capable of detecting hotspot mutation information as well as some rare mutations and undiscovered new mutation types to detect and analyze full-length sequence specificity of target genes, fully covers the mutation types and makes up the defect that a conventional detection method easily causes missing detection of low-frequency mutations and rare mutations greatly.

The invention discloses a hyper-blocking fluorescence quantitative PCR method with high sensitivity for detecting rare mutation. The invention adopts specific primers and a probe technology and can rapidly detect gene point mutation in a blood sample. The method has the advantage that (1) the conventional ARMS primers are modified by LNA or mismatched bases are introduced into 3' tail 2th site totail 3th site so that the sensitivity of the mutation detection is not influenced and the specificity of the single base mutation based on the mutation specific primers is guaranteed, (2) the modifiedblocker probe completely complementary to the wild type gene is introduced so that the wild type background interference is further reduced with keeping the efficiency of the mutant amplification, (3) based on the high specificity modified blocker probe, the ARMS primers can detect different base mutation forms at the same site at the same time so that multiple sites in the same gene can be detected by only a few systems, (4) sensitivity is high and (5) a detection rate is fast.

The invention relates to a genetic detection method, in particular to a quantitative detection method for unusual genetic mutation. The quantitative detection method for unusual genetic mutation comprises the following steps: firstly, corresponding primer sequences are designed and prepared according to the unusual genetic mutation required for quantitative detection, wherein the 3' end of a primer is provided with a mutant site - special nucleotide; except for the final site of the nucleotide, all the other sequences can be synthesized by commercial means; and the final site of the nucleotide is modified nucleotide, so as to make the primer incapable of extending in the general PCR reaction; and the modified nucleotide is added to the tail of the primer; and secondly, in a reaction system which contains special DNA polymerase and pyrophosphate, the primer designed and prepared in the first step is adopted for amplifying the unusual genetic mutation for quantitative detection, and a real-time fluorescent PCR instrument is utilized for quantitative detection; and the quantitative detection information of the unusual genetic mutation can be acquired through analysis of an amplifying curve recorded by the real-time fluorescent PCR instrument.

The invention provides a reagent and a method for detecting the ninth exon mutation of a CALR gene by using a Sanger sequencing technology. The reagent and the method are high in detection specificity, and the mutation types of insertion and deficiency of the ninth exon of CALR can be specially analyzed, so that the false positive problem is eliminated, and missing inspection of rare mutation can also be avoided. The reagent and the method can be used for clinically screening genes of patients suffering from primary thrombocythemia and primary myelofibrosis.

The present disclosure encompasses methods of error corrected sequencing (ECS) that enable detection of very rare mutations well below the error rate of convention next generation sequencing (NGS). Further, the methods disclosed herein enable multiplex targeting of genomic DNA.

A method of identifying a gene or genomic mutation that is linked to causality of a neuropsychiatric disorder is provided. The method comprises identifying exons which exhibit an expression level that is at least within the 75^th percentile of exon expression levels within a nucleic acid-containing sample from a mammal having a neuropsychiatric disorder; comparing the sequence of each identified exon to the sequence of a corresponding exon from a healthy control to identify rare or de novo sequence mutations within the identified exon; calculating the burden of rare or de novo mutations within the exon; and determining the correlation between expression level of the identified exon and burden of de novo or rare mutations in the exon, wherein an inverse correlation indicates that the exon gene is linked to causality of the neuropsychiatric disorder.

The invention discloses primers, method and kit for whole genome detection of human mitochondria. The primers comprise forward and reverse primers for amplifying 37 gene coding regions of a mitochondrial whole genome (16569bp), and the base sequences of the forward and reverse primers are shown in the formulas of SEQ ID NO: 3-222. A detection method based on the NGS technology comprises the following steps: combining specific primers and a target template DNA sequence, amplifying a target region of a to-be-detected sample by using universal primers, purifying a library by using magnetic beads,performing high-throughput sequencing on the obtained library, and analyzing mutation of the mitochondrial gene. According to the primers, the method and the kit disclosed by the invention, the raremutation of mitochondrial DNA can be accurately detected, a high cost performance, least manual operation and ultrahigh sensitivity are realized, guidance for drug selection of familial mitochondrialpatients and genetic susceptibility is provided, mitochondrial diseases caused by mitochondrial DNA mutation assists diagnosis and the onset risk is effectively reduced.

The invention discloses a non-syndromic cleft lip related low-frequency/rare mutation and a detection method thereof. The TAF11 gene mutation is single point mutation c.G144C (chr6:34855591) in a first exon region, the amino acid is changed into p.L48F, and heterozygous mutation is pathogenic. The pathogenic mutation provided by the invention is not reported to be related to the occurrence and development of the non-synthetic cleft lip, and can provide a basis for the screening and detection of disease-causing genes and the formulation of treatment programs of non-syndromic cleft lip. At the same time, the invention constructs a disease-causing gene detection method. The method comprises the following steps: capturing and enriching DNA in a whole genome exon region by using a sequence capture technology, performing high-throughput sequencing, performing bioinformatics analysis and co-separation analysis to screen out candidate genes, and performing Sanger sequencing to verify the mutation.

The present invention relates to highly sensitive and specific methods for detection of nucleic acids, which for example are useful for detection of rare mutations, or for detection of low-abundance variants in nucleic acids sequences. The methods involve an asymmetric incremental polymerase reaction (AIPR) followed by an exponential polymerase chain reaction (PCR).

Bottleneck Sequencing System (BotSeqS) is a next-generation sequencing method that simultaneously quantifies rare somatic point mutations across the mitochondrial and nuclear genomes. BotSeqS combines molecular barcoding with a simple dilution step immediately prior to library amplification. BotSeqS can be used to show age and tissue-dependent accumulations of rare mutations and demonstrate that somatic mutational burden in normal tissues can vary by several orders of magnitude, depending on biologic and environmental factors. BotSeqS has been used to show major differences between the mutational patterns of the mitochondrial and nuclear genomes in normal tissues. Lastly, BotSeqS has shown that the mutation spectra of normal tissues were different from each other, but similar to those of the cancers that arose in them.

The invention discloses a novel FBN2 gene mutation marker and application thereof in CCA auxiliary diagnosis. According to the novel FBN2 gene mutation marker and the application thereof in the CCA auxiliary diagnosis, a whole exome sequencing analysis is carried out on 27 CCA patients of 10 families, it is found that the following rare mutations exist on FBN2: c.3353A > G, c.3350A > G, c.3088G > A, c.2759G > A and c.3467G > A, mutation sites are verified through a CCA clinical scoring system, the accuracy and reliability of the FBN2 gene mutation CCA diagnosis are proved, on one hand, the FBN2 gene mutation is beneficial to explaining the pathogenesis of a CCA patient, on the other hand, the mutation site can be used as a CCA biomarker for diagnosis or auxiliary diagnosis of CCA, assessment of the risk of suffering from CCA and pre-pregnancy early warning.

The invention relates to a method for detecting rare gene mutations. The method utilizes high-selectivity gene mutation detection primers and hinders, to achieve a purpose of enriching rare mutation products by specifically inhibiting wild-type amplification. The method, the primers and the hinders in the invention are particularly suitable for efficient and low-cost detection of rare gene mutations in cfDNA according to an NGS technology.