30 results about "Tert gene" patented technology

The invention relates to a CRISPR/Cas12a gene editing system and application thereof. The invention comprehensively utilizes biochemical, molecular biology, cell biology and other methods to screen and identify novel LiCas12a protein with endonuclease activity from a plurality of different bacteria. Based on LiCas12a, a CRISPR/LiCas12a gene editing system with high editing efficiency and high specificity is established in eukaryotic cells and prokaryotic cells separately, and genetic manipulation such as knockout, insertion and point mutation of genes is achieved; a molecular mechanism that the mutation of a TERT gene promoter promotes the cell proliferation of bladder cancer and liver cancer is disclosed. The genetic editing system based on CRISPR-LiCas12a provides a more accurate and efficient gene editing method for research fields such as disease pathogenesis and metabolic engineering transformation.

The invention belongs to the field of molecular diagnosis and provides a TERT gene combination mutation site detection kit. The kit provided by the invention comprises a primer pair capable of respectively amplifying fragments containing TERT gene polymorphic sites RS34094720, RS61748181, RS121918661, RS121918662, RS121918663 and RS121918664 and a sequencing primer for sequencing amplification products. The invention also provides a method for non-diagnostic detection of TERT gene combination mutation sites by the use of the kit.

The invention discloses a primer for detecting variation of benign and malignant related genes of thyroid nodules, a kit and a detection method. According to the primer, the kit and the detection method, variation of 15 loci of the six genes and variation of the three fusion genes can be detected simultaneously, and the BRAF gene, the KRAS gene, the HRAS gene, the NRAS gene, the TERT gene, the EIF1AX gene, RET/PTC1 fusion, RET/PTC3 fusion and PAX8/PPARgamma fusion are involved. Hotspot mutation and fusion variation of the genes are closely related to the benign and malignant thyroid nodules. Therefore, the primer, the kit and the detection method are used for detecting a sample of a patient and can assist doctors in benign and malignant identification of the thyroid nodules which cannot be clearly diagnosed in the cytology, the accuracy of identifying the benign and malignant thyroid nodules is improved, excessive medical treatment for the patient with the thyroid nodules is reduced, and the primer, the kit and the detection method have important practical significance and high economic benefit for saving medical resources of China.

The present invention discloses a TERT gene polymorphism detection kit through a pyrosequencing method, and a method thereof. With the kit, TERT gene polymorphism can be detected, and specifically rs2736100 (G is more than T) single nucleotide polymorphism is detected. The kit comprises primers represented by SEQ ID NO.2-4. According to the kit, accurate, fast and high-throughput detection can be performed on the TERT gene polymorphism so as to achieve effective prediction and prevention of susceptibility risk of lung cancer.

The invention belongs to the technology of clinical medicine gene detection and particularly relates to a kit for detecting mutation of a TERT gene promoter and a detection method and application thereof. The kit comprises a fluorescence labeling probe specifically recognizing mutation of a lotus C228T and mutation of a lotus C250T or other combinations of the TERT gene promoter, a pair of TERT specific primers and needed Master mix of all reagents. The specificity mutation C228T and the specificity mutation C250T of the TERT gene promoter are provided, and the sum of the specificity mutation C228T and the specificity mutation C250T accounts for about 100% of bladder cancer TERT mutation; a detection method is standardized, the reagents are subjected to optimization verification, the price is low, all ingredients in a reaction system are premixed, the operation process is simplified to the maximum extent, and operation errors are avoided. When the kit is used for detecting mutation of the TERT gene promoter, the number of influence factors is small, detection is precise, operation is easy, detection results are understood easily, and a reliable method for quickly detecting bladder cancer through urine is provided for clinical medicine.

The invention provides a mitochondrion-targeted gene editing complex, a preparation method and application thereof and a mitochondrion genome editing method. The gene editing complex comprises a mitochondria-targeted cationic liposome and an mtCRISPR/Cas9 system whihc are a mitochondrial TERT gene double-stranded homologous template, a crRNA of a specific targeting TERT, an RNA homologous recombination repair template and a Cas9 protein. A gene editing technology based on CRISPR/Cas9 is combined with a mitochondrial targeting means, the mitochondrion-targeted gene editing complex is provided,a normal mitochondrial TERT gene functional fragment is introduced to replace a mismatched gene, and the oxidative stress level is improved by up-regulating a mitochondrial TERT gene.

The invention provides a thyroid cancer auxiliary molecule diagnosis test kit. The thyroid cancer auxiliary molecule diagnosis test kit comprises amplification primers and probes for detecting BRAF, HRAS, KRAS, NRAS and TERT genes, internal control gene ACTB primers and probes, and external control gene BRAF primers and probes; the primers are specifically used for detecting a V600E site of the BRAF gene, a Q61R site of the HRAS gene, a G12C site of the KRAS gene, a G12D site of the KRAS gene, a G13R site of the KRAS gene, a G13D site of the KRAS gene, a Q61K site of the NRAS gene, a Q61P siteof the NRAS gene, a Q61H1 site of the NRAS gene, a Q61H2 site of the NRAS gene, a C228T site of the TERT gene and a C250T site of the TERT gene. The invention also provides a use method of the thyroid cancer auxiliary molecule diagnosis test kit. The problems of sensitivity, cost, operation convenience and the like of an existing thyroid cancer auxiliary molecule diagnosis test kit are solved.

The invention provides sgRNA (small guide Ribonucleic Acid) for specifically identifying a swine Tert (Telomerase Reverse Transcriptase) locus. The sgRNA is formed by pTert-exon1-sgRNA and pTert-exon2-sgRNA, wherein a nucleotide sequence which is responsible for target fragments in the pTert-exon1-sgRNA is SEQ ID NO.1; a nucleotide sequence which is responsible for target fragments in the pTert-exon2-sgRNA is SEQ ID NO.2. The sgRNA for the swine Tert locus is strong in specificity. The sgRNA for the swine Tert locus can utilize CRISPR/Cas9 gene editing system and a bifluorescence enrichment method and is capable of efficiently realizing large-fragment knockout of the swine Tert gene, the homozygote efficiency can be up to 50 percent, and preliminary technical support is provided for follow-up research on swine Tert gene functions.

The invention provides a multiple PCR (Polymerase Chain Reaction) primer composition, a reagent and a multiple PCR-breaking control system used for TERT (Telomerase Reverse Transcriptase) gene whole exome next-generation sequencing. The multiple PCR primer composition used for the TERT gene whole exome next-generation sequencing comprises primers disclosed from SEQ ID NO: 01 to SEQ ID NO: 32. Themultiple PCR primer composition, the reagent and the multiple PCR-breaking control system for the TERT gene whole exome next-generation sequencing solves the following existing technical problems that1: since the point mutation of the BRAF gene is mainly detected in the prior art, a detection range is limited; 2: real-time fluorescent quantitation PCR (Polymerase Chain Reaction) detection and a gene chip only can detect known sites; 3: a Sanger sequencing checkout length is smaller than a range of 800 bp; 4: low-frequency mutation can not be detected; and 5: when a plurality of sites are detected, operation is complex, and cost is high.

The invention discloses a combined marker for early diagnosis of liver cancer and an application of the combined marker. The combined marker for early diagnosis of the liver cancer disclosed by the invention is methylation of FAM78A, MKL1, DCDC2, SR, BCL2L11, CYP1B1, PTPN7, AXIN1, SEMA4D, PROX1, SHH, AK055957, DAB2IP, TBX15, GRASP, PPFIA1 and/or PSD4, and mutation of TERT and/or CTNNB1. According to the invention, liver cancer detection is realized by detecting the methylation level of gene combination in plasma cfDNA, and the combined marker has high sensitivity and specificity; Under the condition of mutation detection after adding genes, the early screening sensitivity is effectively improved by detecting methylation of a gene combination and mutation of TERT genes and CTNNB1 genes at the same time.

The invention belongs to the field of biological medicine, and discloses a biomarker for judging benign and malignant thyroid nodules, a multi-gene joint detection kit and application. The biomarker group comprises at least one of eight DNA single base mutations and four gene fusions; the eight kinds of DNA single base mutations are selected from a BRAF gene V600E, a TERT gene C228T/C250T, a KRAS gene G12C/G12V/Q61R, an NRAS gene Q61R and an HRAS gene Q61R, and the four kinds of gene fusion are selected from CCDC6-RET (Exon1-Exon12), NCOA4-RET (Exon8-Exon12), PAX8-PPARG (Exon10-Exon2) and ETV6-NTRK3 (Exon4-Exon14). The method and the kit are high in detection sensitivity, high in specificity and wide in application scene.

The present invention discloses a microRNA328 for regulating expression of TERT gene and an application of the microRNA328, and provides uses of the microRNA328 or a substance inhibiting expression ofthe microRNA328. The miR328 is found to have a significant inhibitory effect on an expression level of TERT mRNA in Hela cells and an oligonucleotide inhibitor of the miR-328 has a significant up-regulation effect on the expression level of the TERT gene mRNA in the Hela cells. The inhibitor of the miR-328 significantly increases angiogenic capacity of EPC. The EPC transfected by the inhibitor ofthe miR-328 significantly inhibits differentiation of NSCs into glial cells. Studies show that using miRNAs as targets or tools to enhance telomerase activity provide theoretical basis for treatmentof neurological diseases of vascular dementia (VD), etc. and have potential clinical application value.

The invention discloses a primer and kit for evaluating the effect of 131I on treating differentiated thyroid cancer. The kit can be used for simultaneously detecting the variation of nine gene loci of a BRAF gene, a TERT gene, an HRAS gene and an NRAS gene, and the variation of the gene loci is closely related to the treatment effect of 131I on differentiated thyroid cancer. Therefore, when the kit is used for detecting a sample of a patient, a doctor can be assisted in scientifically evaluating the clinical effect of treating a differentiated thyroid cancer patient by using 131I, a basis is provided for making a precise treatment scheme of the patient, and the kit undoubtedly has very important clinical use value.

The invention provides a multiplex PCR primer composition, a reagent and a control system for next-generation sequencing of whole exons of TERT gene. The multiplex PCR primer composition for the second-generation sequencing of the whole exons of the TERT gene comprises primers shown as SEQ ID NO: 01 to SEQ ID NO: 50. The multiplex PCR primer composition, the reagent and the control system for thesecond-generation sequencing of the whole exons of the TERT gene provided by the invention solve the following existing technical problems: 1, the point mutation of a TERT gene promoter is mainly detected in the prior art, and the detection range is limited; 2, real-time fluorescent quantitative PCR detection is carried out, and the gene chip can only detect known sites; 3, the Sanger sequencing test length is in the range of less than 800bp; 4, when multiple sites are detected, the operation is tedious, and the cost is high; and 5, low-frequency mutation cannot be detected.

The invention discloses a method for detecting hot spot mutation of a human TERT gene promoter. The method comprises the following steps: a, enriching templates by adopting DNA enrichment conventionalPCR aiming at different specimens; b, purifying the PCR product, determining the concentration by using an ultraviolet spectrophotometer, and performing concentration calibration and standard substance preparation; and c, carrying out TERT-146 (C/T) and TERT-124 (C/T) mutation rate quantitative detection on the calibrated specimens by adopting qPCR. According to the method, a qPCR technology platform which is conventionally developed clinically is adopted, the high-sensitivity TERT hot spot mutation rate quantitative detection method is established for DNA specimens from various sources, thelower limit of the detected mutation rate reaches 0.05%, and the method has the advantages of being high in detection sensitivity, high in specificity, capable of accurately and quantitatively detecting the mutation rate, low in cost, suitable for conventional clinical development and the like.

The invention particularly relates to a TERT mutation detection kit and an application of the TERT mutation detection kit in non-invasive tumor diagnosis. The invention provides a primer combination,a kit, a method and an application (ddPCR-TERTpm kit) for detecting TERT gene promoter mutation based on a ddPCR method, and aims to solve the problems of complex detection and low sensitivity in theprior art. The ddPCR-TERTpm kit is suitable for detection of tumor and urine mutation TERT promoters of bladder and renal pelvis cancer patients. After urine DNA is extracted, ddPCR amplification analysis is carried out, and the kit has the advantages of rapidness, simplicity, convenience, high sensitivity, specificity and the like. In addition, the detection kit or technology is also suitable fordetection of malignant melanoma, brain glioma, hepatocellular carcinoma and other tumors and blood specimens, and molecular diagnosis of thyroid tumors and DNA derived from fine needle puncture specimens.

The invention discloses miRNA138 and an application of the miRNA138 in regulation of TERT gene expression. The invention provides miRNA138 and an application of an inhibitor of the miRNA138. The invention finds that the miR138 has a remarkable inhibition effect on the expression level of TERT mRNA of Hela cells, an oligonucleotide inhibitor of the miR-138 has a remarkable up-regulation effect on the mRNA expression level of the TERT gene of the Hela cell, the inhibitor of the miR-138 has the advantages that the blood vessel forming capability of the EPC is obviously improved, according to theinfluence of the miR-138 inhibitor on the differentiation of the EPC co-cultured NSC, the EPC transfected by the inhibitor of the miR-138 obviously inhibits the differentiation of the NSC to the glialcells.

The invention discloses a method and a kit used for determining human TERT gene rs2735940 site polymorphism. The method comprises following steps: human genome DNA to be determined is provided; upstream primers and downstream primers used for amplification of sequences near human TERT gene rs2735940 site are provided, wherein the downstream primers possess mismatched base C; the human genome DNA to be determined is taken as a template, and the upstream primers and the downstream primers are used for PCR amplification so as to obtain amplification products containing CYGG or CCYGG segments, wherein Y is used for representing base C or T to be determined on human TERT gene rs2735940 site; a restriction enzyme is provided; the restriction enzyme is used for enzyme digestion of the amplification products so as to obtain corresponding enzyme-digested products; and it is determined that Y is used for representing base C or T based on the enzyme-digested products. The restriction enzyme is a restriction enzyme only used for realizing restriction digestion of one segment selected from CCGG and CTGG, or one segment selected from CCCGG and CCTGG. The method is rapid and reliable, and determination cost is reduced greatly.

The invention discloses a kit for detecting hot spot mutation of a human TERT gene promoter. The method is characterized in that the kit is characterized by comprising a DNA enrichment conventional PCR premixing reagent for detecting mutation sites of TERT gene promoters-124 (C/T) and-146 (C/T), -124 (C/T) and-146 (C/T) mutant plasmids, a human peripheral blood wild type DNA standard substance, and -124 (C/T) and-146 (C/T) mutation rate detection qPCR premix reagents. The kit provided by the invention adopts a fluorescent quantitative PCR technology platform which is conventionally developed clinically, is used for detecting DNA specimens from various sources, and has the characteristics of high detection sensitivity, strong specificity, accurate quantitative detection of mutation rate, low cost, suitability for conventionally developing clinically and the like.

The invention provides a probe composition, reagent and control system for TERT gene all-exon next-generation sequencing. The probe composition for TERT gene all-exon next-generation sequencing comprises sequences as shown in SEQ ID NO:01 to SEQ ID NO:36. The probe composition, the reagent and the control system for TERT gene all-exon next-generation sequencing provided by the invention solve thefollowing existing technical problems: 1, point mutation of a TERT gene is mainly detected in the prior art, so that the detection range is limited; 2, real-time fluorescent quantitative PCR detectionis performed, so that a gene chip can only detect known sites; 3, the Sanger sequencing test length is smaller than 800 bp; 4, low-frequency mutation cannot be detected; and 5, the operation is tedious and the cost is high when a plurality of sites are detected.