Unlock instant, AI-driven research and patent intelligence for your innovation.

Primer-probe composition for detecting TNNC1 gene mutation and application thereof

A primer-probe and composition technology, applied in DNA/RNA fragment, recombinant DNA technology, microorganism determination/inspection, etc., can solve the problems of high equipment requirements, high price, cumbersome process, etc., and achieve high sensitivity and low cost , good repeatability

Active Publication Date: 2021-06-11
DALI UNIV
View PDF5 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Capillary electrophoresis is the gold standard for gene mutation detection, but it is difficult to popularize from the laboratory to the clinic due to its high requirements for equipment and high price. Fluorescence quantitative PCR is fast, simple, economical and accurate.
[0006] CN102965428A discloses a sample preparation kit for detecting genetic mutations related to cardiac hypertrophy, which is aimed at genes ACTC1, ACTN2, BRAF, CALR3, CASQ2, CSRP3, GLA, HRAS, JPH2, KRAS, LAMP2, LDB3, MAP2K1, MYBPC3, MYH6, MYH7, MYL2, MYL3, MYLK2, MYOZ2, PRKAG2, RAF1, SOS1, TCAP, TNNC1, TNNI3, TNNT2, TPM1, TTN, TTR and all exon fragments of VCL, but relying on a large-scale gene sequencing platform, the process is very cumbersome and time-consuming Long time and very expensive

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Primer-probe composition for detecting TNNC1 gene mutation and application thereof
  • Primer-probe composition for detecting TNNC1 gene mutation and application thereof
  • Primer-probe composition for detecting TNNC1 gene mutation and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] Referring to the 2011 American Heart Association guidelines for the diagnosis of hypertrophic cardiomyopathy, a proband with familial hypertrophic cardiomyopathy clinically diagnosed in the Department of Cardiovascular Medicine, the First Affiliated Hospital of Dali University was obtained (the patient signed an informed consent and passed the Dali University The genome was extracted from the peripheral venous blood (approved by the Medical Ethics Committee), and the pathogenic mutation sites related to the pathogenesis of HCM were found through the whole exome sequencing technology, and a new mutation site related to HCM was found through the comparison and analysis of the sequencing results Point TNNC1, c.19G>C (the 19th base G in the coding region of the gene is mutated to C), this mutation leads to the change of A7P in the amino acid of TNNC1 gene, and the mutation screening method of real-time fluorescent PCR is used for the new mutation site Click to verify.

[00...

Embodiment 2

[0084] This example verifies the sensitivity and specificity of the primer-probe composition of the present invention.

[0085] (1) Sensitivity and repeatability of primer probe composition

[0086] The wild type and mutant type of TNNC1 gene c.19G>C were respectively diluted 10 times as a template, and the primer probe composition of the present invention was used for real-time fluorescent PCR detection (ABI7500 real-time fluorescent PCR instrument). The reaction system is shown in Table 3 As shown, the reaction conditions are as shown in Table 4, and the sensitivity of the primer probe composition was tested.

[0087] table 3

[0088] Reactive components Amount added (μL) PCR reaction solution 5 Rox Calibration Dye 0.1 forward primer 0.5 reverse primer 0.5 wild-type probe 0.5 mutant probe 0.5 template 0.8 pure water 2.1

[0089] Table 4

[0090]

[0091] The result is as Figure 4-Figure 7 As shown, as th...

Embodiment 3

[0103] The present embodiment uses the primer probe composition of the present invention to perform TNNC1, c.19G on 22 patients with hypertrophic cardiomyopathy (known genotypes sequenced by capillary electrophoresis, patients with informed consent, and approved by the Medical Ethics Committee of Dali University) >C mutation detection, using the kit of the present invention to carry out real-time fluorescent PCR reaction as described in Table 3 and Table 4, according to the results of fluorescence signal analysis, wherein, GG genotype: has a higher FAM fluorescence signal, HEX fluorescence signal is lower; GC Genotype: FAM fluorescence signal and HEX fluorescence signal have little difference; CC genotype: has higher HEX fluorescence signal, and FAM fluorescence signal is lower. Such as Figure 12 In the genotyping scatter diagram shown, there were 18 cases of GG genotype, 2 cases of GC genotype, and 2 cases of CC genotype. The detection results were consistent with the result...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to a primer-probe composition for detecting TNNC1 gene mutation and application thereof. The primer-probe composition comprises a primer used for detecting a TNNC1 gene mutation site c.19G>C and a probe, the primer comprises nucleic acid sequences shown in SEQ ID NO.1-2, the probe comprises a wild type probe and a mutant type probe, the wild type probe comprises a nucleic acid sequence shown in SEQ ID NO.3, and the mutant type probe comprises a nucleic acid sequence shown in SEQ ID NO.4. T. The primer-probe composition has high specificity and sensitivity, and can be used for carrying out real-time fluorescent PCR on a new mutation site c.19G>C of the TNNC1 gene, so that the genotype can be simply, conveniently, rapidly, accurately and economically judged.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a primer probe composition for detecting TNNC1 gene mutation and its application, in particular to a primer probe composition for detecting TNNC1 gene mutation site c.19G>C and its application . Background technique [0002] Hypertrophic cardiomyopathy (HCM) is a common and complex genetic heart disease. Except for heart transplantation, there is currently no effective treatment, which causes a serious economic burden to the family and society. The main pathological feature of HCM is asymmetric thickening of the left ventricle, double heart, or interventricular septum. Under normal circumstances, cardiomyocytes are regularly assembled into straight bundles of muscle fibers arranged in parallel. disorder, which can cause diastolic dysfunction. The pathogenesis of HCM often occurs in a familial clustering manner, and its molecular genetic basis is gene mutation. [0003] Troponin i...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12Q1/6883C12Q1/686C12N15/11
CPCC12Q1/6883C12Q1/686C12Q2600/156C12Q2561/113C12Q2563/107
Inventor 赵跃张世梅
Owner DALI UNIV