Construction method of gene detection library of hereditary hypertrophic cardiomyopathy and kit
A technology for hypertrophic cardiomyopathy and gene detection, which is applied in chemical libraries, biochemical equipment and methods, and microbial determination/inspection.
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Embodiment 1
[0103] Four peripheral blood samples were used for library construction, using the full coding region and variable splicing region (exon to intron extension) of ACTC1, ACTN2, MYBPC3, MYH7, MYL2, MYL3, TNNI3, TNNT2, TPM1 genes 20bp) as the primer pool of the target region for multiplex reaction PCR, combined with the high-throughput sequencing platform Miseq for DNA sequencing, and then detected point mutations (SNP) and small fragment insertion-deletion (InDel). The specific operation process is as follows:
[0104] (1) Nucleic acid extraction and quality inspection: After nucleic acid extraction and quality inspection of peripheral blood samples, it is required to meet certain quality control standards: DNA integrity is good, no protein RNA pollution, DNA concentration ≥ 20ng / μL; DNA purity: OD260 / 280 1.8-2.0, OD260 / 230>2.0; total DNA input amount: 0.1μg-1μg.
[0105](2) Library preparation: Using the above DNA, make up to 52.5 μl with TRIS-EDTA after sampling, perform ultr...
Embodiment 2
[0113] 16 buccal swab samples were used for library construction, using the full coding region and variable splicing region (exon to intron extension) of ACTC1, ACTN2, MYBPC3, MYH7, MYL2, MYL3, TNNI3, TNNT2, TPM1 genes 20bp) as the primer pool of the target region for multiplex reaction PCR, combined with the high-throughput sequencing platform Nextseq550AR for DNA sequencing, and then detecting point mutations (SNPs) and small fragment insertions and deletions (InDels). The specific operation process is as follows:
[0114] (1) Nucleic acid extraction and quality inspection: After nucleic acid extraction and quality inspection of peripheral blood samples, it is required to meet certain quality control standards: DNA integrity is good, no protein RNA pollution, DNA concentration ≥ 20ng / μL; DNA purity: OD260 / 280 1.8-2.0, OD260 / 230>2.0; total DNA input amount: 0.1μg-1μg.
[0115] (2) Library preparation: Using the above DNA, make up to 52.5 μl with TRIS-EDTA after sampling, pe...
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