Genetyping method based on SNP site nucleic acid mass spectrum detection and application thereof
A genotyping method and nucleic acid mass spectrometry technology, which is applied in the field of genotyping based on SNP site nucleic acid mass spectrometry detection, can solve problems such as probes not being correctly combined with templates, false negative hybridization results, and affecting detection results, etc. The results are clear and easy to understand, the operation time of personnel is short, and the effect of high degree of experiment automation
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[0061] Example 1: Genotyping Mycobacterium tuberculosis using a genotyping method based on SNP site nucleic acid mass spectrometry detection
[0062] 1. Genomic DNA Extraction of Mycobacterium tuberculosis
[0063] (1) Take 2ml of the patient's sputum sample in the biological safety cabinet, add an equal volume of 10% NaOH solution, shake at 150rpm / min, 37°C for 30 minutes, and use the QIAamp DNA Mini Kit (Qiagen) for tuberculosis branching Extraction of bacillus genomic DNA.
[0064] (2) Primer design
[0065] The three sites rpoB516, rpoB526, and ropB531 of the rpoB rpoB of Mycobacterium tuberculosis resistance gene were designed for SNP site primers, and specific PCR primers and single-base extension primers were designed for these three SNPs. The PCR primer sequences are shown in Table 1.
[0066] Table 1
[0067]
[0068] (3) Single-tube multiplex PCR reaction
[0069] The target fragment containing the ropB gene was amplified by single-tube multiplex PCR technolo...
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