Multiple PCR primers for detecting non-small cell lung cancer oncogene mutation based on high-throughput sequencing, kit and method
A non-small cell lung cancer, high-throughput technology, applied in the field of multiplex PCR primers, can solve the problems of inability to meet the needs of non-small cell lung cancer detection of gene mutations, difficult to obtain materials for advanced patients, false positives and false negatives, etc., and achieve a simple detection method. The effect of easy operation, fast detection speed and high accuracy
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[0032] The samples are: Genomic DNA of normal blood, cfDNA of normal human plasma, MCF10A cell line genomic DNA, liver cancer FFPE genomic DNA, using DNA extraction kits to extract genomic DNA from various types of samples, using SEQ No.1~SEQ No.392 performs multiple PCR amplification on the extracted genomic DNA, and then uses the IlluminaNextSeq 500 sequencer for sequencing analysis. The specific implementation method is as follows:
[0033] A. Genomic DNA extraction: use Qiagen DNeasy Blood&Tissue Kit, and refer to the kit instructions to extract genomic DNA. The obtained genomic DNA is subjected to gel electrophoresis and quality detection using NanoDrop 100. It is required that the genomic DNA is not significantly degraded, the concentration is greater than 20ng / μL, A260 / A280>1.8, A260 / A230>2.0, and Qubit 3.0 is used for accurate quantification.
[0034] B. The first round of multiplex PCR amplification: use the amplification primer mixture R1-barcode (SEQ ID NO:1~SEQ IDN...
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