Kit for detecting hotspot mutation of KRAS gene and detection method thereof
A kit and gene technology, which is applied in the field of kits for detecting KRAS gene hot spot mutations, can solve problems such as limiting the application of Sanger sequencing method, and achieve the effects of short operation time, simple operation, and improved sensitivity
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[0033] 1. Synthesis of primers and probe sequences and synthesis of PNA sequences in the PCR reaction solution in the kit
[0034] According to the requirements of KRAS gene hotspot mutation region and sequencing fragments (see figure 1 ), design the following primers, probes and PNA, and synthesize the following primers, probes and PNA by artificial synthesis:
[0035] KRAS12F: ACTGGTGGAGTATTTGATAGTGTA (SEQ. ID NO. 1)
[0036] KRAS12R: CTTTATCTGTATCAAAGAATGGTCCTG (SEQ. ID NO. 2)
[0037] KRAS12P: FAM-ACCTTATGTGTGACATGTTCTAATATAGT-BHQ1 (SEQ. ID NO. 3)
[0038] KRAS12PNA: GGAGCTGGTGGCGTAGGC (SEQ. ID NO. 4)
[0039] The above primers, probes and PNA sequences are all from the 5' end to the 3' end, 12 indicates the codon number of the KRAS mutation hotspot region, F indicates the forward primer, R indicates the reverse primer, P indicates the probe, and FAM indicates the middle of the probe. Fluorescent group, BHQ1 represents a quenching group.
[0040]Insert the universal M...
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