DNA probe library for hybridizing with kras gene and method for enriching kras gene fragments using the same
A DNA probe, gene fragment technology, applied in the field of KRAS gene enrichment and extraction
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Embodiment 1
[0148] Example 1: Enrichment and detection of KRAS gene in MDA-MB-231 cell line
[0149] (1) Prepare the DNA sample bank of the MDA-MB-231 cell line to be tested
[0150] 1. Extract the whole genome DNA of the MDA-MB-231 cell line, and then fragment it (the DNA sample bank obtained in this way is called "DNA sample bank derived from the whole genome")
[0151] 1.1 DNA extraction
[0152] Whole genome DNA was extracted from MDA-MB-231 cell line (human breast cancer cell line, from ATCC cell bank, catalog number: HTB-26) using Qiagen Blood&Tissue DNeasy Kit (Cat. No.: 69506). Operate according to the instructions in the manual.
[0153] Use spectrophotometer and gel electrophoresis system to detect the quality and concentration of DNA. The 260nm absorbance of dsDNA is greater than 0.05, and the absorbance A260 / A280 ratio between 1.8 and 2 is qualified.
[0154] 1.2 DNA Fragmentation
[0155] Dilute 3 µg of high-quality genomic DNA to 120 µl with low TE buffer. According to...
Embodiment 2
[0233] Example 2: Enrichment and detection of KRAS gene in HCT-116 cell line
[0234] Using the probe library consisting of SEQ ID NO.1 to SEQ ID NO.10 obtained in Example 1, the same method as in Example 1 was used to detect the HCT-116 cell line (human colorectal cancer cell line, from ATCC cells Library, product number: ATCC-CCL-247) of the KRAS gene, a single base mutation was found in the KRAS gene of the HCT-116 cell line, which was 38G>A. see results Figure 4 .
[0235] After DNA extraction, PCR amplification and Sanger sequencing, the above mutations have been verified.
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