Human FGFR1 gene copy number variation nucleic acid standard substance, preparation method thereof and kit

Abstract

The invention discloses a human FGFR1 gene copy number variation nucleic acid standard substance, a preparation method thereof and a kit. The human FGFR1 gene copy number variation nucleic acid standard substance comprises background genome DNA and linearized plasmid DNA containing an FGFR1 complete genome, wherein the background genome DNA is extracted from a human normal B lymph suspension cell line. In the invention, the preparation of the background genome only needs to culture one cell line, the cell line is single in type and less in workload, and during mixing, different amounts of linearized plasmid DNA are respectively mixed into the background genome DNA, and the backgrounds of different gradient standard substances are consistent, so that CNV analysis of a high-throughput sequencing method is especially facilitated; and meanwhile, in the nucleic acid standard substance, a fragment inserted by the linearized plasmid DNA is the FGFR1 complete genome, the length reaches 100kbp or above, a real gene amplification situation can be simulated, the nucleic acid standard substance is suitable for various CNV detection methods, and the problem that the application of a standard substance for local region amplification is limited is solved.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a human FGFR1 gene copy number variation nucleic acid standard substance, a preparation method thereof, and a kit. Background technique [0002] Copy number variation (CNV) is an important source of genetic variation, which is a variation of DNA fragments above 1 kb. The specific performance is that the variable fragments are repeated and the number of repeated genome fragments differs between individuals, including deletions, duplications, inversions, and translocations, which greatly enrich the diversity of genetic variation in the genome. CNV is widely distributed in the human genome, and generally manifests in two modes: one is the broad CNV (Broader CNV, bCNV) that occurs due to abnormal chromosome segregation during mitosis, and the repetitive region can occur in most regions of the chromosome arm; the other is One is focal CNV (focal CNV, fCNV) that occurs in...

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Problems solved by technology

[0005] The preparation of existing copy number variant nucleic acid standard substances is mainly based on two methods: 1. Cultivate tumor cell lines from different sources, and extract genomic DNA separately. Different cell lines contain different copy numbers of the target gene, so as to prepare multiple gradients. Copy number variation nucleic acid standard substances, this method has the defects of many types of cultured cell lines and heavy workload, and the genomic backgrounds of different gradient standard substances prepared by this method are inconsistent, which will cause certain problems to the detection of CNV based on high-throughput sequencing methods. Second, use the traditional PCR method to amplify 1-2 kbp target gene DNA fragments, and then mix the fragments into the background cell line genomic DNA to prepare multiple gradient copy number variation nucleic acid standard substances, however , the length of the fragment amplified by this method is small, the amplification of the local region is difficult to represent the amplification of the genomic DNA region of the whole gene, and it is difficult to simulate the real gene amplification situation, and the application of the standard substance prepared by it has limitations, such as based on the PCR method When detecting CNV, if the designed primer probe is not within the scope of the amplified gene fragment, the standard substance is not suitable for this method

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