Construction method of mitochondrial genome sequencing library based on high-throughput sequencing and amplification primer

Abstract

The invention discloses a construction method of a mitochondrial genome sequencing library based on high-throughput sequencing and an amplification primer. The amplification primer comprises a primer pair 1 and a primer pair 2; and the nucleotide sequences of the primer pair 1 are shown as SEQ ID NO.1 and SEQ ID NO.2, and the nucleotide sequences of the primer pair 2 are shown as SEQ ID NO.3 and SEQ ID NO.4. According to the invention, only two pairs of amplification primers are needed, and a sufficient quantity of mitochondrial DNA samples can be obtained through one-time PCR amplification, so that the complex primer quantity and the cost input are greatly reduced, and the operation is simplified. According to the invention, the amplification products of the two pairs of primers are mixed to construct the library, so that the integrity and the accuracy of the mtDNA are improved, and the effectiveness of the data analysis is ensured.

Description

Technical field [0001] The present invention belongs to the field of high-throughput sequencing techniques, and it is related to the construction method of mitochondrial genome sequencing library based on high throughput sequencing and amplification primers. Background technique [0002] The mitochondria is a manufacturing plant of cell energy, and is an important orpatipotome of biological energy metabolism. The human mitochondrial genome is a double-stranded ring structure with a size of 16569 bp, encoding 13 functional proteins involved in the ATP synthesis process, 22 TRNAs, and 2 rRNAs. The mitochondrial DNA does not have a protein protection, and in a highly oxidized environment, it has the characteristics of high mutation rate. These mutations have caused changes in genetic structures, affecting mitochondrial function, causing abnormal phosphorylation and energy metabolism, eventually causing the occurrence of disease, such as common KSS syndrome, Leigh's syndrome. [0003...

AI Technical Summary

Problems solved by technology

However, the probe hybridization capture method is expensive and cumbersome to operate

In the PCR amplification method, multiple pairs of primers need to be designed for short-fragment amplification, and various primer sets are used in combination, which is cumbersome and costly; while for long-fragment amplification, some methods use segmented amplification to collect complete mitochondrial DNA. , but there is a defect that the amplification cannot be performed when the primer binding site is mutated, or there is a segmented amplification method that can correct each other, but it is necessary to carry out subsequent purification and library construction of different amplification products, which makes the workload and Reagent costs doubled

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