Preparation method of genome mixing sequencing library

Abstract

The invention relates to the technical field of molecular biology, and discloses a method for preparing a genome mixed-sample sequencing library, which comprises the following steps: (1) ultrasonic fragmentation of genomic DNA; (2) purification and end repair of fragmented products; (3) ) repair product for purification and adapter ligation; (4) ligation product purification recovery and concentration determination; (5) sample mixing and fragment screening; (6) PCR amplification of the product after fragment screening; (7) purification of the PCR product to obtain Sequencing library; (8) library quality inspection and on-machine sequencing. The present invention provides an adapter compatible with the Illumina next-generation sequencer in the above step (2), a method for purifying the product in the step (2) (3) (4) (7), and a method for mixing samples in the step (4) And the PCR amplification system and program setting in the step (6) ensure that the library building method provided by the present invention can quickly and smoothly carry out multiple sample mixed sample library building, and the method provided by the present invention can be used to obtain good data uniformity. and high-quality sequencing data.

Description

technical field [0001] The invention relates to the field of high-throughput sequencing technology in molecular biology. More specifically, it relates to a method for preparing a genome mixed sample sequencing library, which is suitable for resequencing or simplified sequencing of various samples of all eukaryotic organisms, especially for resequencing of small genes with multiple samples or simplified sequencing of large gene groups. Background technique [0002] Next-generation sequencing technology is currently the most commonly used technology in high-throughput sequencing research. After more than 30 years of development, DNA sequencing technology has made significant progress, and the second-generation sequencing technology characterized by high throughput has gradually matured and become commercialized. Early sequencing technologies mainly relied on first-generation sequencing. The various DNA sequencing technologies developed from the traditional chemical degradatio...

AI Technical Summary

Problems solved by technology

The characteristic of 454 sequencing is that the length of a single read grows, but the amount of data in a single reaction is low

However, for some small genomes, such as fungal genomes with a size of about 2.5-81.15Mb and organelle genomes such as chloroplast genomes with a size of 120K-217K, when the number of sequencing samples is large, if the amount of sequencing data is less than 2G, if a single library is built separately Library sequencing, the construction cost of a single sample library will exceed its sequencing cost

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