Long-fragment DNA (deoxyribonucleic acid) library long paired-end sequencing method

Abstract

The invention belongs to the technical field of complete genome sequencing, and particularly relates to a long-fragment DNA (deoxyribonucleic acid) library long paired-end sequencing method. The long-fragment DNA library long paired-end sequencing method based on a single-molecule sequencing platform comprises the following steps that: extracting complete genome DNA to construct a large-fragment library; through large-fragment library cloning mixing pool DNA, constructing a long paired-end sequencing library and a cloning mixing pool, extracting the DNA of the long paired-end sequencing library and the cloning mixing pool, and carrying out sequencing after a carrier is removed; and utilizing an extracted paired-end double-end sequence to carry out redundancy removal to obtain an unambiguous long paired-end sequence. The method is used for assisting in complete genome assembly, evaluating the assembly quality of an existing genome, authenticating a structural variation site and the like, and therefore, the assembly quality of the genome can be greatly improved.

Description

technical field [0001] The invention belongs to the technical field of whole-genome sequencing, and in particular relates to a long-fragment DNA library paired-end sequencing method, which is used for the assembly, verification, multi-genome comparison and identification of structural variation sites of whole-genome sequences, etc. . Background technique [0002] The development of DNA sequencing technology has a rich history, with many leaps forward in just over 40 years. From Sanger's electrophoretic sequencing technology, which opens the door to biological sequencing with the characteristics of high cost, low throughput, long read length, and high precision, to next-generation sequencing technology (NGS, Next generation sequencing) massively parallel sequencing, with low cost , high-throughput, short read length, and high precision have become the mainstay of biological sequencing, and now leading the new trend of single-molecule real-time synthesis sequencing, creating ...

AI Technical Summary

Problems solved by technology

Although the paired-end technology based on the Fosmid library is widely used due to its advantages of low cost, simple technology and high throughput, its shortcomings are also very obvious. The short read length of the NGS sequencing platform makes the sequence assembly process computationally intensive. Enlarging, it is still difficult to complete the sequence assembly. The assembled sequence may contain many gaps and is difficult to fill. It is difficult to locate the obtained scaffold on the chromosome and determine their relative positions without a reference sequence. Especially when the genome contains a large number of repetitive sequences or contains a large gene family and large fragments of repeats, these shortcomings are even more prominent, so its accuracy is far less than that of BAC terminal sequences

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