Method for constructing sequencing library by cyclizing method

Abstract

The invention relates to the field of genetic engineering and provides a method for constructing a sequencing library by a cyclizing method. The method comprises the following steps: (A) connecting a nucleic acid fragment with a connecting module and cyclizing the nucleic acid fragment and the connecting module without increasing the sizes of the both so as to obtain a first cyclized molecule, wherein the connecting module contains at least one enzyme digestion recognition site; and (B) on the basis of the enzyme digestion recognition site in the step (A), carrying out enzyme digestion on the nucleic acid fragment in the first cyclized molecule by using incision enzyme, connecting an enzyme digestion product with a bridging module, and cyclizing the enzyme digestion product and the bridging module so as to obtain a bridged cyclic compound. The method provided by the invention has a wide application range; known sequences can be inserted into small fragment nucleic acid molecules in an oriented way so as to construct the sequencing library; and the cyclizing efficiency in the library constructing process can be further increased.

Description

technical field

[0001] The invention relates to the field of genetic engineering, in particular to a method for constructing a sequencing library by circularization. Background technique

[0002] In the past few years, second-generation high-throughput sequencing technology has achieved rapid development. Among them, the method of sequencing based on ligase has the advantage of high accuracy, but the application of this type of sequencing method is limited due to its short single read length. In response to this situation, in recent years, there has been a method to improve the sequencing read length by constructing a sequencing library with a special structure, that is, inserting one or more linker sequences in the fragment to be tested, so that the same sequencing library molecule has multiple connections Sequence the starting sites, and then perform ligation sequencing on these different starting sites, thereby increasing the sequencing read length. The core of this met...

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