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A sequencing method using restriction enzyme extension to increase the read length

A sequencing and read length technology, applied in the field of genetic engineering, can solve problems such as limited read length

Active Publication Date: 2014-10-01
盛司潼
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0012] The purpose of the present invention is to provide a sequencing method that utilizes restriction enzyme extension to increase the read length, aiming to solve the problem of limited read length in the prior art restriction enzyme extension method

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  • A sequencing method using restriction enzyme extension to increase the read length
  • A sequencing method using restriction enzyme extension to increase the read length
  • A sequencing method using restriction enzyme extension to increase the read length

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Embodiment Construction

[0066] In order to make the object, technical solution and advantages of the present invention clearer, the present invention will be further described in detail below in conjunction with the accompanying drawings and embodiments.

[0067] The invention provides a sequencing method for increasing the read length by restriction enzyme digestion, which comprises the following steps: first, the first anchor of the anchor primer is combined with the first anchor of the adapter on the original nucleic acid fragment to be tested, and the fluorescent probes with different label positions are used to Needle, repeat the steps of connecting the fluorescent probe, collecting the fluorescent signal, eluting the fluorescent probe, replacing the fluorescent probe and then connecting again, and according to the type of the connected fluorescent probe, read the first one of the original nucleic acid fragments to be tested. The sequence information of M nucleotides after the end of the anchor e...

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Abstract

The invention relates to the field of genetic engineering and provides a sequencing method capable of increasing a reading length through enzyme digestion extension. The method comprises the following steps: firstly, combining a first anchor on a connector 1 of an original nucleic acid fragments to be tested, and reading the sequence information of M nucleotides behind an extension end of the first anchor by a connection sequencing method; secondly, recognizing corresponding enzyme digestion recognition sites by using an incision enzyme, and cutting part or all of the read sequence information; connecting a new connector 2 onto a sequenced fragment which is obtained in the digestion recovery; and finally, anchoring and combining a second anchor on the connector 2, connecting a fluorescent probe to an extension end of the second anchor, and extending forwardly to read nucleotide sequence information. Through repeated operation, the nucleotide sequence information required by the original nucleic acid fragment to be tested is obtained, so that through the adoption of the method, the aims of lengthening and not limiting the sequencing reading length are fulfilled.

Description

technical field [0001] The present invention relates to the field of genetic engineering, and more specifically, relates to a sequencing method for increasing the read length by using enzyme digestion and extension. Background technique [0002] Next-generation sequencing (NGS) mainly includes pyrosequencing (sequencing by pyrosequencing), sequencing by synthesis (SBS) and sequencing by ligation (SBL). Among them, ligation sequencing has the advantages of high accuracy and low cost, and is suitable for transcript and comparative genomics research, especially the detection of single nucleotide polymorphisms (Single Nucleotide Polymorphisms, SNP). [0003] An existing ligation sequencing method uses endonuclease digestion and extension for sequencing, such as figure 1 As shown, the steps of the method include: [0004] (1), using a double-stranded oligonucleotide adapter with an enzyme recognition site to connect to the nucleic acid fragment to obtain the nucleic acid fragme...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68
Inventor 盛司潼
Owner 盛司潼