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Simple and cheap genome sample breaking method applied to second generation sequencing

A sample and sequencing technology, applied in biochemical equipment and methods, combinatorial chemistry, microbial measurement/inspection, etc., can solve the problems of cumbersome operation, limited area occupied, large experimental bench area, etc., and achieve the effect of wide application prospects

Active Publication Date: 2013-09-11
BEIJING NOVOGENE TECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the instrument and consumables are relatively expensive, and this instrument occupies a relatively large area of ​​the experimental bench, and it needs to be equipped with a computer to cooperate with the operation; the Bioruptor instrument is relatively cheap, has no special consumables, occupies a limited area, and does not require computer operation. Under the premise of regular use of the instrument, ideal fragments can be obtained, but the operation is relatively cumbersome

Method used

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  • Simple and cheap genome sample breaking method applied to second generation sequencing
  • Simple and cheap genome sample breaking method applied to second generation sequencing
  • Simple and cheap genome sample breaking method applied to second generation sequencing

Examples

Experimental program
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Effect test

Embodiment

[0044] Example: fragmentation of DNA and construction of a library using an ultrasonic cleaner.

[0045] 1. Fragmentation of genomic DNA

[0046] 1) Determination of sample concentration

[0047] Use Qubit (USA) to measure the DNA concentration of the sample for precise quantification, and use 0.8% agarose gel, 120v voltage, and electrophoresis for 1 hour to test the quality of the sample to ensure that the genomic DNA sample is intact and not degraded.

[0048] 2) Using 100 ng-1 ug of genomic DNA detected in step 1) as a template for sample fragmentation. The sample system and crushing conditions are as follows: sample DNA (150-250ng / μl) 0.1-1μg, add H 2 0 to a total volume of 50 μl, the above mixture was crushed in an ultrasonic cleaner for 90 s, 150 s, and 360 s, respectively, and used to construct libraries with insert sizes of 180, 300, and 500 bp, respectively. 20ul was taken out for electrophoresis detection, the results showed that the fragmentation was sufficient, ...

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Abstract

The invention provides an efficient, fast, cheap and simple sample breaking method, successfully establishes a process for constructing a high-throughput sequencing library by the method. The method can be applied to Hiseq2000 sequencing. The method can be applied to denovo sequencing, re-sequencing, exon trapping or any other high-throughput sequencing library construction.

Description

technical field [0001] The invention relates to the field of high-throughput sequencing library construction, in particular to a method for crushing genomic DNA samples and its application. Background technique [0002] Data quality control in high-throughput sequencing technology is an important link in the sequencing process, and the quality of sample libraries is the premise and basis for obtaining high-quality sequencing data. Obtaining DNA fragments of appropriate size is a necessary step for sample library preparation before sequencing. If the fragments are too long, it will lead to difficulties in splicing data during analysis; if the fragments are too short, it will lead to a reduction in the amount of data, waste and increase the cost of sequencing. Nucleic acid fragmentation techniques are mainly divided into three categories: one is the chemical method, which is mainly used in the sample preparation process of mRNA; the other is the enzyme digestion method, which...

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C40B40/06C40B50/00
Inventor 蒋智刘少卿彭献军王大伟刘运超吴静王秋梅
Owner BEIJING NOVOGENE TECH CO LTD