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A method for typing and identifying mice using fluorescent PCR technology

A type identification, mouse technology, applied in biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of cumbersome, low resolution, false negatives, etc.

Inactive Publication Date: 2018-12-04
DONGHUA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method is too cumbersome and needs to be annealed for a period of time before the next experiment can be carried out; using the enzyme digestion method, if the efficiency of the enzyme digestion is low, it will also produce false negative results; using agarose electrophoresis, the resolution is low

Method used

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  • A method for typing and identifying mice using fluorescent PCR technology
  • A method for typing and identifying mice using fluorescent PCR technology
  • A method for typing and identifying mice using fluorescent PCR technology

Examples

Experimental program
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Embodiment 1

[0034] (1) Mouse breeding and sample collection: The identified Founder mice (I used the Crispr / Cas9 system to knock out mircoRNA 505 (Gene ID: 751545) C57BL / 6J mice) as the Founder mice, and the Founder mice were identified The mouse knocked out 23bp, and it was recorded as Founder17) and crossed with C57BL / 6J mice to produce F1 generation mice. Take 1cm tail tissue of F1 generation mouse and store it at -20℃ for later use.

[0035] (2) DNA extraction: Use the animal genomic DNA extraction kit from Shenggong Bioengineering Co., Ltd. to extract the DNA from the middle tail tissue in (1) according to the instructions, and determine the DNA quality and concentration by 0.8% agarose gel electrophoresis .

[0036] (3) Primer design: Design a pair of PCR primers in the gene locus area knocked out by the Crispr / Cas9 system, according to the sequence of the microRNA505 of the C57BL / 6J mouse in the NCBI database, using the Primer3 online software (http: / / frodo.wi .mit.edu / primer3 / ) for p...

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Abstract

The invention relates to a method utilizing a fluorescence PCR technology to identify mouse genotype. A fluorescence PCR primer is utilized, after PCR amplification, the amplification product is mixed with ROX with a known size, the size of PCR products can be calculated by Genemapper software through 377 sequencing; and thus the mouse genotype can be identified. The provided method can simply, directly, and precisely find out the mice having micro RNA, whose Crispr / Cas9 has been knocked off.

Description

Technical field [0001] The invention belongs to the field of mouse typing and identification methods, and particularly relates to a method for typing and identifying mice by using fluorescent PCR technology. Background technique [0002] Gene targeting technology has been one of the important methods for studying gene function since its birth, revealing the biological functions of many important genes. In addition, researchers also look forward to using gene targeting technology to knock out or modify specific genes to achieve the purpose of curing diseases or improving livestock and poultry production traits. However, the early gene targeting technology is extremely inefficient, and it is difficult to truly apply it to medical treatment or livestock and poultry improvement practices. The CRISPR / Cas9 system, as an emerging gene-directed editing technology, has gradually matured and been successfully applied in many animal and plant species, which has greatly promoted the researc...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6888C12Q1/686
CPCC12Q1/686C12Q1/6888C12Q2600/158C12Q2600/178C12Q2545/101C12Q2563/107C12Q2565/125
Inventor 肖君华李雨王茂春仝莉周宇荀
Owner DONGHUA UNIV