Portunidae mitochondrial COI gene universal primer, design and amplification method thereof

Abstract

The invention relates to the research field of cephalopod mitochondrial genome, and especially relates to a portunidae mitochondrial COI gene universal primer, and design and amplification methods thereof. The universal forward primer of mitochondrial COI gene of Portunidae is SEQ ID NO. 1: 5 '- TCNACAAAYCATAAAGAYATYGG-3 ', and that reverse prim is SEQ ID NO. 2: 5'-TANACYTCWGGRTGHCCRAARAAYCA- 3'.The present invention solves the problem that the prior art does not utilize the DNA barcode technology to identify the family Portunidae, and has the following advantages: 1) the universal primer ofmitochondrial COI gene of the family Portunidae provided by the present invention can amplify a plurality of families of Portunidae in batch at one time with high efficiency, and can provide great convenience for the species identification of the family Portunidae; 2) design universal primers in two conservative region of mitochondrial COI gene of family Portunidae to amplify an intermediate mutation sequence, that amplification length is about 600-700 bp, sufficient information sites are used for identification and differentiation of physical evidence, and a sequencing reaction can be complete, the operation is simple, and the cost is saved.

Description

technical field [0001] The invention relates to the field of cephalopod mitochondrial genome research, in particular to a universal primer and a method for designing and amplifying mitochondrial COI gene of Portunidae. Background technique [0002] As an emerging technology for species identification developed in recent years, DNA barcoding (DNA barcoding) technology has received more and more attention. The principle is usually to use a short mitochondrial COI (cytochrome c oxidase subunit Ⅰ) gene sequence for species identification has been widely used in fish, birds, reptiles, mammals and other biological groups, but it is not very common in crabs. Portunidae crabs are rich in resources, with many species, and have extremely high economic value; however, the classification of crabs is mainly based on traditional morphological and behavioral technical means, and phenotypic variation is significantly affected by environmental factors, resulting in phenotypic Uncertainty...

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Problems solved by technology

[0003] On September 29, 2017, the Chinese Patent Office disclosed an invention authorization for a spider mitochondrial genome full-sequence amplification primer and amplification method, the authorization publication number: CN104531688B. Long-PCR (L-PCR) amplification and sequencing are used to identify spiders, but it can only identify spider mitochondrial DNA, not cephalopod mitochondrial COI genes, and it is necessary to design multiple pairs of primers to amplify the entire genome sequence, which consumes a lot of time. Long time is not convenient and efficient

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