A kind of primer and preparation method of microsatellite dna molecular marker of Taihu new whitebait
A DNA molecule and microsatellite technology, applied in the field of molecular biology, can solve problems such as the undeveloped new silverfish in Taihu Lake, and achieve the effect of high polymorphism and enrichment of genetic basic information.
Active Publication Date: 2017-11-14
JIANGHAN UNIVERSITY
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[0006] In order to solve the problem that the research on the invasion mechanism of the new whitebait in Taihu Lake has not yet been carried out in the prior art, the embodiment of the present invention provides a primer set for microsatellite DNA molecular markers of the new whitebait in Taihu Lake and its preparation method
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[0082] 1. Acquisition of microsatellite sequences in the genome of Taihu Newbait:
[0083] After the first PCR amplification of the biotin repeat probe and the restriction fragment of the genome of Taihu Newbait, the first amplification product was obtained, and the first amplification product was hybridized with the biotin repeat probe to obtain The hybridization product, the microsatellite target fragment in the genome of Taihu New Silverfish was screened out by the magnetic bead enrichment method, and the microsatellite fragment was sequenced after cloning and positive identification to obtain the corresponding microsatellite site sequence.
[0084] Specifically, phenol chloroform was used to extract the genome of Taihu Newbait, and the extracted genome of Taihu Newbay was digested with MseI enzyme to obtain an enzyme-digested fragment of the Taihu genome.
[0085] Specifically, the conditions for the first PCR amplification were: pre-denaturation at 94°C for 5 minutes; 94°...
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The invention discloses a primer for microsatellite DNA molecular marker of Taihu new whitebait and a preparation method thereof. The primers for the microsatellite DNA molecular markers of Taihu New Silverfish include: primers shown in SEQ ID NO.1-SEQ ID NO.38 in the sequence listing. The preparation method comprises: hybridizing the enzyme-cut fragment of the genome of the new whitebait in Taihu Lake with the biotin repeat probe after the first PCR amplification, screening out the microsatellite fragment through the magnetic bead enrichment method of the hybridization product, and the microsatellite fragment. The satellite fragments were sequenced after cloning and positive identification to obtain the corresponding microsatellite locus sequence; primers were designed, and PCR amplification was performed between the primers and the genome of the new whitebait in Taihu Lake; the amplified products were separated by electrophoresis, and then fixed and silver-stained Processing; select microsatellite primers for polymorphic loci. The 19 pairs of primers obtained by this preparation method can amplify microsatellite bands with clear bands and high polymorphism in the samples of Taihu Newbait, which is helpful for the analysis of Taihu Newbait from the perspective of genetics Its invasion adaptation mechanism.
Description
technical field [0001] The invention relates to the field of molecular biology, in particular to a primer set for microsatellite DNA molecular markers of Taihu new whitebait. Background technique [0002] Neosalanxtaihuensis of Taihu Lake belongs to Salmoniformes, Silverfish family, and Neosalanx genus. It is an important small-scale economic fish in the middle and lower reaches of the Yangtze River and its affiliated lakes. It mainly lives in the middle and upper layers of open water areas. Cladocera and claw-footed animals are the main food. Because whitebait is easy to freeze, dry, store, transport and sell, it has a broad market at home and abroad and is an excellent green food. [0003] As the most widely transplanted object in my country, Newbait in Taihu Lake has been transplanted to lakes and reservoirs of different sizes in China. Due to its own biological characteristics, it quickly adapts to the local environment and becomes an invasive alien species. [0004] I...
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IPC IPC(8): C12N15/11C12N15/10C12Q1/68
Inventor 刘红艳李存耀熊飞
Owner JIANGHAN UNIVERSITY