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Microsatellite DNA markers for Trionyx sinensis

A technology of DNA markers and microsatellite markers, which is applied in the direction of recombinant DNA technology, DNA/RNA fragments, microbial determination/inspection, etc., and can solve the problem of not providing enough genetic information

Inactive Publication Date: 2011-03-09
ANHUI NORMAL UNIV
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AI Technical Summary

Problems solved by technology

Some scholars have used different molecular marker techniques (including mitochondrial cytochrome b gene sequence, 12S rRNA, RAPD, etc.) to study the genetic diversity of soft-shelled turtles, but the above genetic markers cannot provide sufficient genetic information, such as Zhang Yongzheng et al., Zhonghua Sequence variation and population genetic structure analysis of cytochrome b gene in five different geographical populations of soft-shelled turtle (Pelodiscus sinensis); Wu Ping, Sequence analysis and phylogenetic study of 12S rRNA gene fragments in Asian freshwater and terrestrial turtles; Xiao Yamei et al., Chinese soft-shelled turtle Population RAPD analysis

Method used

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  • Microsatellite DNA markers for Trionyx sinensis
  • Microsatellite DNA markers for Trionyx sinensis
  • Microsatellite DNA markers for Trionyx sinensis

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Embodiment Construction

[0025] The present invention will be described in further detail below in conjunction with the accompanying drawings.

[0026] 1. Construction of soft-shelled turtle microsatellite DNA enrichment library:

[0027] 1.1 Genome extraction and enzyme digestion: extract the Chinese soft-shelled turtle genome DNA with reference to the method of Sambrook et al. Electrophoresis was performed on 1% agarose gel, a DNA fragment of 400-1000 bp in size was excised under ultraviolet light, and the recovered enzyme-digested product was purified with a gel extraction kit (purchased from Shanghai Sangong).

[0028] 1.2 Add connector:

[0029] Add the two ends of the purified and recovered enzyme-cut product to the adapter:

[0030] SauL-F(5'-GGCCAGAGACCCCCAAGCTTCG-3')

[0031] SauL-R (5'-PO 4 -GATCCGAAGCTTGGGGTCTCTGGCC-3'),

[0032] Under the action of T4 ligase (purchased from Promega), the ligation was performed overnight in a water bath at 16°C.

[0033] 1.3PCR detection adapter conne...

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Abstract

The invention discloses microsatellite DNA markers for Trionyx sinensis, which is characterized in that an enriched library of microsatellites (CA)n, (AAC)n, (ACAG)n and (GATA)n from the Trionyx sinensis is constructed, and positive clones of microsatellite sequences are screened and sequenced to obtain 63 clones containing microsatellite repetitive sequences, thus determining 15 microsatellite markers with rich polymorphism, namely, PSW01, PSW02, PSW03, PSW04, PSW05, PSW06, PSW07, PSW08, PSW09, PSW10, PSW11, PSW12, PSW13, PSW14 and PSW15. The microsatellite DNA markers for the Trionyx sinensis of the invention can be applied to researching genetic diversity and paternity test for the Trionyx sinensis in different geographical populations, have the advantage of good repeatability, and are reliable and effective molecular markers.

Description

technical field [0001] The invention relates to a molecular marker technology, in particular to a Chinese soft-shelled turtle microsatellite DNA genetic marker. Background technique [0002] Microsatellite DNA, also known as short tandem repeats (STR), simple repeats (SSR), simple sequence length polymorphism (SSLP), refers to the tandem repeats of 1-6 nucleotides in the genome. Nucleotide sequence; among them, the most common are dinucleotide repeats, such as ((AC / TG)n and (AG / TC)n. Different organisms show polymorphism due to the number of repetitions of the repeat motif Studies have shown that it may be the "strand slippage" phenomenon in the DNA replication process that causes the high polymorphic information content (PIC) of microsatellite DNA. The high mutation and co-dominance of microsatellite DNA Expression and its ubiquity in eukaryotic genomes make it a superior molecular marker for the analysis of population genetic diversity, identification of kinship, genome m...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/11C12Q1/68
Inventor 聂刘旺卜兴江刘罗
Owner ANHUI NORMAL UNIV
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