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Single nucleotide polymorphism (SNP) molecular markers and method for pork DNA tracing by high resolution melting (HRM) method

A technology of molecular markers and pork, which is applied in the direction of recombinant DNA technology, DNA/RNA fragments, biochemical equipment and methods, etc., can solve the problems of long detection cycle, complex operation and high cost of pork DNA traceability methods, and achieve low detection cost Effect

Active Publication Date: 2014-11-19
NANJING AGRICULTURAL UNIVERSITY
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AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to solve the problems of complex operation, long detection period and high cost in the pork DNA traceability method in the prior art, and detect the SNPs molecular markers from pork by the HRM method, that is, the high-resolution melting curve analysis method, to achieve rapid and accurate detection purpose of pork source

Method used

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  • Single nucleotide polymorphism (SNP) molecular markers and method for pork DNA tracing by high resolution melting (HRM) method
  • Single nucleotide polymorphism (SNP) molecular markers and method for pork DNA tracing by high resolution melting (HRM) method
  • Single nucleotide polymorphism (SNP) molecular markers and method for pork DNA tracing by high resolution melting (HRM) method

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Embodiment 1

[0065] (1) Selection of SNPs sites

[0066] Select SNPs sites from NCBI to ensure that ① the distance between any adjacent SNPs is ≥ 1Mb, and that the SNPs are independent of each other and that there is no linkage; ② there is no relationship between any SNPs and other molecular markers; ③ the flanking sequences of SNPs are not Any variation exists. The SNPs selected in this study are listed in Table 1.

[0067] Table 1 SNP site related information

[0068]

[0069] As shown in Table 1, among the selected 10 SNP sites, SNP1 is located on chromosome 1 of the pig genome, SNP2 is located on chromosome 2 of the pig genome, SNP3 is located on chromosome 12 of the pig genome, SNP4 is located on chromosome 14 of the pig genome, and SNP5 is located on chromosome 14 of the pig genome On chromosome 3, SNP6 is located on chromosome 13 of the pig genome, SNP7 is located on chromosome 17 of the pig genome, SNP8 is located on chromosome 8 of the pig genome, SNP9 is located on chromosom...

Embodiment 2

[0102] 1) Pig DNA sample preparation

[0103] Select the fresh pork individuals of No. 1, 2, 5, 6, 30, 31, 40, 48, 49, and No. 53 in the above-mentioned 299 samples. When the pig muscle tissue and blood were collected in Example 1, 10 pigs were selected here. Cut 10 pig skin samples with pig hair from the individual, record the individual number of each sample, mark the samples, and shuffle the samples to be tested.

[0104] Extract 15 pig hairs with complete hair follicles from the pigskin of each sample, cut as short as possible only the hair follicles, put them into a clean 1.5ml centrifuge tube, add 200ul of DNA lysate and 10ul of proteinase K, and mix well . Digest at 56°C for 2 hours, during which the samples were flicked with fingers to speed up the digestion process. Add 60ul of 5mol / L NaCl and mix thoroughly; add 260ul of saturated phenol / chloroform / isoamyl alcohol (25:24:1), mix by inversion for 5min to fully denature the protein, centrifuge at 1000r / min for 5min, ...

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Abstract

The invention belongs to the technical field of foods, provides single nucleotide polymorphism (SNP) molecular markers for pork DNA tracing and a method for pork DNA tracing by a high resolution melting (HRM) method. The SNP molecular markers comprise 10 SNPs molecular markers which are mutually independent, have gene heterogeneity at least 0.4 and are suitable as pork DNA tracing markers. The 10 SNPs molecular markers are obtained by a HRM analysis method, the effective SNPs sites form a pork SNPs information group and a pork DNA tracing process is carried out. The method for pork DNA tracing by the HRM method based on the SNP molecular markers has the characteristics of simpleness, speediness and low cost. The 10 SNPs molecular markers stably and widely exist in the pig genome DNA and are conducive to large-scale popularization of the DNA tracing technology.

Description

technical field [0001] The invention belongs to the technical field of food, and relates to a SNP molecular marker for traceability of pork DNA, that is, a single nucleotide polymorphism marker, in particular to a method for traceability of pork DNA by using the HRM method, that is, a high-resolution melting curve analysis method. Background technique [0002] With the improvement of people's living standards, the safety of food has attracted more and more attention. As the main source of protein in human food, meat products have naturally attracted much attention to their safety. Traceability technology has increasingly become an important means of meat safety assurance and supervision. [0003] There are many technical methods for meat food traceability, such as label traceability technology, iris traceability technology and DNA traceability technology. Compared with other traceability technologies, DNA traceability technology has the advantages of easy typing and large-s...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12Q1/68
Inventor 周光宏杨萨萨李春保徐幸莲吴洽宇
Owner NANJING AGRICULTURAL UNIVERSITY
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