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Molecular marker for detecting DNA methylation of cow mastitis resistance

A technology of molecular markers and methylation, applied in the direction of recombinant DNA technology, DNA/RNA fragments, microbial determination/inspection, etc., can solve problems such as few reports

Inactive Publication Date: 2015-04-29
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, pyrosequencing has been widely reported in human disease detection, but rarely reported in traditional animal disease research

Method used

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  • Molecular marker for detecting DNA methylation of cow mastitis resistance
  • Molecular marker for detecting DNA methylation of cow mastitis resistance
  • Molecular marker for detecting DNA methylation of cow mastitis resistance

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Example 1 Detection of DNA Methylation Molecular Marker Methylation Levels Related to Chinese Holstein Mastitis Resistance

[0028] 1.1 Extraction of genomic DNA from the blood of Chinese Holstein cows to be tested

[0029] The blood of Chinese Holstein cows to be tested is collected from the tail vein, and placed at room temperature for 3-4 hours until the blood coagulates. At this time, the blood is divided into two parts: serum and clot. The serum is clear yellow, and the clot is dark red. Contains DNA, present in blood clots.

[0030] Use Tiangen Blood / Cell / Tissue Genomic DNA Sample Extraction Kit to extract genomic DNA from blood clots, the specific steps are as follows:

[0031] Use ophthalmic scissors to cut 0.2-0.3mL of clot into a sterilized 2mL round bottom centrifuge tube, add 500μL of cell lysate CL;

[0032] Fully homogenize the blood clot with a hand-held homogenizer, shake for 15s, centrifuge at 12000rpm for 1min, and discard the dark red supernatant in...

Embodiment 2

[0072] Example 2 Association analysis and detection application of methylation status of TRAPPC9 gene promoter region and mastitis resistance

[0073] According to the method of Example 1, 152 Chinese Holstein dairy cows collected from different dairy farms in Beijing were tested for methylation status, and the test results were as follows: figure 1 As shown, it can be seen that each CpG site has a qualitative variation between methylation and unmethylation.

[0074] Using the GLM process of SAS 9.0 software to analyze the relationship between the methylation of each CpG site and the mastitis resistance of Chinese Holstein cattle, the adopted model is as follows:

[0075] y=μ+hys+p+m+e

[0076] Among them, y is the phenotype value, μ is the population mean of the phenotype value, hys is the field year-season effect, p is the parity effect, m is the methylation effect, and e is the random residual effect. The analysis results are shown in Table 1.

[0077] Table 1 Correlatio...

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Abstract

The invention provides a molecular marker for detecting DNA methylation of cow mastitis resistance and a primer pair of the molecular marker. According to an amplification primer pair and a sequencing primer pair designed for the molecular marker, the DNA methylation of cow mastitis resistance can be detected. The detecting method comprises the following steps: treating the genome DNA of a cow to be detected through bisulfite, performing PCR amplification on a TRAPPC9 gene by using the amplification primer pair, and sequencing by using the sequencing primer pair and a pyrophosphoric acid instrument, so as to obtain the methylation level of a TRAPPC9 gene promotor region of the cow to be detected. According to the invention, a theory basis and a hereditary basis are provided for molecular breeding of cows with mastitis resistance.

Description

technical field [0001] The invention relates to a molecular marker, in particular to a molecular marker for detecting DNA methylation. Background technique [0002] Cow mastitis is one of the main diseases affecting the development of the dairy industry in the world, and it is also the most important disease causing the loss of dairy farming. Cow mastitis will not only affect the milk production of dairy cows and reduce the quality of raw milk, but also affect the normal physiological functions of dairy cows, increasing the abortion rate and culling rate of cows. The heritability of cow mastitis is low, only between 0.02-0.04, and the selection effect of traditional breeding methods is not obvious. With the rapid development of molecular genetics, quantitative genetics and molecular biology techniques, molecular breeding techniques have been gradually applied to livestock and poultry breeding, and achieved certain results. Dairy cow mastitis is greatly affected by the envi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q1/6858C12Q1/6869C12Q2600/124C12Q2600/154C12Q2523/125C12Q2565/301
Inventor 俞英董易春王晓
Owner CHINA AGRI UNIV
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