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Paternity test method for goats and microsatellite primer and kit thereof

A paternity test and kit technology, applied in the field of molecular genetics research, can solve problems such as unsuitable Chong white goat paternity test, and achieve the effects of reliable test results, high accuracy and low cost

Inactive Publication Date: 2012-05-23
TONGJI UNIV
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  • Abstract
  • Description
  • Claims
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AI Technical Summary

Problems solved by technology

With the in-depth research on microsatellite markers, it has been introduced into the pedigree identification of cattle and sheep (using microsatellite DNA technology for sheep paternity testing, 2005, Journal of Anhui Agricultural University; using 15 microsatellite loci for parentage of Holstein cows Identification, 2010, Shandong Animal Husbandry and Veterinary Medicine; etc.), Wang Jie et al. used 10 microsatellite loci to carry out paternity identification on plateau Tibetan goats (Application of microsatellite DNA in plateau Tibetan goat paternity identification, 2010, Southwest University for Nationalities Acta Sinica Sinica), but the Chongming white goat belongs to the Yangtze River Delta white goat, which has breed specificity. This system is not suitable for paternity testing of Chongming white goats. Therefore, it is necessary to establish a suitable paternity testing system for Chongming white goats by screening microsatellite markers

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  • Paternity test method for goats and microsatellite primer and kit thereof
  • Paternity test method for goats and microsatellite primer and kit thereof
  • Paternity test method for goats and microsatellite primer and kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Example 1. Screening of microsatellite primers for paternity testing of Chong white goats

[0042] The choice of microsatellite markers

[0043] According to the existing literature and the relevant research results of others (analysis of microsatellite genetic diversity of six local goat breeds in China, 2004; microsatellite analysis of genetic diversity of 10 goat breeds in China, 2008), the Food and Agriculture Organization of the United Nations (FAO ) recommended 58 microsatellite loci, their complete nucleotide sequences, primer sequences, PCR reaction conditions, population polymorphism information, and chromosome genetic map positions can all be obtained from the following website: http: / / www.ncbi. nlm.nih.gov / . All primers were synthesized by Shanghai Jierui Bioengineering Co., Ltd., and 19 pairs of primers with clear amplification bands and good reproducibility were screened out. The specific information is shown in Table 1.

[0044] Table 1 Primer sequence, ...

Embodiment 2

[0102] Embodiment 2, verification of microsatellite detection system

[0103] Theoretically speaking, as long as there is a genotype of a genetic marker that does not conform to the Mendelian law of inheritance, a conclusion can be made to exclude patriarchal relationship. However, there is a possibility of wrong exclusion by detecting DNA polymorphisms to identify paternity. In order to prevent wrong conclusions, the present invention follows the following principles in paternity exclusion: there are 3 or more microsatellite markers that do not conform to Mendelian The law of inheritance makes a conclusion that excludes parent-child relationship.

[0104] 13 father-daughter pairs and 1 unrelated father were selected from the Chong white goat parent-child group. Assuming that the 4 rams (numbers 11♂-14♂, see Table 6) are the hypothetical fathers of the 13 ewes, use the results of the 8 microsatellite marker genotypes screened out in the above experiment to carry out simulated...

Embodiment 3

[0118] Example 3. Chong white goat paternity test kit and its composition

[0119] The above test results show that the paternity identification of Chong white goats can be carried out by using 8 microsatellite markers LSCV043, BMS 1724, BM6526, Oarfcb48, BMS1248, BMS1678, BMS574, and BMS875, and the exclusion probability of the real parents and suspected parents can be greater than 99.9%. The relative chance of paternity is greater than 0.99, and the identification result is more credible. So use these 8 microsatellite primer combinations to make a kit for paternity testing of Chong white goats.

[0120] The kit includes the following reagents:

[0121] (1) Taq DNA polymerase 2.5U / ul

[0122] (2) 10×PCR Buffer (Mg 2+ Free) buffer

[0123] (3) dNTPs25mmol / L

[0124] (4) MgCl 2 Solution 25mmol / L

[0125] (5) Eight microsatellite markers screened in the above experiment: LSCV043, BMS1724, BM6526, Oarfcb48, BMS1248, BMS1678, BMS574, BMS875 were used as special primers for ...

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Abstract

The invention belongs to the technical field of research on molecular genetics and particularly relates to a method for utilizing a microsatellite marking technology to perform a paternity test on goats and a microsatellite primer and a kit thereof. The invention discloses a paternity test method for the goats, which comprises the following steps of: (1) extracting a blood sample from a goat breed and performing DNA (Deoxyribonucleic Acid) extraction and detection; (2) selecting a microsatellite locus: selecting the microsatellite locus with a stable PCR (Polymerase Chain Reaction) augmenting result and a clear strip as well as a polymorphism but without non-specific augmentation; (3) utilizing each microsatellite locus to perform PCR augmentation on different goat DNA templates, and performing electrophoresis detection on augmenting fragments; and (4) performing genetic polymorphism analysis on the microsatellite locus. The invention also discloses the microsatellite primer and the kit used for the paternity test method for the goats. The paternity test method for the goats, provided by the invention, is simple in operation, high in speed, low in cost, and high in accuracy.

Description

technical field [0001] The invention belongs to the technical field of molecular genetics research, and in particular relates to a method for paternity testing of goats using microsatellite marker technology, as well as a microsatellite primer and a kit. Background technique [0002] In modern animal breeding, accurate pedigree records are very important due to the use of phenotypic information of various relatives. However, in some cases, people cannot accurately identify the father of certain goat individuals. Practical examples include Inaccurate pedigree records are likely to lead to inaccurate pedigree records of lambs from ewes bred with mixed semen and from ewes crossed with multiple rams. slow. Therefore, it is more important to confirm the pedigree of the breeding ram. Therefore, paternity testing is required to correct errors in pedigree records and ensure as much genetic progress as possible. At present, the effectiveness and accuracy of goat paternity identifi...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 陈士超刘志学罗倩倩王海鹏杨姣郑佩培
Owner TONGJI UNIV
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