Method for sifting Holstein cow BLAD carrier and special primer and reagent case

A technology of carriers and kits, applied in the field of screening BLAD carriers of Holstein cows and their special primers and kits

Inactive Publication Date: 2007-09-26
CHINA AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0009] The advantage of PCR-RFLP is that it overcomes the defect that the standard RFLP analysis requires a large amount of DNA, and can study a small amount of DNA samples, and the products amplified by...

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  • Method for sifting Holstein cow BLAD carrier and special primer and reagent case
  • Method for sifting Holstein cow BLAD carrier and special primer and reagent case
  • Method for sifting Holstein cow BLAD carrier and special primer and reagent case

Examples

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Embodiment 1

[0028] Example 1. Screening Holstein cows for BLAD carriers and preparation of special kits thereof

[0029] 1. Design of primers for screening BLAD carriers in Holstein cows

[0030] The CD18 gene of BLAD-affected cattle or carriers is mutated from A to G at the 1200th base at the 5' end, and Taq I restriction endonuclease specifically recognizes T*CGA. When this site is mutated, the sequence is changed from T* CGA becomes TCGG, and the Taq I recognition site disappears. According to this principle, refer to the partial genome sequence of the cow CD18 gene published by Kriegesmann, B. (Kriegesmann B., Jansen S., Baumgartner B.G. and Brenig B. Partialgenomic structure of the bovine CD18 gene and the refinement of test of bovineleukocyte adhesion deficiency [J]. J. Dairy. Sci. 1997, 80: 2547-2549.), using Oligo6.0 software at its 1200th A / G point mutation site A pair of primers for PCR amplification of the nucleotide fragment containing the BLAD-causing mutation site were desi...

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Abstract

The invention discloses a sieving method of hesitant cow BLAD carrier and specific primer and agent box, wherein the primer is composed of primer couple with nucleic acid sequence in the list 1 and list 2. The sieving method comprises the following steps: adopting detected cow genome DNA as mold; proceeding PCR augumentation guided by primer; recycling PCR augumented segment; using restriction enzyme TaqI to cut the segment; detecting the segment at 193bp and 131bp as normal cow and 324bp, 193bp and 131bp DNA as BLAD carrier and 324bp DNA as BLAD ill cow; fitting for selecting superior cow and detected one with BLAD gene effectively.

Description

technical field [0001] The invention relates to a method for screening BLAD carriers of Holstein cows by using PCR-RFLP technology, as well as special primers and a kit. Background technique [0002] Bovine leukocyte adhesion deficiency (Bovine leukocyte adhesion deficiency, BLAD) is an autosomal recessive genetic disease controlled by a single gene in Holstein cows. Caused by point mutations (Shuster D.E., Bosworth B.T., Kehrli M.E.Jr. Sequence of the bovine CD18-encoding cDNA: comparison with the human and murine glycoproteins. Gene. 1992, 114(2): 267-271.). The CD18 gene has been located on chromosome 1. Shuster et al. analyzed the cDNA sequence of the CD18 gene of a Holstein cow suffering from BLAD, and found that the 383rd base from the 5' end was mutated from A to G. This mutation was a missense mutation, resulting in a Tianmen Aspartic acid is mutated to glycine (D128G), causing β on the surface of leukocytes 2 Significant reduction or lack of integrin expression c...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 张沅孙东晓孙艺
Owner CHINA AGRI UNIV
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