Primer, reagent kit and detection method of detecting short spine syndrome of dairy cattle

A syndrome and kit technology, applied in the field of molecular biology, can solve problems such as difficult to meet the needs, cumbersome detection of dairy cow short spine syndrome, etc., and achieve the effect of rapid detection, high sensitivity, and simple reaction procedures

Inactive Publication Date: 2018-06-08
博迪泰(厦门)生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to overcome the deficiencies of the prior art and provide a primer, a kit and a test method for detecting short-spine

Method used

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  • Primer, reagent kit and detection method of detecting short spine syndrome of dairy cattle
  • Primer, reagent kit and detection method of detecting short spine syndrome of dairy cattle
  • Primer, reagent kit and detection method of detecting short spine syndrome of dairy cattle

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1: primer and test kit

[0035] 1. Primer design

[0036] The primers were designed according to the pathogenic gene sequence of respiratory tract infection in the NCBI gene database, referring to the TwistDx instructionmanual and Primer-BLAST primer design principles. The primer length is about 30-35nt. Since there is no primer design software for RPA at present, a large number of primers were designed and synthesized in the previous work of the present invention, and the primers with high sensitivity and good specificity were screened out for use in the present invention (Table 1).

[0037] 2. Primer synthesis

[0038] According to the primer sequences shown in Table 1, sequence synthesis was carried out. It is characterized in that the sequence of the forward primer of the FANC1 gene is shown in SEQ ID NO: 1, modified and added with Biotin; the sequence of the reverse primer is shown in SEQ ID NO: 2, modified and added with DIG. The internal quality con...

Embodiment 2

[0042] Example 2 Detection of Dairy Cow Short Spine Syndrome (FANC1 Gene Deletion)

[0043] The present embodiment adopts the kit described in Example 1 to detect dairy cow short spine syndrome (FANC1 gene deletion). The method used in the detection is the recombinant enzyme polymerase constant temperature gene amplification method combined with lateral flow chromatography. The specific steps are as follows:

[0044] 1. Clinical sample preparation

[0045] Collect dairy cow whole blood samples, and the collection conditions are: positive clinical samples (10 cases) confirmed to have dairy cow short spine syndrome (FANC1 gene deletion) after laboratory inspection (fluorescent quantitative RT-PCR), and confirmed to be No negative clinical samples (10 cases) of short spine syndrome (FANC1 gene deletion). Clinical samples were stored at -80°C.

[0046] 2. Nucleic acid extraction from samples

[0047] In this embodiment, whole blood DNA column chromatography extraction or automa...

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Abstract

The invention discloses a primer, a reagent kit and a detection method of detecting a short spine syndrome of dairy cattle and belongs to the field of molecular biology. The primer comprises a forwardprimer with a nucleotide sequence shown as SEQ ID NO (sequence identifier number): 1, and a reverse primer with a nucleotide sequence shown as SEQ ID NO: 2, wherein the forward primer is labeled withbiotin; and the reverse primer is labeled with digoxin. A recombinase-polymerase nucleic acid amplification technology and a lateral flow chromatographic technology are adopted; isothermal amplification is conducted on a clinical sample after DNA (deoxyribonucleic acid) extraction; a PCR (polymerase chain reaction) instrument is not required for a thermal circulation reaction; and a detection result can be directly read from a lateral flow chromatographic test strip by naked eyes. The primer has strong specificity and high amplification efficiency for detecting the short spine syndrome of thedairy cattle. The reagent kit and the detection method are simple in reaction procedure, can effectively and quickly detect the short spine syndrome of the dairy cattle and are applicable to simple,convenient, quick and accurate diagnosis of the short spine syndrome in a cattle farm.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a primer, a kit and a detection method for detecting short spine syndrome in dairy cows. Background technique [0002] Brachyspina syndrome (BS) was first reported in the Danish Holstein herd in 2006. It is a congenital recessive genetic fatal defect in dairy cows, which can lead to malformed embryos and even death or stillbirth. Short spine syndrome deficient calves may be severely underweight and stunted despite normal or long gestation periods. When stillborn calves are examined, a combination of severely shortened spines, long and thin limbs, and abnormal viscera are found. The molecular genetic mechanism for the formation of this symptom is a deletion mutation (Deletion) of 3329 bp in the FANC1 gene on bovine chromosome 21 (BTA21), resulting in the loss of exons 25, 26, and 27, and at the C-terminus of amino acid 877 The original 451 amino acid peptide chain is replaced by...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/156C12Q2521/507C12Q2522/101C12Q2565/137C12Q2565/625
Inventor 张睿李博安詹尔昌
Owner 博迪泰(厦门)生物科技有限公司
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