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Specific primer and fluorescent quantitative detection kit detecting cattle ADIPOQ gene mRNA expression level

A technology of expression level and real-time fluorescence quantification, applied in the field of genetic engineering, can solve problems such as reducing feed utilization rate and affecting consumer health, and achieve the effect of improving carcass quality, simple and fast quantitative detection, and good stability.

Inactive Publication Date: 2015-10-07
LANZHOU INST OF ANIMAL SCI & VETERINARY PHARMA OF CAAS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In animal production, fat deposition and fat content in animals are important factors affecting meat quality. Excessive fat content in animal products not only affects the health of consumers, but also reduces feed utilization

Method used

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  • Specific primer and fluorescent quantitative detection kit detecting cattle ADIPOQ gene mRNA expression level
  • Specific primer and fluorescent quantitative detection kit detecting cattle ADIPOQ gene mRNA expression level

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Embodiment 1

[0032] The fluorescence quantitative PCR detection kit of ADIPOQ gene mRNA expression level of the present invention is made up of following components:

[0033] 2×SYBR GREEN MIX 5mL;

[0034] Positive control: standard ADIPOQ gene template 500μL;

[0035] Primer mixture 500μL;

[0036] Ultrapure water 5mL.

[0037] Among them, 2×SYBR GREEN MIX consists of the following components: Taq DNA polymerase 0.1 U / μL, dNTPs substrate 0.4 mmol / L, Mg 2+ 5.0 mmol / L, KCl 100 mmol / L, Tris·HCl 20 mmol / L at pH 8.3, 0.02% gelatin and SYBR Green dye.

[0038] Primer mixture: upstream primer 8 μmol / L, downstream primer 8 μmol / L. Wherein, the sequence of the upstream primer is: 5'-AGAAAGGGAGAACCTGGAGA-3',

[0039] The sequence of the downstream primer is: 5'- GGTAAAGCGAATGGGAACA -3'.

[0040] Standard ADIPOQ gene template: the concentration is 0.8 μmol / L, and the ADIPOQ gene template sequence is

[0041] 5'-AGAAAGGGAGAACCTGGAGAAAGTGCCTATGTATACCGCTCAGCATTCAGTGTGGGACTGGAGAGACAGGTCACTGTCCCCAA...

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Abstract

The invention provides a specific primer detecting the cattle ADIPOQ gene mRNA expression level. According to the nucleotide sequence of the specific primer, the upstream primer sequence is 5'-AGAAAGGGAGAACCTGGAGA-3', and the downstream primer sequence is 5'-GGTAAAGCGAATGGGAACA-3'. The invention further provides a corresponding detection kit. The purpose of easily, conveniently and fast detecting the ADIPOQ gene transcription level is achieved, and the mRNA expression state of ADIPOQ genes of different cattle breeds (including dairy cows, cattle and yaks) and different kinds of tissue at different periods can be detected. The specific primer has the advantages of being high in sensitivity, good in stability and low in experiment cost when the mRNA expression level of the ADIPOQ genes is detected.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to specific primers and a fluorescent quantitative PCR detection kit for detecting the expression level of bovine ADIPOQ gene mRNA. Background technique [0002] Polymerase chain reaction (Polymerase Chain Reaction, PCR) is the most commonly used technique in DNA manipulation technology in vitro. PCR technology puts template DNA, specific primers, dNTPs substrates, heat-resistant DNA polymerase, magnesium ions, etc. in the same buffer reaction system, and performs repeated thermal cycles of high-temperature denaturation, low-temperature annealing, and medium-temperature chain extension to achieve target DNA Fragments appear as 2 in the reaction solution n Double amplification (where n is the number of thermal cycles). [0003] Fluorescent quantitative PCR technology is based on ordinary PCR reaction, combined with real-time fluorescence detection technology...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q1/6851C12Q2600/124C12Q2600/158C12Q2531/113C12Q2545/113C12Q2563/107
Inventor 裴杰阎萍包鹏甲郭宪褚敏丁学智冯瑞林梁春年朱新书王宏博
Owner LANZHOU INST OF ANIMAL SCI & VETERINARY PHARMA OF CAAS