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Primers and detection method for detecting the expression level of 5-ht receptor gene related to pork quality

A 5-HT and receptor gene technology, applied in the field of agricultural biology, can solve problems such as difficult to achieve accurate detection, and achieve the effect of improving detection accuracy, high detection efficiency, and good repeatability

Active Publication Date: 2015-09-16
HANGZHOU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, 5-HT 2A , 5-HT 7 The receptor has been detected in highly expressed tissues such as the uterus and gastrointestinal tract, and it has been found that as an excitatory receptor, it has the effect of contracting the uterus, but it is difficult to achieve accurate detection in tissues with low expression such as muscle (Sumner et al .,2008; Nakamura et al.,2008)

Method used

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  • Primers and detection method for detecting the expression level of 5-ht receptor gene related to pork quality
  • Primers and detection method for detecting the expression level of 5-ht receptor gene related to pork quality
  • Primers and detection method for detecting the expression level of 5-ht receptor gene related to pork quality

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1 5-HT receptor gene detection method and its optimization

[0052] 1 Experimental instruments and materials

[0053] Real-time fluorescent quantitative PCR instrument (Bio-Rad OPTICON2, the United States); ultra-low temperature refrigerator (Thermo, the United States); high-speed refrigerated centrifuge (Eppendorf5417R, Germany); gradient PCR instrument (ABI PCR system9700, the United States); adjustable micropipette ( Eppendorf, Germany); GEL EQ electrophoresis gel imaging system (Bio-Rad, USA); UV-Vis spectrophotometer (BioSpec-nano, Japan).

[0054] 2 RNA extraction and reverse transcription

[0055] Total RNA was extracted from each tissue by Trizol method. RNA samples were detected by 1.4% agarose-formaldehyde denaturing gel electrophoresis and ultraviolet spectrophotometer to ensure reliable quality of RNA samples, and the total RNA concentration of samples was calculated. Take 2-4 μg of total RNA, and synthesize the first strand of cDNA according to t...

Embodiment 25

[0078] The expression level of embodiment 25-HT receptor gene in different pig species, different tissues

[0079] According to the method of embodiment 1, respectively detect Chun'an flower pig (fat type Chinese local pig breed, n=12), outer Sanyuan pig (lean meat type fast large commodity pig, n=12) in fat and muscle tissue 5 -HT receptor gene expression level, the results are shown in Table 4. It can be seen from Table 4 that 5-HT 2A , 5-HT 7 The expression of receptor gene in adipose tissue was significantly higher than that in muscle tissue (P2A , 5-HT 7 The expression level in the muscle of Wai Sanyuan pig was significantly lower than that in Chun'an Hua pig (P<0.05), while the expression in fat was opposite, but the difference was not significant.

[0080] Table 4 5-HT receptor gene expression in each tissue

[0081]

[0082] Note: * indicates significant difference between groups (P<0.05).

[0083] See Table 5 for the slaughter and meat quality performance inde...

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Abstract

The invention discloses primers for detecting 5-HT receptor gene expression quantity related to pork quality. The primers include (1) a reference gene beta-actin amplification primers: Forward: 5'-TCTGGCACCACACCTTCT-3'; Reverse: 5'-TGATCTGGGTCATCTTCTCAC-3'; (2) target gene 5-HT amplification primers: 5-HT2A: Forward: 5'-AGGTGCTGGGCATAGTCTT-3'; Reverse: 5'-GACGGCTGAGGAGAGGTAAC-3'; 5-HT7: Forward: 5'-CAGATCAACTACGGCAGAGC-3'; Reverse: 5'-GCAGACCGAAATCACCACCAG-3'. The invention further provides a kit and fluorogenic quantitative PCR method for detecting the 5-HT receptor gene expression quantity related to the pork quality. By adopting the primers and the fluorogenic quantitative PCR detection method based on different template concentrations, pig 5-HT receptor gene mRNA expression quantities in different tissues can be detected accurately.

Description

technical field [0001] The invention relates to a method for detecting the expression level of a 5-HT receptor gene related to pork quality, belonging to the field of agricultural biotechnology. Background technique [0002] 5-hydroxytryptamine receptor (5-hydroxytryptamine receptor) is a kind of transmembrane protein combined with 5-hydroxytryptamine (5-hydroxytryptamine, 5-HT) on the cell membrane, belonging to the G protein-coupled receptor superfamily. It includes 5-HT 1 ~5-HT 7 There are 7 types of receptors, and most types can be subdivided into several subtypes. As the first messenger, 5-HT activates or inhibits the activity of transmembrane protein-adenylate cyclase (AC) after binding to the corresponding receptor, thereby increasing or decreasing the intracellular cyclic adenylate (cAMP, second Messenger) concentration, and then affect protein, sugar and fat metabolism, play its physiological role. After that, 5-HT is transported into the cell through the 5-hydr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6851C12Q2531/113C12Q2545/101C12Q2563/107
Inventor 王欢欢章学东李庆海楼立峰张雷
Owner HANGZHOU ACAD OF AGRI SCI
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