Cloning,Bioinformatics Analysis and Tissue Expression Analysis of GnRHR Gene in Jinghai Yellow Chicken

doi:10.7668/hbnxb.2014.05.009

Abstract

Abstract: Based on the published mRNA nucleotide sequence of Gallus gallus GnRHR gene,two pair of primers were designed to clone the GnRHR gene coding sequence of Jinghai yellow chicken by RT-PCR.A variety of software and online tools were used to analyze the homology among different species,physical and chemical properties,transmembrane region,subcellular localization,hydrophilic,potential phosphorylation locus,conserved domain database,secondary structure and tertiary structure of GnRHR protein.Two pair of primers were designed to detect the tissue expression of GnRHR gene in twelve tissues of Jinghai yellow chicken by RT-qPCR.Finally,GnRHR gene was cloned which contained CDS region,part of promoter region and 3'region.Result of Blast showed that GnRHR gene of Jinghai yellow chicken shared 99.7%,86.7%,55.7%,54.6%,52%,51.6%,50.8%,50%,49.9%,49.6%,49.4%,47.4% and 39.3% identity with Gallus gallus,Cairina moschata,Pantholops hodgsoni,Sus scrofa,horse,mice,rats,rabbits,sheep,cows,chimpanzees and zebrafish.Phylogenetic tree was constructed.Analysis of GnRHR protein structure showed that the molecular weight was 45.432 kDa and pI was 9.55.GnRHR protein was consisted of twenty kinds of amino acids,in which leucine accounted for the highest content of 13.8% and Lysine accounted for the lowest content of 0.7%.The instability index was computed to be 65.35 which classified the protein as unstable.The grand average of hydropathicity was 0.312 which showed that the protein was Non-water-soluble protein.GnRHR protein did not belong to secreted protein,mainly presented on membrane,with no signal peptide and contained 16 phosphorylation sites and 11 glycosylation sites.Analysis of conserved domains showed that GnRHR protein included two low complexity sequences and seven transmembrane segments which agreed with the transmembrane analysis.The secondary structure of GnRHR was mainly composed of random coil.The tertiary structure of domain area of GnRHR protein showed a helix and single strand structure.The result of tissue expression showed that GnRHR was mainly expressed in pituitary.The expression quantity of GnRHR in other eleven tissues was low.

Key words: Jinghai yellow chicken, GnRHR, Cloning, Bioinformatics analysis, Tissue expression

CLC Number:

ZHANG Tao, ZHANG Gen-xi, WANG Jin-yu, FAN Qing-can, WANG Wen-hao, HAN Kun-peng, WANG Yong-juan. Cloning,Bioinformatics Analysis and Tissue Expression Analysis of GnRHR Gene in Jinghai Yellow Chicken[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2014, 29(5): 45-53. doi: 10.7668/hbnxb.2014.05.009.

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