天柱番鸭NPY基因克隆及其在组织中的表达分析

doi:10.16431/j.cnki.1671-7236.2019.04.002

摘要/Abstract

摘要：

试验旨在克隆天柱番鸭神经肽Y（neuropeptide Y，NPY）基因，并分析其在天柱番鸭不同组织中的转录水平。以天柱番鸭产蛋期组织混合cDNA为模板，通过RT-PCR扩增天柱番鸭NPY基因的完整CDS区进行克隆测序，并结合生物信息学分析工具分析其同源性及遗传进化关系，同时进行NPY蛋白理化特性、亚细胞定位、信号肽、糖基化与磷酸化位点、二级结构、三级结构等预测，并对NPY基因在天柱番鸭不同组织中的转录水平进行检测。结果表明，天柱番鸭NPY基因CDS区全长294 bp，编码97个氨基酸，核苷酸序列与氨基酸序列同源性比对显示，NPY基因在不同物种间具有一定的遗传多样性，与绿头鸭亲缘关系最近；NPY蛋白为酸性不稳定蛋白，存在1个信号肽（第1-28位氨基酸），亚细胞定位为100%位于细胞外；存在1个功能结构域PAH，同时含有2个O-糖基化和丰富的磷酸化位点，空间结构以α-螺旋和无规则卷曲为主。实时荧光定量PCR结果表明，NPY基因mRNA在天柱番鸭各组织中均有分布，在大脑中表达水平相对较高，在胰腺、腺胃中表达量次之，与其他组织间差异极显著（P<0.01），在肌胃中表达量最低。本试验结果为进一步研究NPY基因在调控家禽能量平衡、生长发育、脂肪沉积、繁殖性能等多种生理功能提供了参考。

关键词: 天柱番鸭; NPY基因; 克隆; 生物信息学分析; 组织表达

Abstract:

This study was aimed to clone neuropeptide Y (NPY) gene in Tianzhu muscovy,and analyze its transcriptional level of different tissues in Tianzhu muscovy.The mixed cDNA of Tianzhu muscovy tissues during lying was used as PCR template,then the entire CDS region of NPY gene was cloned and sequenced by RT-PCR.The homology and relationship of genetic evolution of NPY gene,the physical and chemical properties,subcellular localization,signal peptide,glycosylation site and phosphorylation site,the secondary and tertiary structures of NPY protein were analyzed by bioinformatics tools,and the transcriptional level of NPY gene mRNA in different tissues were detected.The results showed that the whole-length coding region of NPY gene in Tianzhu muscovy was 294 bp,which coded 97 amino acids.The homology alignment results of nucleotide and amino acid sequences showed that there was a certain genetic diversity of NPY gene in different species,and Tianzhu muscovy had a close relationship with Anas platyrhynchos.NPY protein were acidic and unstable protein,which had one signal peptide (1-28 amino acids),and the subcellular localization was 100% outside the cell.In addition,there were two O-glycosylation sites and abundant phosphorylation sites of NPY protein,which existed a functional domain named PAH.Spatial structure of NPY protein were mainly assembled with alpha helix and random coil.Real-time PCR results showed that the NPY gene mRNA were distributed in all tissues,there was the highest expression level of NPY gene mRNA in cerebrum,and there was the secondry highest expression level of NPY gene mRNA in pancreas and glandular stomach,which were extremely significant difference with others tussues (P<0.01),and there was the lowest expression of NPY gene mRNA in gizzard.The results would provide references for further research on the physiological regulatory functions of NPY gene on energy balance,growth and development,fat deposition,reproductive performance in poultry.

Key words: Tianzhu muscovy; NPY gene; cloning; bioinformatics analysis; tissues expression

中图分类号:

谭斌, 杨胜林, 杨汝才, 周璇, 杨世皓, 罗林丽, 祝永才. 天柱番鸭NPY基因克隆及其在组织中的表达分析[J]. 《中国畜牧兽医》, 2019, 46(4): 967-976.

TAN Bin, YANG Shenglin, YANG Rucai, ZHOU Xuan, YANG Shihao, LUO Linli, ZHU Yongcai. Cloning and Expressing Analysis of NPY Gene of Different Tissues in Tianzhu Muscovy[J]. , 2019, 46(4): 967-976.

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