马鹿β-防御素-1cDNA全长克

发布时间：2018-04-15 16:42

本文选题：马鹿β-防御素- + RACE　；参考：《畜牧兽医学报》2017年02期

【摘要】：为了研究马鹿β-防御素-1(Red deerβ-defensin-1,redBD-1)基因的结构与功能,揭示该基因的组织表达规律。本研究利用PCR结合RACE(Rapid-amplification of cDNA ends)技术从马鹿舌黏膜中克隆redBD-1基因的cDNA全长序列并对其进行了生物信息学分析,同时采用Real-time quantitative PCR(RT-qPCR)技术检测该基因在各组织的表达情况。结果表明,redBD-1基因的cDNA全长序列为455bp,开放阅读框(ORF)为192bp,编码64个氨基酸。生物信息学分析表明,redBD-1蛋白的理论分子量为6.94ku,有10个带正电荷的氨基酸残基,无带负电荷的氨基酸残基,理论等电点为10.85。预测redBD-1蛋白有一个分泌信号肽结构,无跨膜区,主要在细胞外发挥生理功能;6个保守的半胱氨酸残基分别以Cys1-Cys5、Cys2-Cys4和Cys3-Cys6连接形成3个分子内二硫键;成熟蛋白的三级结构是由β-折叠、延伸和无规则卷曲构成。redBD-1基因编码的氨基酸序列同源性最高的是梅花鹿β-防御素(siBD-1)为98.4%,其次是水牛肠β-防御素(BEBD)为92.2%,与人β-防御素-2(HBD-2)同源性最低仅为35.9%。RT-qPCR结果得出,redBD-1在被检器官中均有表达,在消化系统、呼吸系统以及生殖系统的大部分器官表达量较高,肝、肾和脾等实质性器官表达量相对较低。本试验为今后深入研究防御素基因功能以及马鹿黏膜免疫系统提供理论依据。
[Abstract]:In order to study the structure and function of the 尾 -defensin-1 red deer 尾 -defensin-1 (BD-1) gene of Wapiti, the expression pattern of the gene was revealed.In this study, the full-length cDNA sequence of the redBD-1 gene was cloned from the tongue mucosa of Wapiti by PCR and RACE(Rapid-amplification of cDNA ends, and its bioinformatics analysis was carried out. The expression of the redBD-1 gene in various tissues was detected by Real-time quantitative PCRRT-qPCR.The results showed that the full-length cDNA sequence of the BD-1 gene was 455bpand the ORF of ORAF was 192bpencoding 64 amino acids.Bioinformatics analysis showed that the theoretical molecular weight of BD-1 protein was 6.94ku. there were 10 positively charged amino acid residues and no negative charge amino acid residues, and the theoretical isoelectric point was 10.85.It was predicted that the redBD-1 protein had a secretory signal peptide structure with no transmembrane domain and mainly played a physiological function outside the cell, and that six conserved cysteine residues were joined by Cys1-Cys5Cys2-Cys4 and Cys3-Cys6 to form three intramolecular disulfide bonds.The tertiary structure of a mature protein is a 尾 -fold.The highest homology of amino acid sequence of the. RedBD-1 gene encoding by extension and irregular curl is 98.4% for sibbd 1, 92. 2 for buffalo intestinal 尾 -defensin, and 92. 2 for human 尾 -defensin-2HbD-2. The result shows that the lowest homology with human 尾 -defensin-2 HBD-2 is only 35.9%.RT-qPCR.The expression of redBD-1 was found in all the tested organs.In digestive system, respiratory system and reproductive system, the expression of most organs is higher, but the expression of parenchymal organs such as liver, kidney and spleen is relatively low.This study provides a theoretical basis for further study of defensin gene function and mucosal immune system of Wapiti.
【作者单位】：内蒙古农业大学兽医学院;农业部动物疾病临床诊疗技术重点实验室;
【基金】：内蒙古教育厅人才基金
【分类号】：S825

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