摘要： 基于物种同源性,应用PCR和RT-PCR技术,克隆得到藏山羊瓣胃溶菌酶基因cDNA编码序列,并命名为TGOLyz,应用生物信息学软件对TGOLyz基因核苷酸序列及预测的蛋白序列进行了分析,并预测该蛋白的三级结构;应用qRT-PCR技术检测了TGOLyz在5个组织中的定量表达。结果表明,藏山羊瓣胃溶菌酶基因TGOLyz cDNA编码区长444 bp(Accession No.KC558501),编码147个氨基酸,分子量为16.29 kDa,等电点为6.08。同源性对比显示,TGOLyz与牛胃1(Cow stomach 1)的同源性最高,达95.24%,与牦牛胃(Yak stomach)的同源性最低,仅为67.61%;系统进化树显示,该基因属于c型溶菌酶(c-type lysozyme)家族;qRT-PCR分析表明,TGOLyz基因在5个组织中均有表达,在瓣胃中表达量最高。这为进一步深入研究TGOLyz的抗菌及消化功能的分子调控机制提供了理论基础。
关键词:
藏山羊,
TGOLyz基因,
克隆,
定量表达,
生物信息学
Abstract: Based on species homology，the lysozyme cDNA was cloned from omasum in Tibetan goat by PCR and RT-PCR，and named TGOLyz gene． The sequence of nucleotides and deduced amino acids of TGOLyz was analyzed through bioinformatics methods，and the gene expression levels at five tissues of Tibetan goat were detected through qRT-PCR． The result showed that the coding region of the TGOLyz cDNA was 444 bp，and encoded 147 amino acids with a molecular weight of 16.29 kDa and isoelectric point of 6.08． The deduced amino acids sequence of TGOLyz shared 95.24% homologies with that from cow stomach 1 and had 67.61% homologies with that from yak stomach． The phylogenetic analysis showed that the TGOLyz gene belongs to c-type lysozyme． Real-time quantitative PCR assay demonstrated that the TGOLyz could be detected in all 5 tissues with the highest expression level observed in omasum． All these would establish a foundation for TGOLyz of its molecular mechanism of antibacterial and digestive function．
Key words:
Tibetan goat,
TGOLyz gene,
Clone,
qRT-PCR,
Bioinformatics
中图分类号:
张鹏, 江明锋, 马晓瑞, 方毅, 王永. 藏山羊瓣胃溶菌酶基因TGOLyz的克隆及组织表达分析[J]. 华北农学报, 2013, 28(3): 35-42. doi: 10.3969/j.issn.1000-7091.2013.03.007.
Zhang Peng, Jiang Mingfeng, Ma Xiaorui, Fang Yi, Wang Yong. Cloning and Expression Analysis of Omasum Lysozyme Gene(TGOLyz) in Tibetan Goat[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2013, 28(3): 35-42. doi: 10.3969/j.issn.1000-7091.2013.03.007.