绿尾虹雉(Lophophorus lhuysii)Toll样受体5基因的克隆和适应性进化分析

Cloning and Adaptive Evolution Analysis of Toll-Like Receptor 5 Gene in Chinese Monal(Lophophorus lhuysii)

【目的】获得绿尾虹雉(Lophophorus lhuysii)Toll样受体5基因(TLR5)编码区序列,并对其编码区蛋白的特征及适应性进化进行分析。【方法】以绿尾虹雉基因组DNA为模板,采用PCR扩增克隆获得TLR5基因编码区序列。使用MEGA、PAML等软件对其序列特征及选择压力进行分析。【结果】绿尾虹雉TLR5基因编码区序列全长2 583 bp,共编码860个氨基酸,以亮氨酸含量最高(133/860),色氨酸含量最低(11/860)。编码蛋白为典型的Ⅰ型跨膜结构,包括富含LRRs结构域的胞外区、跨膜区和胞内TIR结构域。绿尾虹雉TLR5基因极其保守,与雉鸡(P. colchicus)的同源性最高(97.75%),与小白鹭(E. garzetta)同源性最低(85.17%)。采用位点-特异模型在胞外区共检测到3个正选择氨基酸位点(263F, 280K和645I)。【结论】绿尾虹雉TLR5基因编码区受到较强的纯净化压力,胞外区LRRs结构域可能为识别病原微生物的区域。

【Objective】The objective of this study was to obtain the full coding DNA sequence (CDS) and to explore the mechanisms for the adaptive evolution of toll-like receptor 5 (TLR5) gene in the Chinese monal (Lophophorus lhuysii).【Method】The Chinese monal genomic DNA was used as a tem-plate to amplify and clone the TLR5 gene by PCR. The sequences characterization and adaptive evolu-tion of TLR5 gene encoded protein were analyzed by MEGA and PAML software.【Result】The results showed that the full coding DNA sequence (CDS) of Chinese monal TLR5 was 2 583 bp,encoding 860 amino acids, among which leucine (133/860) occupied the highest content and tryptophan (11/860) with the lowest content. The encoded protein of TLR5 was a typical type I transmembrane receptors, which were composed of a extracellular leucine-rich repeats (LRRs),a transmembrane domain, and a cyto-plasmic Toll/interleukin-1 receptor (TIR) signaling domain, and the entire coding area is subject to strong purification selection. Homology analysis showed that the TLR5 gene of Chinese monal had rela-tively conservative, and the closest relationship with Phasianus colchicus (97.75%),the lowest relation-ship with Egretta garzetta (85.17%). Additionally, three amino acid sites (263F,280K and 645I) were found in the extracellular domain under positive selection by site-specific model of PAML software.【Conclusion】The coding regions of the Chinese monal TLR5 gene were subject to strong purification pressures. It is speculated that the extracellular LRRs domain of the Chinese monal TLR5 gene may play an important role for identifying pathogenic microorganisms.