杨树HSP90基因的克隆及生物信息学分析

Cloning and Bioinformation Analysis of HSP90 Gene of Poplar

【目的】为了揭示热激蛋白90(Heat shock protein 90,HSP90)基因在杨树抗溃疡病中的功能,克隆获得毛白杨HSP90基因的全长序列,以期明确HSP90基因的表达与溃疡病菌侵染间的关系。【方法】采用RT-PCR技术和Gateway克隆的方法,获得了毛白杨的HSP90基因序列,并利用相关软件对该基因编码的蛋白的理化性质、疏水性、结构域、功能及亚细胞定位等进行了生物信息学分析。【结果】毛白杨的HSP90基因序列(NCBI登录号:AGU99972.1)全长为2 100 bp,其中A+T占52.90%,C+G占47.10%,共编码699个氨基酸。毛白杨HSP90基因的生物信息学分析结果显示,该基因编码的蛋白相对分子质量为80.08 kDa,理论等电点为4.96。毛白杨HSP90蛋白主要位于细胞核和细胞质膜中,且在N端含有1个基因保守结构域HATPase_c,可与ATP结合,具有内源ATPase活性,属于HSP90家族,为亲水性蛋白,可能具有离子通道的功能。同时,对毛白杨的HSP90基因进行系统进化分析,发现毛白杨的HSP90基因与毛果杨(gi 539331543)、大豆(gi 358248990和gi 356552478)的亲缘关系较近,而与毛果杨(gi 224124864)的亲缘关系相对较远,说明同一物种的HSP90进化可能有所不同。【结论】首次从毛白杨中成功克隆得到与抗溃疡病相关的HSP90基因,并对其序列和生物学信息进行了分析,为下一步研究HSP90基因在杨树抗溃疡病中的功能奠定基础,也为进一步研究HSP90基因在杨树抗逆胁迫中的生理功能提供了理论支持。

【Objective】In order to reveal the function of heat shock protein 90(HSP90)gene in poplar anti-ulcerative disease,the sequence of HSP90 gene of Populus tomentosa was cloned to clarify the relationship between the expression of HSP90 gene and the infection of canker.【Method】The HSP90 gene sequence of P. tomentosa was obtained by RT-PCR and Gateway cloing. Meanwhile,the physicochemical properties,hydrophobicity,domain,function and subcellular localization of the protein encoded by the gene were obtained by bioinformatics analysis.【Result】The results showed that the sequence of HSP90 gene was 2100 bp(gene accession number: AGU99972.1),A+T was 52.90% and C+G was 47.10%,which encoded 699 amino acids. Bioinformatics analysis of HSP90 gene in P. tomentosa showed that the HSP90 gene encoded a protein with a relative molecular mass of 80.08 kDa and a theoretical isoelectric point of 4.96. Besides,the protein was located in the nucleus and plasma membrane,and contained a gene conserved domain HATPase_c at the N-terminus,which had endogenous ATPase activity to bind ATP. It belonged to the HSP90 family,which was a hydrophilic protein. The function of the protein may be ion channels. Meanwhile,phylogenetic analysis found that the HSP90 gene of P. tomentosa was closely related to P. trichocarpa(gi 539331543)and Glycine max(gi 358248990 and gi 356552478),but it was far to P. trichocarpa(gi224124864),which indicated that the evolution of HSP90 in the same species may be different.【Conclusion】In this studythe HSP90 gene was cloned from P. tomentosa for the first time and was analyzed by bioinformatics. The results provide a basis for the further study of the function of the HSP90 gene in poplar canker resistant and theoretical support to investigate the function of HSP90 gene in poplar stress resistance.