不同镉积累基因型烟草CAX2基因克隆及序列分析

摘要

Objective]The present study was conducted to clone and analyze the cation exchanger gene (CAX2) from two nicotiana genotypes in order to provide references for CAX2 function and mechanism in cadmium transportation of nicotiana . [Method]Based on CAX2 sequence of potato in NCBI and EST sequences of Nicotiana the primers CAX2-F and CAX2-R were designed to clone CAX2 gene from K326 (Nicotiana tabacum) and Nicotiana. rustica by PCR technique. The sequence of CAX2 was analyzed by bioinformatics methods. [Result]Both of CAX2 coding region in K326 and N. rustica (named as NtCAX2 and NrCAX2) were 1314 bp in length, their nucleotide sequences shared with 98.0% of homology. Their deduced amino acids were 437 aa long with only existing 5 residues differences. Nt-CAX2 and NrCAX2 proteins shared with 98.9% of homology, had two identical Na+-Ca2+ exchange protein domains in 102-249 aa and 293-426 aa regions, and were characterized by common structure characteristics of CAX family. The theoretical pI of NtCAX2 and NrCAX2 were 5.54 and 5.45, respectively. Both of them had 11 transmembrane spans with obvious hydrophobicity and two repeat regions of α-1 and α-2, and richened in alpha helix. Phylogenetic analysis results found that , NtCAX2 and NrCAX2 had the closest relationship to potato (StCAX2) , and their protein homolo-gies were higher than 92 . 0%. [Conclusion]The homology of CAX2 gene was very high in N. tabacum and N. rustica with different Cd accumulating pattern. The CAX2 gene was highly conserved in Solanaceae.%【目的】克隆不同镉积累基因型烟草中阳离子转运基因（CAX2），为研究烟草CAX2基因在镉转运过程中的作用机理提供参考。【方法】根据NCBI中马铃薯的CAX2序列，BLAST烟草EST数据库，设计引物克隆普通烟草（Nicotiana tabacum）K326和黄花烟草（N. rustica）的CAX2基因，使用生物信息学方法进行序列分析。【结果】K326和黄花烟草CAX2基因（分别命名为NtCAX2和NrCAX2）编码区长度均为1314 bp，核苷酸序列一致性达98.0%；二者均编码437个氨基酸，仅有5个氨基酸位点存在差异。NtCAX2和NrCAX2蛋白同源性为98.9%，均有两个完全相同的Na+-Ca2+交换蛋白结构域，分别位于CAX2102~249 aa和293~426 aa区域，具有CAX家族共有的结构特征。NtCAX2和NrCAX2理论等电点分别为5.54和5.45，均有较强疏水性，包含11个跨膜结构域和两个重复区域α-1、α-2，三级结构以α-螺旋为主。系统进化分析结果表明，NtCAX2和NrCAX2与马铃薯（StCAX2）的亲缘关系最近，蛋白同源性均高于92.0%。【结论】K326和黄花烟草CAX2基因同源性较高，CAX2基因在茄科具有较高保守性。