海州香薷不同抗性种群液泡转化酶基因 EhvINV 序列趋异及表达差异分析

摘要

Specific primers were designed to clone the cDNA sequences according to the EhNvINV(JX500755)and EhCvINV (JX500756)from GenBank. The DNA and deduced amino acid sequences were analyzed by bioinformatics methods. The three-dimensional structures were constructed by homologous modeling. The structures of vacuolar invertase from two populations of E. haichowensis and non-tolerant population with each single point mutation in complex with sucrose were simulated by AutoDock 4.0. Transcript expression of EhNvINV and EhCvINV under copper tress was analyzed by real-time PCR. The results showed that there were two divergent amino acids at position 114 and 346 between EhNvINV and EhCcINV. The three-dimensional structures were exactly similar between EhNvINV and EhCvINV. It showed difference at Glu114/Gln114 and Leu346/Pro346,which were divergent sites. The structures of catalytic active center of EhNvINV,EhCvINV, and simulated mutants(EhNvINV-E114Q,EhNvINV-L346P)binding with sucrose showed no significant differences. However,there were differences on spatial position. The result of real-time PCR indicated that the transcript expression of EhCvINV induced by copper stress after 7 days,however,the transcript expression of EhNvINV inhibited by copper stress after 7 days.%根据 GenBank 中海州香薷（Elsholtzia haichowensis）液泡转化酶基因 EhNvINV（JX500755）和 EhCvINV（JX500756）的序列设计特异性引物，克隆 cDNA 全长序列，并通过生物信息学分析基因及推导的蛋白序列，利用 SWISS-MODEL 进行同源建模，并与蔗糖分子进行模拟对接，实时荧光定量 PCR 分析 EhNvINV 和 EhCvINV 在铜胁迫下的转录表达。结果表明，海州香薷非抗性和抗性种群液泡转化酶蛋白 EhNvINV 和 EhCvINV 在114和346处存在氨基酸趋异位点，模拟3D 结构相似，仅在趋异位点 Glu114/Gln114和 Leu346/Pro346处有差别。EhNvINV、EhCvINV 和模拟突变体（EhNvINV-E114Q 和 EhNvINV-L346P）与蔗糖分子对接形成的活性中心构象基本一致，但在空间位置上存在细微差异。实时荧光定量 PCR 结果表明铜胁迫7 d 后，EhCvINV 的表达受铜的诱导，而 EhNvINV 受铜的抑制。