莲藕Na^+/H^+逆向转运蛋白基因-LrNHX的克隆及表达分析

Cloning and expression analysis of LrNHX in Lotus Loot (Nelumbo Nucifera Gaertn)

[目的]寻求解决莲藕栽培种普遍对盐敏感的途径,从耐盐野生种中克隆了一个候选耐盐基因-LrNHX的全长c DNA序列,并对其表达进行分析,为今后利用基因工程技术改良莲藕耐盐性奠定基础。[方法]根据NCBI数据库中的莲藕Na^+/H^+逆向转运蛋白基因序列设计引物,利用PCR技术在莲藕叶片中克隆LrNHX,再利用半定量RT-PCR研究了250 mmol/L Na Cl处理0 h、6h、12 h、18 h、24 h和30 h时莲藕叶片中LrNHX的表达情况。[结果]LrNHX cDNA全长1 861 bp,编码526个氨基酸。Blast比对后发现该基因和与胡杨(ACX46912.1)、毛白杨(AAX37333.1、番茄(CAC83608.1)、拟南芥(NP 178079.2)的同源性分别达88%、87%、83%、82%。半定量RT-PCR结果表明,该基因在250 mmol/L Na Cl处理12 h后表达显著增高,说明LrNHX受盐诱导。[结论]LrNHX参与了莲藕耐盐信号的转导。

[Objectives] Cultivars of lotus root used in product are all sensitive to salt. In this study,a candidate salt resistant gene-LrNHX was isolated from a salt-tolerated wild lotus,and expression profiling was also analyzed using semi-RT-PCR method with aim to improve lotus salt resistant by genetic engineering technology in further. [Methods]Primers were designed according to the existing sequence of lotus＇ s NHX,and the full-length of LrNHX c DNA was obtained in lotus leaf using PCR method. In addition,LrNHX expression in leaf was also identified by the semi RT-PCR method under 250 mmol/L NaCl treatment for 0 h,6h,12 h,18 h and 24 h. [Results] The full length c DNAa of LrNHX was 1 861 bp encoding 526 amino aicd. The gene showed 88%,87%,83%,82% similarity with Populus euphratica（ ACX46912. 1）,Populus Tomentosa（ AAX37333. 1）,Solanum lycopersicum（ CAC83608. 1）,Arabidopsis thaliana（ NP 178079. 2） respectively after compared against to NCBI database. The results of RT-PCR elucidated that LrNHX was induced by NaCl,and the expression was enhanced after 250 mol/L NaCl treatment for 18 h. [Conclusion] LrNHX probably involved in signal transduction for salt resistance in lotus.