白菜NRT2基因的克隆及表达模式分析  

Cloning and Expression Pattern Analysis of NRT2 Gene in Non-heading Chinese Cabbage

作　　者：孔敏[1] 杨学东[1] 侯喜林[1] 刘同坤[1] 任君[1] 

机构地区：[1]南京农业大学作物遗传与种质创新国家重点实验室,农业部华东地区园艺作物生物学与种质创制重点实验室,南京210095

出　　处：《园艺学报》2011年第12期2309-2316,共8页Acta Horticulturae Sinica

基　　金：国家科技支撑计划项目(2009BADB8B03-1);江苏省自然科学基金项目(BK2009311);江苏省农业科技自主创新资金项目[CX(09)604]

摘　　要：以白菜(Brassica campestris ssp.chinensis Makino)品种‘苏州青’为试材,采用RT-PCR技术,获得1个高亲和硝酸盐转运蛋白基因(NRT2)的cDNA序列,全长1 593 bp,推断其编码530个氨基酸,命名为BcNRT2。序列分析表明:BcNRT2基因与甘蓝型油菜BnNRT2基因和拟南芥AtNRT2.1基因核苷酸序列的相似性分别为98%和90%,氨基酸序列的相似性分别为99%和95%,表明植物中NRT2基因保守度较高。实时定量PCR表达分析表明,BcNRT2在白菜根部的表达量最高,为诱导型表达。低浓度NO3-(0.2 mmol.L-1 KNO3)处理0.5 h后其表达量迅速上升,BcNRT2可能为NO3-感受器。高浓度NO3-(20 mmol.L-1 KNO3)处理后其表达量更高,持续时间较长,可能是受到低亲和硝酸盐转运蛋白NRT1.1的调控而产生的高水平响应。原生质体的瞬时表达显示,BcNRT2蛋白位于细胞膜上。In this study,a full-length of cDNA sequence of a high-affinity nitrate transporter gene BcNRT2 was cloned from non-heading Chinese cabbage（Brassica campestris ssp.chinensis Makino） cultivar‘Suzhouqing’using reverse transcript PCR（RT-PCR）.Sequence analysis showed that the length of nucleotide sequence of this gene is 1 593 bp,containing a complete open reading frame to encode 530 amino acids.Nucleotide and amino acid sequence comparison indicates that BcNRT2 has a certain high similarity with the orthologous gene BnNRT2 in Brassica napus（98%,99%）and AtNRT2.1 in Arabidopsis thaliana（90%,95%）,respectively.Conclusion was made that NRT2 gene is highly conserved among several plant species.Quantitative real-time PCR analysis showed that the expression of BcNRT2 has the highest level in root cells,and expression pattern of this gene belongs to induced system.After 0.5 h treatment of low concentrations of NO3-（0.2 mmol.L-1 KNO3）,the expression of BcNRT2 was upregulated rapidly in root and shoot,suggested that BcNRT2 may acts as a NO3-sensor or signal transducer.After treatment of high concentrations of NO3-（20 mmol.L-1 KNO3）,the expression of BcNRT2 was highly up-regulated and lasted longer in root and shoot,which may be resulted from a high-level response of the regulation of a low-affinity nitrate transporter NRT1.1.BcNRT2 protein was located at the plasma membrane supported by subcellular localization assays experiment.

关 键 词：白菜 硝酸盐 NRT2 实时定量PCR 亚细胞定位 

分 类 号：S634.3[农业科学—蔬菜学]