茶树硝态氮转运蛋白NRT1.1基因的克隆及表达分析  

Cloning and Expression Analysis of Nitrate Transporter NRT1.1 Gene in Tea Plant(Camellia sinensis(L.))

作　　者：杨亦扬[1,2] 胡雲飞[1] 万青[1] 李荣林[1] 王枫[3] 阮建云[2] YANG Yiyang HU Yunfei WAN Qing LI Ronglin WANG Feng RUAN Jianyun(Institute of Horticulture, Jiangsu Academy of Agricultural Science, Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement, Nanjing 210014, China Tea Research of Institute, Chinese of Academy of Agricultural Science, Key Laboratory of Tea Biology and Resources Utilization, Ministry of Agriculture, Hangzhou 310008, China College of Horticulture, Nanjing Agricultural University, State Key Laboratory of Crop Genetics and Germplasm Enhancement, Nanjing 210095, China)

机构地区：[1]江苏省农业科学院园艺研究所江苏省高效园艺作物遗传改良重点实验室,江苏南京210014 [2]中国农业科学院茶叶研究所农业部茶树生物学与资源利用重点实验室,浙江杭州310008 [3]南京农业大学园艺学院作物遗传与种质创新国家重点实验室,江苏南京210095

出　　处：《茶叶科学》2016年第5期505-512,共8页Journal of Tea Science

基　　金：国家自然科学基金(31400587);江苏省自然科学基金(BK20160590);茶树生物学与资源利用国家重点实验室开放基金(SKLTOF20150114);江苏省农业科技自主创新资金(CX(16)1003)

摘　　要：以茶树(Camellia sinensis(L.))品种龙井43为试材,采用PCR结合RACE技术,克隆得到硝态氮转运蛋白基因(NRT1.1)的c DNA全长序列,基因序列全长1 880 bp,其中开放阅读框(ORF)1 788 bp,编码595个氨基酸,预测蛋白质分子量为65.9 k D,理论等电点为8.99,命名为Cs NRT1.1。序列分析表明,Cs NRT1.1与葡萄NRT1.1氨基酸序列的相似性最高。通过生物信息学分析,对Cs NRT1.1的氨基酸理化性质、亲/疏水性、跨膜区域及亚细胞定位进行了预测。实时定量PCR表达分析表明,茶树根和叶片中Cs NRT1.1在1 mol·L^(-1) NO3-处理5 min内均受到抑制,叶部Cs NRT1.1表达量0.5 h后即达到最大值,24 h内各个时间点均高于根部,根中表达量Cs NRT1.1始终低于对照。本研究结果为研究茶树对NO3-的吸收、转运和调控机理提供了分子生物学基础。A full length cDNA sequence of Nitrate transport gene （NRT1.1） was obtained from tea plant （Camellia sinensis （L.）） cultivar ‘Longjing 435 by polymerase chain reaction （PCR） and rapid amplification of cDNA ends PCR （RACE-PCR）. The length of nucleotide sequence of this gene was 1 880 bp, containing a complete open reading frame （1 788 bp） to encode 595 amino acids. The putative protein had an isoelectric point of 8.99 and a calculated molecular weight of 65.9 kD. CsNRTl.1 was highly homologous to the gene NRT1.1 in Vitis vi^ifera by sequence alignment. Several parameters of these sequences, including sequences composition, physicochemical property, topological structure of transmembrane regions, hydrophobicity or hydrophilicity, subcellular localization were predicted by bioinformatics tools. Quantitative real-time PCR analysis showed that the expression of CsNRT1.1 in roots and leaves were inhibited after incubation in 1 molL-1 NO3 - for 5 min. The expressions of CsNRT1.1 in roots were always lower than that of CK within 24 h. Its expressions in leaves were higher than those in roots with its peak at 0.5 h. Our results provides favorably help to reveal NO3 ＂ uptake and utilization in tea plants.

关 键 词：茶树 硝态氮 NRT1.1基因 实时定量PCR 

分 类 号：S571.1[农业科学—茶叶生产加工] Q51[农业科学—作物学]