Abstract: A  putative lowaffinity nitrate transporter, OsNRT1.2   gene, was identified based  on the rice genomic database and bioinformatics analysis. By Agrobacterium tumefaciens  transformation with a Ubiquitin (Ubi) promoter, the gain of function of rice OsNRT1.2  was tested by overexpression in Wuyunjing 7. The OsNRT1.2  cDNA from Knowledgebased Oryza Molecular Biological Encyclopedia (KOME) database encoded 533 amino acids with a fulllength of 2178 bp and an open reading  fragment (ORF) of 1732 bp. The expression  pattern of OsNRT1.2  in Wuyunjing 7 was investigated and the results showed that its mRNA was only expressed in roots but not in shoots.  And the overexpressed  plants were generated by the transformation of pUbiOsNRT1.2  into Wuyunjing 7. Wild type (WT) and three T2 generation transgenic lines, named as OE1, OE2 and OE8, were used in the growth experiment at 0.2 and 5.0  mmol/L  nitrate  for 30 days and the plant height and root length were recorded every 10 days. The results showed that the plant height  of  WT and transgenic plants had no difference except OE8 line at 5.0 mmol/L  NO3-  , while the root length of transgenic lines significantly  decreased at both 0.2   and 5.0 mmol/L  nitrate. Nitrogen (N) content in shoots  and roots  was not different between WT and transgenic lines at 0.2 mmol/L  NO3-. But at 5.0  mmol/L  NO3-, compared with WT, the shoot nitrogen content of OE2 and OE8 except OE1 was increased and the root N content of all transgenic lines was significantly decreased.  The rootshoot ratio of transgenic lines was significantly decreased at 0.2 and 5.0 mmol/L nitrate levels. The dry weight of transgenic lines was significantly increased by from 9.4% to 31.1% at 0.2 mmol/L  nitrate and from 12.5% to 43.8% at 5.0 mmol/L  nitrate. It indicated that OsNRT1.2  in Wuyunjing 7 might  be involved in the N translocation from plant roots to shoots and therefore shoot biomass was increased.

Key words: rice, lowaffinity nitrate transporter gene, overexpression, nitrogen , translocation

摘要： 应用公开的植物基因组数据库和生物信息学分析确定了一个水稻低亲和硝酸盐运输蛋白候选基因OsNRT1.2。利用农杆菌介导Ubiquitin启动子过量表达OsNRT1.2，研究了该基因在武运粳7号中的获得性功能特征。OsNRT1.2的cDNA序列从KOME网站中获得，该序列全长为2178 bp，编码阅读框为1732 bp，编码533个氨基酸。采用半定量RTPCR技术分析，OsNRT1.2在武运粳7号中表达存在器官特异性，主要在根系表达，地上部分几乎不表达。通过将pUbiOsNRT1.2 在武运粳7号中转化，得到OsNRT1.2基因超量表达材料。0.2 mmol/L NO3-和5.0 mmol/L NO3-处理野生型（WT）和T2转基因植株OE1、OE2 和OE8  30 d，每10 d跟踪记录株高和根长。转基因植株（除OE8在5.0 mmol/L NO3-供氮水平外）与WT株高无明显差异，根长增加量显著降低。在0.2 mmol/L NO3-供氮水平下，转基因植株和WT地上部分和根系的全氮含量均无显著差异；在5.0 mmol/L NO3-供氮水平下，除OE1外，OE2和OE8地上部分全氮含量显著增加，3个转基因株系比WT根系全氮含量显著降低。在两个氮素水平处理条件下，转基因植株根冠比都显著降低；生物量比WT都增加，在0.2 mmol/L NO3-供氮水平下增加了9.4% ~ 31.1%，在5.0 mmol/L NO3-供氮水平下增加了12.5% ~ 43.8%。研究表明，OsNRT1.2基因在武运粳7号中可能参与了氮素从根系向地上部的转运，从而导致地上部生物量增加。

关键词: 水稻, 低亲和硝酸盐运输蛋白候选基因, 超量表达, 氮素转运