| Nitrate (NO3) is the primary nitrogen source for most plants. In order to improve nitrate use efficiency, a more complete understanding of the transport of NO3ˉfrom the soil to the plant and within the plant itself is required. NO3ˉacquired by plants from soil is mainly transported via nitrate transporters from the NRT family. In Arabidopsis, the NRT2family has seven members which appear to be high-affinity transporters.Non-heading Chinese cabbage (Brassica campestris ssp. chinensis Makino), originated in China, is commonly grown vegetables species in the southern region of China. Understanding the complexity of the regulatory networks that control NO3ˉuptake, metabolism, and associated responses has the potential to provide solutions that will address the major issues of NO3ˉpollution and toxicity that threaten agricultural and ecological sustainability as well as human health.The main results are as follows:A full-length of cDNA sequence of a high-affinity nitrate transporter gene BcNRT2was cloned from non-heading Chinese cabbage cultivar’Suzhouqing’using reverse transcript PCR (RT-PCR). Sequence analysis showed that the length of nucleotide sequence of this gene is1593bp, containing a complete open reading frame encoding530amino acids. Nucleotide and amino acid sequence comparison indicates that BcNRT2has a high similarity with the orthologous gene BnNRT2in Brassica napus (98%,99%) and AtNRT2.1in Arabidopsis thaliana (90%,95%), respectively. Conclusion was made that NRT2gene is highly conserved among several plant species. Quantitative real-time PCR analysis showed that the expression of BcNRT2has the highest level in root cells, and this gene’s expression pattern belongs to induced system. After0.5h treatment of low concentrations of NO3ˉ, the expression of BcNRT2was up-regulated rapidly in root and shoot, suggesting that BcNRT2may act as a NO3ˉsensor or signal transducer. After the treatment of high concentrations of NO3ˉ, the expression of BcNRT2was highly up-regulated and lasted longer in root and shoot, which may be result from a high-level response of the regulation of a low-affinity nitrate transporter NRT1.1. BcNRT2protein was located at the plasma membrane supported by subcellular localization assays experiment.Synteny analysis approach and sequence similarity was used to identify putative orthologoues of the Arabidopsis NRT2genes in other eight fully sequenced genomes (Chinese cabbage, soybean, western poplar, grape vine, maize, rice, sorghum, and purple false brome). Sequence analysis indicated that these sequences belong to the Nitrate/Nitrite Poter family (NNP) of the Major Facilitator Superfamily (MFS). However, there is significant difference in the NRT2gene structure between monocots and dicots. Phylogentic tree analysis showed that, most members of the NRT2family developed primarily following the divergence of the monocots and dicots. Simultaneity, the evolution models of NRT2in different kinds of plants are different. NRT2family of Chinese cabbage (Brassica rapa) generated the genome triplication and gene loss. The NRT2family in Chinese cabbage has a relatively close evolutionary relationship with that of Arabidopsis. Evolution analysis provides useful reference clues for further function research of NRT2in Chinese cabbage to utilize existing research results of NRT2in Arabidopsis. |
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