Cloning and Expression Analysis of Potato Nitrate Transporter Gene
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    Abstract:

    In this study, we used potato double haploid ‘DM’ as material to clone the complete cDNA of highaffinity nitrate transporter gene StNRT2.1(JGI accession number PGSC0003DMT400002924). Expression pattern and bioinformatics analysis of the StNRT2.1 gene were constructed in this study, which provided a theoretical foundation for further exploration of the biological functions of the StNRT2.1 gene and improving the efficiency of nitrogen utilization in potato. The results showed that: (1) complete cDNA fragment of StNRT2.1 gene was obtained by homologous cloning and PCR amplification, and the expression vector pCEGFPStNRT2.1 was constructed. From the sequencing results, it was found that the actual encoded protein sequence was identical with the target protein sequence in the database, indicating that StNRT2.1 gene was cloned successfully without missense mutation. (2) StNRT2.1 gene is located on the chromosome 11 of potato and cDNA sequence was 1 593 bp in length encoding 530 amino acids. The relative molecular mass of the protein is about 57.60 kD, and the theoretical isoelectric point is 9.36. (3) Bioinformatics analysis showed that StNRT2.1 consisted of 20 different amino acids, of which glycine (Gly) accounted for the most, reaching 10.8%, and mainly consisted of 228 αhelix, 27 βsheet, 87 extended chain and 188 irregular coils. Moreover, the functionally conserved structure MFS_1 (PF07690) and 12 transmembrane helix domains are present, and the Nterminus and Cterminus are located in the cell membrane. StNRT2.1 is predicted to be located in the cytoplasm without signal peptide. Therefore, it may be a nonsecret membrane protein. (4) Compared with sufficient nitrogen level (7.5 mmol/L), potato seedlings showed obvious phenotypic changes including yellowed leaves and shorter plant with the three week treatments of nitrogenfree (0 mmol/L) and lownitrogen (0.75 mmol/L) level. (5) The results of realtime PCR showed the expression of StNRT2.1 gene in root tissue increased 3.98 times with the nitrogenfree treatment, which suggesting that StNRT2.1 may be an inducible highaffinity transporter.

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NAN Yunyou, L Tingting, CAO Minxuan, LIU Hengzhi, CHEN Yue, CHEN Qin. Cloning and Expression Analysis of Potato Nitrate Transporter Gene[J]. Acta Botanica Boreali-Occidentalia Sinica,2019,39(4):588-594

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  • Online: May 17,2019
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