ABSTRACT
The purpose of this study is to investigate the function of a novel potassium transporter gene (NrHAK1) isolated from Nicotiana rustica roots using yeast complement and real-time PCR technique. The complementary DNA (cDNA) of NrHAK1, 2 488 bp long, contains an open reading frame (ORF) of 2 334 bp encoding a protein of 777 amino acids (87.6 kDa) with 12 predicted transmembrane domains. The NrHAK1 protein shows a high sequence similarity to those of high-affinity potassium transporters in Mesembryanthemum, Phytolacca acinosa, Arabidopsis thaliana, and so on. We found that the NrHAK1 gene could complement the yeast-mutant defect in K+ uptake. Among several tissues surveyed, the expression level of NrHAK1 was most abundant in the root tip and was up-regulated when exposed to potassium starvation. Moreover, the transcript accumulation was significantly reduced by adding 5 mmol/L NH4+ to the solution. These results suggest that NrHAK1 plays an important role in potassium absorption in N. rustica.
Subject(s)
Cation Transport Proteins , Chemistry , Genetics , Physiology , Plant Proteins , Chemistry , Genetics , Physiology , Potassium , Metabolism , Quaternary Ammonium Compounds , Pharmacology , Sodium , Pharmacology , Tobacco , Genetics , MetabolismABSTRACT
The DNA fragment sized 2 139bp, the same Sequence with AtKup1 gene from Arabidopsis thalianna was used as the templates for DNA family shuffling. The shuffeld AtKup1 gene library was expressed in the mutant of 5. cerevisae in which potassium transporter gene TRK1 and TRK2 were knocked out by homologous recombination. Then the screening was carried out in the low potassium media containing 5. 0 mmol/L KC1 and no histidine in it. it was found that both of diverse and wild AtKup1 gene can rescues the trk1△trk2△yeast mutant strain in low [ K + ] medium. The growth of 2 clones yeast containing diverse AtKup1 were beter than that of AtKup1 wild gene transformant. The sequencig results of the shuffeld AtKup1 showed that there were 2 nucleotide changed, which resulted in 2 amino acid variations in it compared with the original AtKup1. The potassium uptaking capacity of shuffled AtKup1 gene increased significantly when it was transformed into tobacco.