关键词: 烟草, 钾转运体基因, 生物信息学, Northern杂交

Abstract:

【Objective】 The aim of this study is to clone K+ transporter gene in tobacco, analyze its genetic relationship, and predict its structure, nature, function and to provide a basis for K+ gene function studies of tobacco. 【Method】 Cloning tobacco K+ transporter gene by means of the combination of laboratory work and e-cloning techniques. Using tobacco K+ transporter gene 3' end sequence, cloned by RACE, as a probe to search the tobacco EST databases, filter out the ESTs sequences with the homology of 95% of the probe, and spliced the two segments using DNAStar software. Then, analyze gene structure and properties of the full cDNA sequence, and predict its function by applying a variety of bioinformatics databases and softwares. 【Result】A new tobacco K+ gene (TPK1) with full cDNA sequence was found. The molecular weight of the protein encoded by this gene is 26.21 kD, and the theoretical isoelectric point pI is 9.72. TPK1 is a hydrophobic membrane protein, containing six distinct transmembrane helix topologies. In secondary structure, a total of 11 α-helix, and 16 β-fold were found in TPK1. At subcellular level, TPK1 protein located in the plastid, vacuole and endoplasmic reticulum. The signal peptide splice site and the serine, threonine and tyrosine kinase phosphorylation sites in TPK1 amino acids sequence were found as well. In the function, TPK1 protein contains a typical conserved K+ uptake transporter function domain, and is highly of homology to the major K+ uptake transporter genes in higher plants. Northern blotting analysis indicated that TPK1 was expressed in root, stem, leaf and flower.【Conclusion】A new K+ transporter TPK1 was successfully cloned in tobacco, and its function maybe involved in K+ transportation in tobacco plants. Therefore, further investigation still needed to understand its role in tobacco K+ uptake and transportation.

Key words: tobacco, potassium transporter, bioinfomatics, Northern blotting