Cloning And Expression Of Wheat Low-affinity Cation Transporter Gene LCT1 And Its Splice Variant In Tobacoo Isolated From Full-length CDNA Pool

Wheat is one of the most important cereal foods in the world and its production has significant effect to world food supply. Full-length cDNA library plays an important rule in wheat fuctional genome research, such as physical map construction , gene identification .structure and function analysis, expression and mapping study in wheat genome project. Full-length cDNA pool is the most important step in construction of cDNA library.In this experiment, we use modified oligo-capping method to construct full-length cDNA pool, which is based on effective RACE technique. The PCR products of cDNA pool were ligased to T-vector and transformed to E. coli. White clones were picked randomLy and sequenced. After blasting them in NCBI database, we make sure the quality of full-length cDNA pool is quite satisfaction. In this way, the method can be used in construction of full-length cDNA library.LCTl gene of wheat can be expressed as low-affinity cation transportation protein which permits the uptaking of cations (e. g., Li⁺, Na⁺, K⁺, Ca²⁺, Cd²⁺ etc) into the cell. When this gene expresses in yeast mutant of Na⁺ transport deficiency (yeast strain G19), it shows hypersensitive to salt due to absorption of more cation. Low concentration of calcium can block cadmium transport into cell of root when transgenic tobacco expresses LCTl gene (Daniel, 1997 ). With respect to the LCTl mediated salt tolerance in plant, relevant research has seldom been made. In order to determine the function of LCTl gene in plant under salt stress and to improve wheat resistance to salt, we cloned LCTl gene from full-length cDNA pool and obtained a splicing variant of it, which is lack of a 115 base pairs fragment (perhaps intron). We also cloned LCTl genomic sequence. By comparing them, we find that this LCTl splicing variant (we call it LCT1-2 because it comes from number 2 clone) is the constitutive splicing product. While the reported LCTl gene is...