Cloning And Squence Analysis Of Genes Related To Glutelin In Wild Rice

Objective:To clone cDNA encoding glutelin from wild rice (Zizania latifolia Turcz), acquire the property right of the genes, establish the foundation of function study related to glutelin in wild rice, probe into evolution between wild rice and rice by comparison with the sequence of glutelin, enrich the gene pool of rice and do favor to the characters improving of rice.Methods:(1) A CTAB-LiCl-based extraction method which is especially useful for the RNA extraction of plant material plenty of polysaccharide, hydroxybenzene and lipids is improved. Extract total RNA of wild rice embryo samples through this method.(2) Design the primers based on conserved sequence of rice glutelin genes reported previously and two partial cDNA sequences of wild rice glutelin genes named Glu-1 and Glu-2 is obtained from RT-PCR.(3) Design gene-specific primers based on the two specific fragments and the 5'-end and 3'-end sequences of glu-1 and glu-2 are obtained from RLM-RACE.(4) Both Glu-1 and Glu-2 complete cDNA sequence are obtained by overlapping 5'RACE and 3'RACE products and analyzed with bioinformatics.Results:(1) High-quality total RNA of wild rice embryo samples is obtained.(2) The length of specific fragments obtained from RT-PCR are 1381bp (Glu-1) and 1279bp (Glu-2). BLAST research with database (www.ncbi.nlm.nih.gov) shows that they are highly homogenous with rice glutelin genes.(3) The 5'-end and 3'-end sequences of Glu-1 and Glu-2 are obtained from RLM-RACE. The 5'RACE and 3'RACE products size of Glu-1 are 433bp and 358bp respectively, and Glu-1 are 447bp and 695bp respectively. Sequence analysis shows...