Cloning, Expression And Characterization Of Genes Related To Glutelin In Rice

Rice seeds accumulate large storage proteins (seeds storage protein, SSP) in endosperm during seed development, including glutelins and prolamins which account for up to 90% of the total proeins of the seeds. Both glutelins and prolamins were deposited in protein bodis (PBs), prolamins deposit in PB-Ⅰ while glutelins in PB- Ⅱ.In this study, three glutelin genes and one protein disulfide isomerase (PDI) gene cDNA were cloned with cDNA library sellection and reverse trascription polymerase chain reaction (RT-PCR) in addation to 3' rapid amplification of cDNA ends (3 'RACE). Then, the characterization structure and expression pattern of genes' were analysized. In order to identify the the function of PDI, a prokaryotic expression vector and one plant expression vector were constructed respectively.The results were as followings:(1) Cloning, expression, and charactorization of three cDNA encoding glutelin from rice. After screening the cDNA library with α -32P labeling GluB2 partial cDNA sequence obtained from RT-PCR, a full-length glutelin cDNA was isolated, named GluB-7. The genome sequence of GluB7 was obtained by PCR with the DNA template from XiuShui 11 and the sepecial primers designed with the cDNA seqeucne. The full-length cDNA of GluB-7 is 1588bp, which containing a 1488bp OREAfter BLAST research with conserved sequence of rice glutelin genes reported previously, many identical contigs were obtained. We predict these contig sequence with gene prediction software (www.softberry.com ) and analysized the putavie gene by BLAST with the database (www.ncbi.nlm.nih.gov ) . The results shows that a contig, GenBank accession number AP005511 may contain two new glutelin genes. According to the prediction sequences, we design two gene-specific primers. Two cDNA was obtained by RT-PCR using as template totaol RNA extracted from endosperm 12DAF, and the products size were 1517bp (GluB-6) and 1519bp (GluC-1) . Based on the GluB-6 and GluC-1 fragment we designed primers for 3'RACE. Both GluB-6 and GluC-1 complete...