Screening And Characterization Of Promoters In Rice Promoter Trap Lines

Promoter trap vectors were constructed using a promoterless reporter genewhich was located next to the right (left) border of T-DNA. The reporter gene can notbe expressed until the reporter gene was just inserted inside gene extron ordownstream of a plant promoter, and directions of the promoter and reporter gene wasconsistent. Promoter trap strategy provided us a powerful tool for analyzing newgenes and their regulatory elements. This study was focused on screening and analysisof rice promoters based on rice promoter trap lines from Fujian Key Laboratory ofGenetic Engineering for Agriculture. The main results were as follows:Author detected the GUS expression in different tissues in 940 plants from ricepromoter trap lines at seedling stage, vigorous tillering stage and heading stage. Theresults was found that 26 plants showed GUS activity at least one tissue and the GUSexpression rate was 2.77%. Of which 16 showed special expression in root, culm/leaf,flower or seed of rice, and the others in more than two tissues (organs).By TAIL-PCR technique, flanking sequences of 25 T-DNA inserted sites wereobtained in 22 of 26 rice promoter trap plants above. Informatiaon analysis of thesesequences showed that the eighteen T-DNA fragments were integrated inside genes,and the two in downstream of promoters. Five flanking sequences were not annotatedor they might be located between two genes.Author selected to analyze a rice promoter P8513 which was speculated onspecial expression in culm and leaf of rice based on GUS expression pattern ofpromoter trap line w8513 and its flanking sequence. A full-length of P8513 promoterwas cloned and a fusion expression vector was constructed with gus gene. Analysis oftransgenic Minghui 86 and Nipponbare showed expression pattern of gus gene inthese transgenic plants seedling phase was similar to that of the trap line w8513.Further analysis was under way in tillering and reproductive phase.