Cloning, Construction Of Expression Vector And Tabacoo (Nicotiana Tabacum L.)Transformation Of A Pollen Specific Gene OsPSG076Promoter In Rice (Oryza Sativa L.)

Both wheat and rice are the most important food crops in the world. Previous studyshowed that TaPSG076is a pollen-specific gene that is expressed in the developmentalprocess of pollens in wheat. However, the function of TaPSGO76remained unknown. Theresearch of PSG076gene promoters will help us to understand the gene regulation inpollen development and the growth of wheat.Since wheat has hexaploid genome and it is very large, the research of genes andtheir promoters in wheat will be time-consuming and heavy tasks, and work efficiency isnot high.By comparison, we found a homologous gene of TaPSG076in the rice genome andthe gene was designated as OsPSG076. Therefore, the present research focused onisolation and functional analysis of OsPSG076gene promoter, the results of which shouldbe helpful for understanding the regulating characterization of TaPSG076gene promoterin wheat.We successfully cloned a1266bp upstream sequence of OsPSG076from ricegenomic DNA using inverse PCR, based on the sequence of OsPSG076. Putativefunctional promoter elements were analyzed by the PLACE database. The results showedthat this upstream sequence contained the basic elements such as TATA-box and pollenspecific cis-elements such as AGAAA、GTGA, indicating that these motifs may conferpollen specific expression for the promoter driven gene.In order to analyze expression specific functions of various cis-elements contained inthis OsPSG076gene promoter, six vectors respectively named RP13-pBI、RP10-pBI、RP7-pBI、RP4-pBI、RP3-pBI、RP2-pBI, respectively, with different5’-deletions:-1258bp,-986bp,678bp,-379bp,-334bp and-236bp were inserted into pBI121.These vectorswere introduced into tobacco (Nicotiana tobaccum L.) plants by Agrobacterium-mediatedtransformation method.Transgenic tobacco was screened by PCR using GUS gene and different lengths ofpromoter fragments as targets. Results showed that only RP2-pBI was successfullytransformed into tobacco. Histochemical GUS staining on transgenic tobacco plantsshowed that the0.2kb promoter fragment neither possesses the pollen-specific function,nor has the basic promoter activity. The functional analyses of other promoter fragmentsare underway.