Sucrose transporter is a functional protein family related to the sucrose transportation and signal transduction in plants. Using DNA recombination technology, it was prepared the specific antibody of rice sucrose tra...Sucrose transporter is a functional protein family related to the sucrose transportation and signal transduction in plants. Using DNA recombination technology, it was prepared the specific antibody of rice sucrose transporter (OsSUT2) based on its typical sequence of hydrophilic non-transmembrane to provide efficient molecular tool in the research on the physiological function of this protein. The cDNA sequences of the hydrophilic non-transmembrane region of central loop (CL) and TACL (Transmembrane and Apoplast regions and CL) of OsSUT2 were sub-cloned into the fusion expression vector pQE40 having dihydrofolate reductase (DHFR) to construct recombinant plasmid pQE40-OsSUT2-CL and pQE40-OsSUT2-TACL, respectively. The pQE40-OsSUT2-CL can be highly expressed in E. coli M15, especially in E. coli BL21 plysS. However, the pQE40-OsSUT2-TACL cannot be expressed in E. coli M15. DHFR-OsSUT2-CL purified with SDS-PAGE gel was injected into rabbit to obtain the produce antiserum. The specificity of obtained antibody was tested by Western blotting. It was indicated that the raised antibody can specifically identify OsSUT2, and it can be efficiently used in the research on the physiological function of OsSUT2.展开更多
文摘Sucrose transporter is a functional protein family related to the sucrose transportation and signal transduction in plants. Using DNA recombination technology, it was prepared the specific antibody of rice sucrose transporter (OsSUT2) based on its typical sequence of hydrophilic non-transmembrane to provide efficient molecular tool in the research on the physiological function of this protein. The cDNA sequences of the hydrophilic non-transmembrane region of central loop (CL) and TACL (Transmembrane and Apoplast regions and CL) of OsSUT2 were sub-cloned into the fusion expression vector pQE40 having dihydrofolate reductase (DHFR) to construct recombinant plasmid pQE40-OsSUT2-CL and pQE40-OsSUT2-TACL, respectively. The pQE40-OsSUT2-CL can be highly expressed in E. coli M15, especially in E. coli BL21 plysS. However, the pQE40-OsSUT2-TACL cannot be expressed in E. coli M15. DHFR-OsSUT2-CL purified with SDS-PAGE gel was injected into rabbit to obtain the produce antiserum. The specificity of obtained antibody was tested by Western blotting. It was indicated that the raised antibody can specifically identify OsSUT2, and it can be efficiently used in the research on the physiological function of OsSUT2.
