The Expression Of Analgesic-antitumor Peptide (AGAP) From Chinese Buthus Martensii Karsch In Transgenic Tobacco And Tomato

Scorpion is a traditional Chinese drug. It has been applied to treatment of convulsion and epilepsy in Chinese traditional medicine. Scorpion venom, the main active component of scorpion, contains various toxic proteins with different functions. Analgesic-antitumor peptide(AGAP) has been purified from the scorpion venom of Buthus martensii Karsch which is widely distributed in China. Pharmaceutical tests showed that AGAP plays roles in both analgesic and antitumoral activities such as prolonging the survival days of mice engrafted Ehrlich ascites tumor cells, inhibiting the S-180 fibro sarcoma efficiently and also inhibiting the twisting action of the mice induced by acetic acid and increasing tolerance in the hot-plate mice model. So there was promising that AGAP was put to use at clinical situations as antitumor drug.The present study aimed to obtain Analgesic-antitumor peptide (AGAP) gene expression in plants. AGAP gene was cloned into the plasmid pBI121 to obtain plant expression vector pBI-AGAP. By tri-parental mating and freezing transformation methods, pBI-AGAP was transformed into Agrobacterium tumefaciens LBA4404, and pBI-AGAP/ LBA4404 was obtained by verifying.Tobacco (Nicotiana tabacum)and tomato(Lycopersicom esculentum) was transformed by Agrobacterium-Mediated leaf disc transformation method. Transformants were screened to grow and root on media containing kanamycin. Then, transformations were confirmed by analysis of PCR, and the results confirmed that the AGAP gene was integrated into the tobacco and tomato genomic DNA first. Further results verified that the transcriptions of AGAP gene were carried out in resistance plantlets by analysis of RT-PCR. Finally.it was confirmed that AGAP gene was successfully expressed in transgenic tobacco and tomato plantlets by analysis of western blotting.