EXPRESSION ANALYSIS AND PURIFICATION OF HUMAN RECOMBINANT TISSUE TYPE PLASMINOGEN ACTIVATOR (rt-PA) FROM TRANSGENIC TOBACCO PLANTS

Abstract

Recombinant tissue-type plasminogen activator (rt-PA) has been produced in different hosts. In this research, transgenic tobacco was selected for production of human rt-PA. Transgenic plants were analyzed by polymerase chain reaction (PCR) and reverse-transcription (RT)-PCR. The protein was extracted by Lysine Sepharose chromatography column and was further purified by HiTrap desalting column. The function of eluted protein was analyzed on zymography gel. The results showed that the 1.7-kb cDNA of tissue-type plasminogen activator (t-PA) (as well as a shortened 650-bp transcript of t-PA) has been expressed in transgenic plants. The anticipated 63-kD protein band and an additional 53-kD protein were observed in transgenic plants. Finally, zymography assay revealed that the purified rt-PA has anticipated appropriate activity comparable to a positive control drug (Alteplase). On the whole, we can say that transgenic tobacco is a good alternative host for production of t-PA.

Keywords:

• HiTrap desalting column
• Lysine Sepharose
• tissue-type plasminogen activator
• transgenic tobacco

ACKNOWLEDGMENTS

Transgenic tobacco seeds are a gift from Mr. Masoumi Asl. This work was supported by a grant from International Center for Science, High Technology and Environmental Science, Kerman, Iran.