Unexpected Transcription of Human t-PA cDNA in transgenic tobacco plants

Document Type : Research Paper



Recombinant tissue-type plasminogen activator (rt-PA) is used as a fibrin-specific thrombolytic agent for the treatment of various thromboembolic diseases. In this study, we tried to show the expression of transferred cDNA of human t-PA gene in transgenic tobacco plants produced previously by this group. RT-PCR with specific primers for t-PA, were used to assay the transgenic plants. RT-PCR products were observed on agarose gel electrophoresis after ethidium bromide staining. The results showed that the t-PA gene is stably expressed in second generation transgenic plant at the level of mRNA, while no expression was seen in non transgenic plants. These results also indicate an unfamiliar 650 bp transcript from t-PA transgene in plants. The results of homology search by CDD at NCBI database demonstrate that in addition to complete cDNA, an incomplete transcript so observed. This truncated transcript is a part of cDNA of t-PA that has lost the kringles domain while tyrosine-like serine protease is remained. This may results from alternative RNA splicing. The serine protease domain can carry out the role of t-PA alone. So many scientists are trying to clone this fragment only as a drug. Interestingly the plant produces this fragment naturally. This result may be because of alternative splicing of RNA.


 Domain, Kringle, recombinant t-PA and transgenic tobacco.


Bachmann F (1994). Molecular aspects of plasminogen, plasminogen activators and plasmin. In Hemostasis and Thrombosis. Bloom AL, Forbes CD, Thomas DP, Tuddenham EGD (eds), third edition, Churchill Livingstone, Edinburgh. 575-613.
Brondyk W H (2009). Chapter 11 Selecting an Appropriate Method for Expressing a Recombinant Protein. Methods in enzymology 463: 131–147.
Brown TA (1998). Gene Cloning. Blackwell Science LTD. PP. 50-56. 3rd edition
Bruen KJ, Ballard JR, Morris SE, Cochran A, Edelman LS, Saffle JR (2007). Reduction of the incidence of amputation in frostbite injury with thrombolytic therapy. Surgery 6: 546–51.
Collen D, Lijnen HR (1991). Basic and clinical aspects of fibrinolysis and thrombolysis. Blood 12: 3114-3124.
Daniell H, Stephen JS and Wycoff K (2001). Medical molecular farming. Prodaction of antibodies biophamaceuticals and edible vaccines in plants. plant Science 615: 219-26.
Fischer R, Stoger E, Schillberg S, Christou P and Twyman RM (2004). Plant-based production of biopharmaceuticals. Plant Biology 7: 152–158.
Frank A (1995). t-PA, Product Monograph Copyright © 1995 Chromogenix AB. Version 1.1. Sweden.
Gaffney PJ, Longstaff C (1994). An overview of fibrinolysis. In: Hemostasis and Thrombosis. Bloom AL, Forbes CD, Thomas DP, Tuddenham EGD (eds), third edition, Churchill Livingstone, Edinburgh. 549-571.
Henkin J, Marcotte P, Yang H (1991). The plasminogen-plasmin system. Cardial Disease 2: 135-164.
Masoumi asl A (2009). Cloning and transformation of human t-PA gene to tobacco plants and analysis of transgenic plants. Ph.D. Thesis. Department of plant breeding, faculty of agriculture, tarbiat mofares university. Tehran-Iran.
Mayer M (1990). Biochemical and biological aspects of the plasminogen activation system. Clincal Biochemistry 23: 197-211.
Mosher DF (1990). Blood coagulation and fibrinolysis: an overview. Clinical Cardiology 13: 5-11.
Pan H, Yang CP, Wei ZG and Jiang J (2006). DNA extraction of birch leaves by improved CTAB method and optimization of its ISSR system. Forestry Research 17: 298-300.
Schillberg S, Zimmermann S, Voss A, and Fisher R (1999). Apoplastic and cytosolic expression of full size antibodies and antibodies fragments in Nicotiana tabacum. Transgenic Research 8: 217-221.
Sonya N (2005). Molecular farming. Science and Technology Division Canadian Library of Parliament. http://www2.parl.gc.ca/Content/LOP/ResearchPublications/prb0509-e.htm. Retrieved 2008-09-11.
Von FT, McDiarmid T,  MacKler L,  Zolotor  A (2008). Clinical inquiries: Can recombinant growth hormone effectively treat idiopathic short stature? family practice. 57: 611–612.
Wiman B, Hamsten A (1990) The fibrinolytic enzyme system and its role in the etiology of thromboembolic disease. Seminar Thromb Haemostas 3: 207-216.