Abstract

Plants offer a promising alternative to microbial fermentation and animal cell cultures for production of recombinant proteins, the major advantages are safety, low cost, post-translation modifications and high volume of production. t-PA (tissue Plasminogen activator) is a trypsin-like serine proteinase and a superior thrombolytic agent. In this report, recombinant cDNA of tissue Plasminogen activator was transformed to tobacco plants. t-PA recombinant protein was expressed under the control of CaMV35S promoter and NOS terminator. A high-expression sequence (Kozak sequence) and KDEL signal (for endoplasmic reticulum retention of recombinant protein) were linked to amino and carboxy-termini of t-PA gene, respectively. The vector containing t-PA (pBIt-PA) was transferred to Agrobacterium tumefaciens and the t-PA gene was inserted into the plant genome by agrobacterium-mediated transformation. Transgenic plants were selected on kanamycin (100 mgL^-1), maintained in perlite and then the soil, subsequent generations were obtained. The presence and expression of the transgene was confirmed in the transformants by PCR, SDS-PAGE, RT-PCR, Zymography and Western blotting. This report examines the transformation and expression of t-PA gene in tobacco plants.

Key words: Recombinant protein, tissue plasminogen activator, tobacco, zymography.