Human Trefoil Factor 3 Gene Synthesized And Transformed Into Tomato

Intestinal trefoil factor 3 (TFF3) is a member of the trefoil family of peptides, named from its special trefoil structure, small molecules constitutively expressed in epithelial tissues, including the gastrointestinal tract. TFF3 has been shown to promote migration of intestinal epithelial cells in vitro and to enhance mucosal healing and epithelial restitution in vivo. The trefoil structure can protect TFF3 from digesting by proteinase and being destroyed by acid and thermal. So it can keep the whole structure and function in human gastric tract.Gene splicing by overlap extension (gene SOEing) is a technique that can be used in gene recombination and site specific mutation. Four oligonucleotides are synthesized for using in hTFF3 gene SOEing. After two reactions, we got the whole 183bp of hTFF3 gene. Plant as bioreactor exploited to produce heterologous protein possess many advantages such as expressing and folding the heterologous protein exactly, lower cost, easier to grow up, and can be mass-produced, and safety. For these, researches on it have been carried out extensively and exhibits more attraction.In this research, tomato was exploited as plant bioreactor to produce recombinant hTFF3, a plant expression vector was constructed, it under the control of CaMV35S promoter. After co-cultivation with Agrobacterium Tumefaciens GF3101 and several times kanamycin-resistant selection, eight regenerated tomato plantlets were obtained. One plantlet was ruled out by PCR, and then seven of them were analyzed further by Southern blot, these were confirmed that the hTFF3 gene was co-integrated with CaMV35S and Npt II into plant genome by Southern hybridization. The result of RT-PCR of transgenic plants showed: in transcriptional level, hTFF3 was expressed in leaves of tomato under control of CaMV35S promoter.