Genetic Transformation Of Tomato With VP7 Gene From A Group A Rotavirus G1 Chinese Strain

Rotaviruses (RV) are the most common cause of severe diarrhea in children all over the world,VP7 is major outer capsid protein and is a primary candidate for inclusion in a subunit or recombinant. Advances in genetic engineering in the past decade have accelerated the expression, in plant, of foreign proteins with pharmaceutical value. Antigens from infectious bacterial or viral diseases have been introduced into plants. So introducing VP7 gene into tomato to develop edible plant vaccine of rotaviruses would change the traditional means of production of vaccines and the cost of vaccine production would be reduced greatly. It is provided with commercial meaning and applicative prospect.In this study, two plant expression vectors pBIVP7 and pEVP7 were constructed. In pBIVPT, it is under control of CaMV35S promoter. In pEVP7, it is under control of tomato-fruit-specific TFP promoter. After co-cultivation with Agrobacterium Tumefaciens EHA105(containing pBIVP7 or pEVP7) and several times kanamycin-resistant selection, eighteen regenerated tomato plantlets were obtained . One plantlet was ruled out by PCR , then 9 of 17 plantlets selected randomly were analyzed further by PCR-Southern blot and Southern blot, eight plantlets were confirmed that the VP7 gene had been integrated into plant genome by Southern hybridization. The result of RT-PCR of transgenic plants showed: in transcriptional level, VP7 gene was expressed in leaves and fruits of 4 transgenic tomato plants under control of CaMV35S promoter; VP7 gene was expressed in the tomato fruits not in the leaves of 3 transgenic tomato plants under control of fruit-specific TFP promoter.One TO offspring (T1) was selected for further genetic analysis from two types of self-pollinated transgenic plants and screened for the presence of the recombinant gene by the PCR respectively. The result suggested that the resistance to kanamycin can be a stable heredity by genetic analysis, separation ratio of T1 was 3:1 and T1 kanamycin-resistant tomato plants can carry VP7 gene.