Transfer Of Para-caroene Biosynthesis Enzyme Gene(LycB) Into Tomato And Chrysanthemum Mediared By Agrobacterium Tumefaciens

LycB gene is one of the key enzyme during the course of biosynthesis of para-carotene biosynthesis. As a medicinal and healthful agent, the function of para-carotene have been paid more and more attention such as postponing ageing, enhancing human immunological competence, preventing cardiorascular diseases, and antitumous effect, especially P -carotene, prisoma is also of vitamin A.antitumous effect, Para-caroene concludes almost important pigment from red to yellow, which can alter flower coulor and be used in floribreeding.LycB gene was transferred into tomato and chrysanthemum respectively Mediatied by Agrobacterium tumefaciens, and the genetic transformation system were studied.Using tomato species including Dongnong 704,708,709 as material and their cotyledons, petiol segments as acceptor, Target lycB gene contained in A. tumefaciens (EAH101) was transformed into the cells. The factors influencing the transformation have been studied and regenerated tomato plants were examined by PCR and PCR-Southern analysis . The results were as follows:Acceptor (cotyledon and petiol segment) after pre-culture was more suitable for the infection and transformation of A. tumefaciens compared to un-culture ones.The frequency of resistant calli after pre-culture 5d was hijher than pre-culture 2d.The infection time was suitable for cotyledon 10 min and petiol segment 15 min.The fittest OD600 of engineering bacterial suspension was 0.7.The addition of AS could increase the frequency of resistance calli and the frequencyreached the highest when the concentration of AS was 100 mol/L.The moderate postponement of selection time was beneficial to transformation.It was proved that target LycB gene had been integrated into the genome of tomato after PCR, PCR-Southern analysis. Five transgenic tomato plants were obtained, at present the research in this fields has not been reported at home.Using chrysanthemum(30,33,34)as material and the calli from their acceptor. The factors influencing genetic transformation were optimized through the frequency of resistance calli. The results were as follows:The suitable time was 5min for chrysanthemum calli to be infected by A tumefaciens.The suitable OD600 of engineering bacterial suspension was 0.55.The most effective could been obtained when the co-culture time was 3d.The addition of AS could increase the induction frequency of resistance calli and obtain the best frequency was the highest when the concentration of AS was 100 mol/L.The resistance calli rate could be increased effectively by increasing Hyg concentration from the low to the high.It was originally proved by the PCR examination that the target gene had been transferred into chrysanthemum calli.