Genetic Transformation Of Tomato With Apple α-Farnesene Synthase Gene

α-Farnesene, an acyclic sesquiterene existed in organisms, has been proved to play an important role in codling moth,chilling injury and development of scald on cold-storage apples. As one of aroma related volatiles in apple fruit,α-farnesene appears to be predominantly through classical mevalonate pathway. 3-Hydroxy-3- methylglutaryl CoA reductase (HMGR), AFS is the final, rate-limiting enzyme that converts farnesyl diphosphate (FPP) toα- farnesene in the mevalonic acid (MVA) pathway of terpene biosynthesis.Specific primers ac cording to the full sequence ofα-farnesene synthase gene were designed, sense eukaryotic vector was transformed. A highly efficient regeneration system and genetic transformation system of tomato were established,and physiological characteristic of transgenic plant was studied.The main results were as follow:1. A recombinant plant expression vector pBI-AFS was constructed and was used to transform to Tomato.2. Adventitious bud regeneration from the cotyledon of tomato and efficient gene transformation system was optimized.The results showed that the optimum medium for adventitious bud regeneration was MS+ZT2.0mg/L+IAA 0.5mg/L+sucrose30.0g/L+ agar 6.0g/L, MS+ IAA 0.5mg/L+sucrose30.0g/L+agar 6.0g/L for root regeneration.The suitabale concentration of kan is 50mg/L,and cef is 100mg/L.3. A recombinant prokaryotic expression vector pEASY-E1-AFS was constructed by cloning the fragment of AFS and was transformed to E.Coli. Western blot analysis indicated that AFS gene expressed in tomato.4. The transgenic plants ,with higher adaptability to high temperature than wild type, could keep running of photosynthesis in high stress. 5. Constructed pBI-AFS-GFP eukaryotic expression vector by the cloned the cDNA of AFS. Transient expression of AFS-GFP fusion protein in protoplast showed that AFS was localized in the chloroplast.6. Monoterpenes, sesquiterpenes, and other volatiles were found in relatively high amounts in transgenic tomato leaves, but no farnesene was detected in the both samples.