Molecular Cloning And Expression Of Tomato Flower Pedicel Abscission Specific Gene SlPP2C And Obtaining The Transgenic Tomato Plants

Tomato(Solarium lycopersicum)is one of the largest cultivated horticultural crops in the world,but the flower abscission and fruit abscission is seriously restricted its yield,seriously damage it's planting benefits,so it has a great significance to study the abscission of tomato flower pedicel.Studies have shown that protein phosphatase 2C may affect the abscission of tomato flowers.In previous study,we identified a SlPP2C gene while sequencing the transcriptome of tomato flower pedicel abscission,which gene expression increasing gradually during the flower pedicel abscission,so this experiment uses "zhongshu 6" as the test material to study the effect of SIPP2C gene on the tomato flower pedicel abscission in vitro.Investigating abscission rate of tomato flower pedicel explants in the protein phosphatase 2C inhibitor treatment.Meanwhile the characteristic of SIPP2C gene,encoding protein,promoter sequence and gene expression pattern is studied.Construct SIPP2C gene silencing and promoter GUS expression vectors,The vectors is transferred into the cultivated tomato,respectively.This transferred tomato laied the foundation of the study in tomato flower pedicel abscission.The main results in this research are as follows:1.By investigating abscission rate of tomato flower pedicel explants in protein phosphatase 2C inhibitor EDTA and NaF treatments,the results show the rate of flower pedicel abscission is lower than that of the control,we conclude that protein phosphatase 2C significantly inhibits the flower pedicel abscission.2.The full length and the promoter sequence of the SIPP2C gene is successfully obtained.Bioinformatics analysis shows that SlPP2C has the same conserved domain as other PP2C and the promoter sequence has a variety of putative cis-element responeding to light,drought,heat,IAA,ABA,JA,GA with online promoter analysis software.3.The expression level of SIPP2C gene is analized with qRT-PCR,the relative expression of SlPP2C gene is increasing gradually and the expression in the abscission zone is higher than that of non abscission zone during the flower pedicel abscission,suggesting that the SIPP2C gene may participate in the flower pedicel abscission,which may accelerate flower pedicel abscission.Exogenous hormones IAA,ABA,ETH and 1-MCP are used to treat flower pedicel explants.The treatment of IAA and 1-MCP inhibit the abscission,while ABA and ETH promote the abscission of flower pedicel.4.The recombinant expression vector of SlPP2C-RNAi and ProPP2C::GUS was constructed,and are introduced into the "zhognshu 6" tomato by using the Agrobacterium-mediated genetic transformation on the basis of the established tomato efficient regeneration system.PCR,RT-PCR and qRT-PCR was used to identify the TO lines silencing positive transgenic tomato,and access to obtain TO lines of SlPP2C silencing positive transgenic tomato.The tomato lines grow slowly and their leaves are dark green,a little round and more thicker.The flower pedicel remove difficultly and it's flower pedicel rate decreases compared to the wild type tomato.The SlPP2C promoter positive transgenic tomato has been identified by GUS result and access to have TO lines ProPP2C::GUS.transgenic tomato.The two transgenic plants would provided important experimental materials for the further study of the SlPP2C role in the tomato flower pedicel abscission.In this paper,we conclusion that the S1PP2C gene is specifically expressed in the flower abscission zone,which promotes the process of flower pedicel abscission and obtain the S1PP2C silencing and ProPP2C::GUS transgenic tomato plants,which provide important experimental material and basis for the further study of the function of SlPP2C.