Exploration Of CRISPR Genetic Transformation System And Transfer Verification Of Flowering Gene FT/TFL1

The flower transformation process of higher plants is the basis for the continuation of species reproduction,and is regulated by the signals of plants inside and outside.Among them,FT and TFL1 are important genes in the flowering regulatory pathway of plants,and they are at the core of the flower-regulating network.This study used superexpression and CRISPR gene knockout technology to study the role of FT and TFL1 genes in citrus development.Grafting is used to verify the signalling of flower-related genes,In order to change the status of citrus juvenile period,cultivate new germplasm of early flower or late flowering citrus.The results of this study are:1.Based on the research rules of CRISPR/Cas9 system in herbaceous plants,try to establish an application system in citrus.The transformation and regeneration process was optimized by Agrobacterium-mediated genetic transformation of the epiphyseal axis.The statistical results showed that the growth time of the sterile buds was 42d,the concentration of OD₆₀₀00 was 0.8,the concentration of As was 30mg/L,and Explants used a slitting method can effectively improve the regeneration rate of resistant buds.2.Using the overexpression vector 35S::Pt FT-EGFP to genetically transform the cultivated tobacco to obtain overexpressing PtFT tobacco.PtFT and CiTFL1transgenic tobacco T2 generation were planted,and it was found that PtFT can significantly promote the flowering period of tobacco,and promote the development of plant side branches and increase the number of flowering and fruiting.It was found that CiTFL1 was differentially expressed in different tissues of Poncirus trifoliata L.Raf and was able to significantly delay the flowering time of cultivated tobacco.3.Wild type tobacco?WT?was grafted onto the rootstock of transgenic tobacco?PtFT,CiTFL1?,and the related traits of tobacco plants after grafting were observed and analyzed.The corresponding early flower or late flower phenotype transmission of transgenic tobacco rootstocks could not be found.No PtFT and CiTFL1 mRNA molecules were detected in the scion phloem at 0.5 cm on the graft interface,and mRNA of citrus plants FT and TFL1 may not move between the herbaceous tobacco rootstock and the scion.