Preliminary Study On Anthocyanin Biosynthesis Of Rubber Tree(Hevea Brasiliensis)

Genetic improvement and gene function characterization of rubber tree urgently need genetic transformation system with high efficiency;however,the selection efficiency of current transformation system is low.Anthocyanin is a simple,safe and naked visual selection marker.However,due to its bad university between species,the tissue accumulated anthocynin being unpredicted and over-accumulation being negative effect on plant development,anthocyanin is not widely utilized for plant transformation until now.In order to establish a visual tracking system for rapid screening of transgenic rubber plants using anthocyanin as selection marker.The main researches and results are as follows:1.Isolation and identification of anthocyanin:the rubber tree anthocyanins from different tissues were extacted by 3mol/1 hydrochloric acid solution,found that anthocyanins can be extracted in the all tested tissues,and subsequent analysis showed that cyanidin is main anthocyanin synthesyed in rubber tree.2.Cloning and RT-PCR analysis of structural gene related to anthocynanin synthesis:28 anthocyanin structural genes and its homologous genes were cloned by transcriptome innotation and genome anlignment.And qRT-PCR analysis found HbCHS,NbF3H,HbF3’H,HbLDOX and HbUFGT were closely correlated with the content of anthocyanins,indicating that these genes may be critical to anthocyanin biosynthesis in rubber tree.3.Stimimulation of anthocyanin accumulation in somatic embryo:140 mg L-1MeJA could successfully induce rubber tree somatic embryos anthocyanin accumulation,and cyanidin is the main molecue accumulated in rubber tree after MeJA stimulation.RT-PCR analysis showed overexpression of genes in jasmonic acid biosynthesis pathway and anthocyanin biosynthetic pathway was obviously changed,indicating that the accumulation of anthocyanins is induced by jasmonic acid.4.Screening R2R3-MYB critical to anthocyanin accumulation in rubber tree:114 R2R3-MYBs were analyed by RT-PCR,showing that 5 were critical to anthocyanin biosynthesis and 2 had conserved motif of anthocyanin synthesis.Overall,this study provided a good basis for establishment of visual selection system for rubber tree transformation based on anthocyanin accumulation.