The Roles Of External Environment And Chemicals In Anthocyanins Biosynthesis Of Red Leaf Peach In Vitro Based On RNA-Seq

Peach(Prunus persica),originates from China,is a very important fruit tree of Rosaceae prunus.Edible peach is the main body of the development.There are many types and varieties for edible.Besides edible peach,red-leaf peach(Prunus persica f.atropurpurea)as a member of peach for ornamental,having a great application prospect in garden,because of its red leaf in spring,green leaf in summer,and dark red leaf in autumn.Our laboratory had found in physiological that red-leaf peach and purple-leaf plum color rendering is caused by the accumulation of anthocyanins,but the types of anthocyanins are not alter in different seasons.And a large number of studies have also shown,the external environment,chemicals,endogenetic and exogenous hormones also play an important role in regulating the biosynthesis of anthocyanins.In order to further study on mechanism of red-lef peach color changing,transcriptome sequencing technology was used to study differentially expressed genes at the transcriptional level taken red-leaf peach ’tsukuba-5’ as the experimental material.Deal with leaves in vitro in September,expression analysis of anthocyanins in the process of synthesis structure gene(CHI,UFGT)and regulation(MYB10、MYB15、bHLH33 and WD40)expression level by qRT-PCR.The main results were as follows:1.Using RNA-Seq to get complete coding region information of 27865 genes,including 5467 genes are undetected in Peach gene sequencing and annotation,which belong to the newly discovered genes.Comparativing analysis of the leaves in May and July,we get 2738 differentially expressed genes;Contrasting analysis in May and September,we get 2419 differentially expressed genes;Comparativing analysis in July and September,there are 1540 differentially expressed genes.GO function analysis shows that the genetic difference between cells,organelles,cell metabolism,catalytic activity,cell physiological process and combination,etc.KEGG annotation shows that differentially expressed genes of higher frequency involved in carbohydrate metabolism,lipid metabolism and other secondary metabolites biosynthesis.Sequencing results also find that temperature,illumination,IAA,ABA,ethylene,jasmonic acid and salicylic acid related genes expressed significant difference in leaf growth period,and speculate that has certain contact with anthocyanins synthesis of leaf.Also find that UFGT(ppa005162m),CHI(ppa011476m)and MYB protein 4(ppa010227m)may be have a promoting effect in anthocyanins synthesis of peach leaf.2.According to the result of RNA-Seq,the leaves of ’Tsukuba No.5’ peach in vitro was as the test materials,using the qRT-PCR to analysis related gene(CHI and UFGT)expression level of anthocyanins biosynthesis via different concentrations of temperature,light and hormone treatment.The result showed,low temperature prormotes the expression level of CHI gene,and inhibits UFGT,in contrast to the high temperature;Light promote CHI UFGT gene expression;Hormone(ABA,IAA,JA and SA)treatments can promote CHI and UFGT gene within a certain amount of concentration and time.But have the sensitivity differently of the two genes to the hormone.ABA processing of three concentration can improve CHI gene expression,and the longer the better,but 50(μM and 100μM promoted UFGT express,150 μM high concentration suppressed UFGT gene expression;50 μM IAA treatment can obviously promote the CHI and UFGT gene expression,100 μM and 150 μM high concentration is inhibiting factors of CHI and UFGT expression;100 μM JA can increase the expression level of two genes;The most suitable concentration of SA was 50 μM,and for UFGT is 100 μM.3.The leaves of Tsukuba No.5’ peach in vitro was as the test materials,using the qRT-PCR to analysis related gene expression level of anthocyanins biosynthesis via different concentrations of NaCl,sucrose and KH2PO4 treatment.The result showed,low concentration(100 and 200 mM),short time(4 h)of NaCl treatment can all improve the expression level of CHI,UFGT,MYB10,bHLH33 gene;0.3%,8 h of cane sugar processing promoted expression of UFGT,MYB10 bHLH33 gene significantly;0.2%,8 h of KH2PO4 treatment also have a significant role in promoting all genes expression,besides MYB15.The effect of CHI gene expression is more obvious with the increased concentration and the extended time of sucrose and KH2PO4 treatments,in a certain range.Three exogenous substances have significant inhibitory effect on MYB15 and WD40,in addition to the 0.2%,8 h KH2PO4 treatment improved the WD40 transcription.