| The anthocyanin played an important role in the red or purple colour of the plant tissue and would be accumulated when the plants suffered from stress,such as low temperature and high light condition,because of which we can treat it as a signal of stress that the plants suffer from.With the development of transcriptome sequencing technology,it is possible to explore the different and similar parts between the low temperature and high light stress on anthocyanin biosynthesis,respectively.In this study,the transcriptome sequencing was performed on the leaves of ‘Olmpia' Begonia semperflorens in control(CK),low temperature(LT)and high light(HL)conditions.Then we got 21.09 Gb Clean Data,and there were 74,779 and 83,699 unigenes in LT and HL treatment and the average length of the uningene were 684 and 640 pb,respectively.In the low temperature treatment,the N50 of transcript and unigene were 2,200 and 1,249 pb,while the value in high light treatment was 2,113 and 1,098 pb.We also do the analysis of genetic structure of uningene and got 6,567 SSR markers.The uningenes were all annotated in Nr,Swiss-Prot,KEGG,COG and GO database.Furthermore,many of the differentially expressed genes were related to factors associated with anthocyanin metabolism,transportation and regulation,which influenced the expression of leaf color.We also analysis the enrichment pathway,and discuss the difference in mechanisms of anthocyanin induced under low temperature and high light conditions.This study is the first time to explore broad-scale gene expression level in B.semperflorens.By identifying DEGs regulated by both LT and HL conditions,we found that anthocyanin accumulation was employed as a common strategy by Begonia seedlings in resisting LT and HL stress.By identifying DEGs regulated differently by LT and HL conditions,we found that Begonia seedlings also had some different strategies for resisting LT and HL stresses:anthocyanins were biosynthesized under HL condition,while lignin,proanthocyanidins and anthocyanins under LT condition.Our study was the first research of the transcriptome sequencing in Begonia semperflorens.Based on the transcriptome sequencing results,we found that the Bs Rboh D gene was significantly up-regulated while the expression of the PAL?4CL?CHS?F3H?ANS ? 3GT gene were up-regulated.So we got the sequence of Bs Rboh D from the transcriptome sequencing of B.semperflorens.Then we identified its sequence through many bioinformatics analyses and investigated its chemicophysical properties.We found that the full length gene of Bs Rboh D weselected was 3796 bp and the graph of CDS was 2784 pb.We used the q RT-PCR method to analyze the different expression level of Bs Rboh D,Bs PAL,Bs CHS,Bs F3 H and Bs ANS in B.semperflorens at different conditions.Furthermore,we measured the H2O2,O2.-,chlorophyll and anthocyanin content of them.All the result indicated that the expression of the Bs Rboh D started being up-regulated in low temperature at 3h and reached the first peak at 9h,while the Bs PAL,Bs CHS,Bs F3 H and Bs ANS started being up-regulated at 5h,7h,7h,9h,and up to the first peak at 9h,respectively.In the treatment group of CK,LT,LT+NADH,LT+DPI and LT+DMTU,the trend of H2O2,O2-,chlorophyll and anthocyanin content of them was LT+NADH>LT>LT+DPI>LT+DMTU.And the relative expression level of Bs Rboh D,Bs PAL,Bs CHS,Bs F3 H and Bs ANS were all up-regulated in the treatment group of CK,LT,LT+NADH,LT+DPI and LT+DMTU and the trend of them positively correlated with the change of H2O2,O2.-,chlorophyll and anthocyanin content.In the CK?MV and H2O2 treatment group,the all indexes of MV and H2O2 were significantly up-regulated compared with CK treatment.The above analysis show that the environment stress such as low temperature and high light condition could promote the expression of Bs Rboh D and increase the content of ROS and anthocyanin in the leaves of B.semperflorens.The expression of Bs Rboh D and the ROS which was induced by Bs Rboh D was a signal that could promote the anthocyanin biosynthesis. |
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