Transcriptome Analysis Under Drought Stress And The Cloning And Expression Analysis Of MP5CS Gene In Mulberry (Morus L.)

Mulberry (Morus alba) is an economically important plant used for sericulture. Chinahas the most abundant mulberry germplasm resources. However, the research of genes andgenome from mulberry is in a backward state compared with other plants, and the exactfunction of some functional genes and their encoded proteins under stress conditions inmulberry is still not fully understood.The growth and productivity of mulberry is adverselyaffected by abiotic and biotic stresses. Efforts to investigate the functional genes and theirmolecular adaptation mechanisms under stresses plays an important roles in strengtheningstress tolerance and improving production of mulberry, and are of fundamental importanceto the preservation of these genetic resources. Therefore, the research of functional genes inmulberry is an urgent and meaningful work.In our research, we cloned the P5CS gene involved in the drought resistancemechanism of mulberry based on the homology cloning, and carried out the analysis of thesequence. Furthermore, we conducted the stress expression patterns of the gene by themeans of stress-induced method based on the prediction of their functions. In the secondpart, a large-scale RNA sequencing (RNA-seq) of mulberry (Morus L.) was carried outbetween two samples in regular and drought stress condition. Then seven unique genesshowing different expression level in control and drought stress group were subsequentlyanalyzed and identified by real-time PCR.The main results were summarized briefly asfollows:1. Cloning, sequence analysis and induced expression studies of P5CS gene inmulberry.Drought is one of the important factor limiting plant growth.1-Pyrroline-5-Carboxylate Synthase is the key enzyme closely related in the synthesis ofProline under drought stress. Cloning and function analysis of mulberry gene P5CS has avery important significance. In this report, we cloned the P5CS gene of mulberry by meansof homology cloning, RT-PCR and RACE (rapid amplification of cDNA ends), and we gotthe full sequence of cDNA named MP5CS(GenBank accession number：KC202259).Homology and phylogenetic analysis of P5CS were carried out using Clustal X andMEGA4.1software. In addition, the expression level of P5CS in drought and salt stresseswere performed by qRT-PCR. Result shows that the expressiong level of P5CS isup-regulated.2. Drought stress RNA-seq of mulberry and identification of differential expression genesSolexa RNA-seq was carried out for transcriptome analysis of mulberry between twosamples in regular and drought stress condition. Data analysis was performed including denovo assembly, differential expression analysis, database annotation, GO functionalclassification, KEGG pathway mapping and annotation analysis of SSR repeat. Total54,736contigs were got from the reads including the scaffolded regions. As determined byGO annotation and KEGG pathway mapping, database annotation was also performed.1,051genes were identified that were significantly differently expressed between twosamples and33,285microsatellites were identified from assembled unique sequences.QRT-PCR was carried out to detect the expression level of seven different genes that have adifferent expression in drought stress and control samples.