| Sweet potato(Ipomoea batatas)is an important food,feed,vegetable,industrial raw material and new energy crop in the world.Improving the yield of sweet potato has become one of the main directions of sweet potato breeding.MADS-box gene is a very important group of transcription factors,it plays an important role in the process of plant growth and development.We cloned eight MADS-box genes by using transcriptome data and I.trifida genome database in sweet potato(Xu22).The real-time quantitative RT-PCR was used to study their expression characteristics.Abiotic stress and hormone treatments were performed on sweet potato seedlings.Real time quantitative RT-PCR was used to analyze the responses of eight MADS-box genes to abiotic stresses and hormones.A MADS-box gene IbMBP4,which might be related to the storage root development and stress resistance of sweet potato was cloned.And an over-expressed construct was created and transformed into tomato,in order to analyze the function of IbMBP4.The main results are as follows:(1)Eight MADS-box genes were screened by using transcriptome data and I.trifida genome database.The eight genes were cloned by using the mixed cDNA templates(leaf,stem,root)of sweet potato(Xu 22).(2)The basic properties of eight proteins were analyzed by bioinformatics.The subcellular localization of the MADS-box genes was predicted.The results showed that the eight genes were all located in the nucleus,which was consistent with the subcellular localization of MADS-box gene.We compared the protein sequences of eight genes by DANMAN.The results showed that the similarity in MADS and K regions was high.MEGA5.1 and Cluatal X were used to construct the phylogenetic tree of eight genes,the results showed that eight genes belonged to six different MADS-box subfamilies.(3)The expression of eight MADS-box genes in different organs and tissues,including leaves,stems and storage roots of sweet potato were studied by real-time quantitative RT-PCR.The expression of IbMBP4 gene was significantly increased at the initial stage of storage root development,and its expression was decreased with the development of storage root,this suggests that IbMBP4 gene might be involved in the development of tuberous roots.(4)Abiotic stress treatments(including drought,NaCl,low temperature and high temperature)were conducted on sweet potato seedlings.We found that the eight MADS-box genes exhibited different expression in response to to different treatments.The results showed that,the high salt and drought induced the expression of IbMBP4 sigenificantly,suggesting that the gene might be involved in the process of salt and drought tolerance.(5)In order to further verify the results of abiotic stress treatment,and to analyze the function of MADS-box gene in the development of sweet potato tuberous roots,we conducted the hormone treatments,including hormones related to stress resistance(abscisic acid(ABA),brassinolide(BR)and jasmonic acid(JA))and hormones related to the growth of tuberous root(indole acetic acid(IAA),gibberellic acid(GA)and zeatin(ZT)).Real time quantitative RT-PCR analysis revealed that the eight MADS-box genes showed different response to different hormones.IbMBP4 showed a strong response to ABA,JA and BR.It further indicated that this gene may be involved in the process of stress resistance in sweet potato.In addition,IAA and ZT,which are related to the development of tuberous roots,greatly induced the expression of IbMBP4,which further indicated that the gene may regulate the formation and expansion of sweet potato tuberous roots.(6)To further analyze the function of IbMBP4,we constructed the RNAi and over-expressed vectors of IbMBP4 gene,and the function of IbMBP4 in tomato was analyzed based on tomato genetic transformation system.The results showed that IbMBP4 affected the stress related genes(SlCAT1 and SlPR5)and root development related genes(IbEXP1 and IbAGPL1)greatly.These results indicated that the gene might regulate the stress response and the development of roots. |
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