Biological Function Of The Auxin Early Response Gene Mul-SAUR15 In Mulberry

Mulberry is not only an important food plant of the silkworms,but also has important medicinal,feeding and ecological value.Using genetic engineering technology to cultivate new mulberry varieties will contribute to realize the economic and ecological values of mulberry trees.As a kind of auxin early response gene,small auxin-up RNA(SAUR)gene is closely associated to the growth and development of plants,but its specific biological function and regulatory mechanism remain largely unknown.At present,there are few reports about SAUR gene of mulberry.In this study,the mulberry Mul-SAUR15 gene,which is one of the SAUR family genes was cloned for the first time,and its physiological and biochemical characteristics were analyzed.Moreover,its biological function was studied by transgenic technology.In addition,the promoter of Mul-SAUR15 gene was cloned and its expression activity was analyzed to reveal the expression profile of Mul-SAUR15 gene.The research results will provide a candidate gene for mulberry genetic engineering breeding and lay a foundation for further study on the biological function and regulatory mechanism of SAUR gene.The main results are as follows:(1)Cloning and bioinformatics analysis of Mul-SAUR15 geneAn auxin early response gene of mulberry designed as Mul-SAUR15 was cloned by PCR.The gene contains an open reading frame of 318 bp and encodes a peptide composed of 105 amino acids.The theoretical isoelectric point(pI)of Mul-SAUR15 protein is 8.98,and the predicted molecular weight(Mw)is 11.516 kDa.The protein contains more random coil(39.05%)and ?-helix(37.14%)but only a few extended strand(17.14%)and ?-turn(6.67%).Mul-SAUR15 protein does not contain transmembrane domain,O-glycosylation site,N-glycosylation site,and signal peptide,but it contains an auxin-inducible superfamily domain.Mul-SAUR15 protein has higher amino acid sequence consistency with SAUR15 proteins of other plants,and has the closest genetic relationship with SAUR15 of Morus notabilis.(2)Expression characteristics of Mul-SAUR15 geneThe promoter sequence of Mul-SAUR15 gene was cloned by PCR and named as pMul-SAUR15.pMul-SAUR15 was analyzed with the online promoter analysis tool of PlantCARE.The results showed that it contains not only the core components of promoter,but also auxin,gibberellin and a variety of environmental factor response elements.The agrobacteria-mediated transient expression test of tobacco leaves showed that pMul-SAUR15 had significant auxin,gibberellin and light induced activities.The expression vector of GUS gene initiated by pMul-SAUR15 was transformed into Arabidopsis.The results of GUS histochemical analysis showed that pMul-SAUR15 had different expression activities in different tissues or organs.Meanwhile,the expression characteristics of Mul-SAUR15 in mulberry various tissues were analyzed by fluorescence quantitative PCR and the results showed that Mul-SAUR15 was expressed in all tissues,but it expression level was different tissues among different tissues.Its expression abundance in the stem(phloem)is the highest.On the contrary,its expression abundance in the root is the lowest.(3)Biological function of Mul-SAUR15 geneThe plant expression vector of Mul-SAUR15 gene was constructed,and the transgenic Mul-SAUR15 Arabidopsis thaliana plants were successfully obtained.It was found that overexpression of Mul-SAUR15 gene in Arabidopsis could promote the elongation of hypocotyls under dark conditions and the development of roots under normal conditions.Moreover,overexpression of Mul-SAUR15 gene in Arabidopsis promoted the growth and development of transgenic plants and the transgenic plants showed enhanced tolerance to high concentrations of auxin.In addition,the transgenic Arabidopsis plants were inoculated with Pseudomonas syringae pv.Tomato DC3000(Pst.DC3000)and treated with salt stress and drought stress.It was found that the transgenic Arabidopsis had enhanced resistance to Pst.DC3000,but showed increased sensitivity to salt stress and drought stress than wild type plants,indicating that the Mul-SAUR15 gene not only plays an important role in the regulation of plant growth and development,but also was involved in the response process of plant to environmental stresses.