Study On Secondary Growth By PsnSHN2 Transcription Factor From Populus Simonii×P.nigra

SHN2(SIHNE2),a member of the plant-specific AP2/EREBP transcription factor family,is an important transcription factor which related to the formation of plant gum and wax closely and could control of cellulose synthesis,lignin synthesis and secondary cell wall biosynthesis directly or indirectly.In recent years,it has attracted more and more attention.However,the study of the SHN2 gene almost focused on the model plant Arabidopsis thaliana;and some other cash crops.So far,it has not been reported on woody plants.To help people learn more about the mechanism of secondary growth of poplars,use the PsnSHN2 gene to change the material composition of poplars and cultivate high quality pulpwood,in this research,we preliminary analysis function and expression profiles of the PsnSHN2 gene in Populus simoniixP.nigra.In this research,first,we clone the PsnSHN2 gene which homologous with AtSHN2 using PCR technology,construct the plant expression vector containing the CaMV 35S strong promoter and analyze the gene sequence.Second,construct the GFP expression vector,analyze the properties of the PsnSHN2 protein by gene gun method and transgenic Nicotiana benthamiana.Third,a real-time PCR is used to quantify the PsnSHN2 gene expression of different tissues of two-year-old Populus simonii ×P.nigra seedlings.Then,analyze PsnSHN2 and cis-acting elements of DNA interactions by Yeast one-hybrid technology,while using the PLACE web,we analyze sites in the promoter region of target genes from Populus trichocarpa.At last,we fuse the PsnSHN2 gene into Nicotiana benthamiana and observe the phenotype changes and molecular mechanism of the transgenic Nicotiana benthamiana strains which have high-level expression of the PsnSHN2 gene.The results indicate that:1.Forecast showed that the PsnSHN2 gene contained 540bp of full-length sequence,protein analysis and comparison find that Populus simonii×P.nigra has 99%similarity with Populus trichocarpa,molecular 43291.1k Da,Theoretical pI:5.21.2.PsnSHN2 encoded protein localization in the nucleus,in line with the transcription factor encoded proteins located in the nucleus of the general features;3.There was a significant difference of the PsnSHN2 gene expression among different tissues of Populus simonii ×P.nigra seedlings.In descending order,the expression level of the PsnSHN2 gene were xylem,phloem,root.4.The PsnSHN2 specifically binding to GCC-box,JREB-box and DRE category Cis-acting elements.Some secondary growth-related gene promoter region were discovered that contain GCC-box or DRE class cis-acting element by PLACE web.5.The phenotype of PsnSHN2-overexpressing plants exhibited increased height,number of leaves,length and width of leaves,stem diameter,length and cellulose content,secondary cell wall thickening etc.while improve expression about cellulose synthesis,lignin synthesis and secondary cell wall biosynthesis genes.The results suggest that the PsnSHN2 gene mainly expresses in Xylem.Throughout the growth cycle of Populus simonii ×P.nigra,it plays an important role in secondary growth.