| Poplar is an important renewable source and a major model plant for tree research.Succinate dehydrogenase(SDH)is the binding enzyme of the mitochondrial inner membrane.Recent studies show that SDH acts as an indirect modulator of superoxide production by complex Ⅰ and Complex Ⅲ.Complex Ⅰ and Complex Ⅲ are the production sites of reactive oxygen species(ROS)in mitochondria.Therefore,SDH plays an important role in the generation of ROS in mitochondria and is the source of reactive oxygen species in plants,which is involved in regulating plant development and stress response.In this study,SDH gene of poplar was taken as the research object,and its primary structure,secondary structure,domain and gene expression were analyzed.Gateway technology was used to construct 35S::PagSDH4 overexpression vector,and leaf disk method was used to construct PagSDH4 overexpression transgenic poplar(PagSDH4-OE).Through phenotypic analysis and anatomical analysis,this paper preliminarily revealed its role in poplar growth and development,thus providing an important reference for relevant regulatory networks in wood formation.The main research results are as follows:(1)BLAST was used to obtain SDH gene family homologous sequences of Populus trichocarpa and Arabidopsis thaliana from popgenie database.the phylogenetic tree analysis showed that there were 12 SDH genes in poplar,and the SDH in Populus alba×P.glandulosa was named as PtrSDH1～PtrSDH7,and the SDH in silver gland poplar(Populus alba × P.glandulosa,84K poplar)was named as PagSDH1～PagSDH7.Bioinformatics analysis of the SDH family of Populus chinensis was carried out.Subcellular localization analysis showed that PtrSDH1～PtrSDH4 and PtrSDH6 genes were located in mitochondria,PtrSDH5 genes were located in chloroplasts,and PtrSDH7 genes were located in the nucleus.Physicochemical analysis showed that PtrSDH3 was a hydrophobic protein,and other PtrSDH were hydrophilic proteins.Domain analysis results showed that PtrSDH1 and PtrSDH4 mainly contained FAD_binding domains,while other PtrSDH contained FAD_binding,MAM33.HSP70 and other domains.PtrSDHl and PtrSDH4 genes are evolutionarily conservative,which is reflected in stable protein properties and structure,gene structure conservation,evolutionary distance conservation and functional domain conservation.(2)Tissue specific expression of PtrSDH gene was analyzed through AspWood website of poplar.The results showed that PtrSDH1-2 and PtrSDH4 genes were highly expressed in cambium and xylem.Therefore,the corresponding homologous genes PagSDH1-2a and PagSDH4 in Populus alba × P.glandulosa were selected for fluorescence quantitative PCR to analyze their expressions in different tissues of poplar.The results showed that the expression levels of PagSDH1-2a and PagSDH4 were the highest in young stems,and gradually decreased with the maturity of stem tissues.Both PagSDH1-2a and PagSDH4 were highly expressed in young leaves,but decreased in mature leaves.At the same time,both PagSDH1-2a and PagSDH4 are highly expressed in cambium and xylem,and the expression level of PagSDH4 in xylem and cambium is higher than that of PagSDH1-2a.Therefore.PagSDH4 gene is selected for further study.(3)PagSDH4 gene was cloned from 84K poplar,PagSDH4 overexpression vector was constructed,and PagSDH4-OE transgenic poplar was obtained by Agrobacterium tumefaciens mediated leaf disk transformation.PagSDH4-OE-1 and PagSDH4-OE-4,which had the highest expression,were selected for phenotype observation.It was found that PagSDH4-OE poplar showed tall,robust and large leaves,and the plant height to ground diameter ratio of 84K was significantly increased in the control group without transgenic.The results of tissue sections showed that the xylem width and number of catheters of PagSDH4-OE were higher than those of the control group 84K,indicating that overexpression of PagSDH4 promoted xylem differentiation and development of plants.These results indicated that overexpression of PagSDH4 gene could promote xylem cell differentiation in poplar vascular tissue and significantly increase the number of xylem conduit cells in plants.In conclusion,this study identified the structure and function of PtrSDH gene in Populus sinicum,analyzed the role of PagSDH4 gene in the growth and development of poplar,and found that overexpression of PagSDH4 gene can promote xylem differentiation and development of poplar,providing important clues for the construction of relevant regulatory networks in wood formation.At the same time,it provides theoretical reference for forest genetic breeding. |
