The Initiation And Activity Of Vascular Tissues In Populus Tomentosa Carr.

Chinese white poplar(Populus tomentosa Carr.)is one of the main wood species in northern China.It has the advantages of fast growth and fertility,and can be used as building materials,paper making and biological energy,and the carbon sink in the ecosystem,butthe structureof poplar wood is relatively loose,low hardness and compressive ability is poorer,so it belongs to the cork,these factors have restricted the poplar cultivation application more widely.Different cross sections were taken from the top of the cuteness that grew from 30 cm in April and observe the continuous development of the vascular tissue.In this study,we explored the cellular and molecular mechanism underneath the initiation and activity of vascular cambium in P.tomentosa,by immunohistochemistrical methods,bioinformatics analysis and gene expression change techniques.Finally six pectase genes may be found who have regulated the changed of the wood hardness(Potri.014G117100.1,Potri.008G104800.1,Potri.003G076900.1,Potri.004G128900.1,Potri.018G068400.1,Potri.007G015700.1).Which has certain scientific significance for further research on the utilization and development of poplar wood.The main conclusions are as follows:(1)Examination of serial sections of apex and stem tissues showed that the procambium–cambium meristematic continuum in P.tomentosa could be subdived into provasular,initiating layer,metavasular tissue and secondary vascular tissue,similar to P.deltoids.The procambium is derived from the residual meristem in the form of acropetally developing strands.(2)Twenty four members in pectin methylesterase(PME)gene family have been found through the bioinformatics analysis on PME family in the poplar.The locating information of the chromosome distribution showed that all other members(Potri.006G120100,Potri.006G137100,Potri.001G364200,Potri.001G365700,Potri.006G186000,Potri.006G186100,Potri.012G114900,Potri.012G112800,Potri.004G156300,Potri.T007200,Potri.004G128900,Potri.012G115200,Potri.004G033700)were located in the segment repeat or tandem repeat regions except 12 genes(Potri.002G145700,Potri.003G076900,Potri.005G012800,Potri.007G015700,Potri.008G104800,Potri.013G008100,Potri.014G117100,Potri.015G110700,Potri.016G017700,Potri.017G054700,Potri.018G068400,Potri.T007200).Therefore,it was predicted that the twelve gene members might come from the retrotransposition.The gene function enrichment analysis was performed on PME gene family in the poplar through GOEAST.KEGG pathway analysis showed that the gene family mainly gathered and distributed in the pentose-glucuronic acid conversion passageway and the metabolic pathway of starch and sucrose.The phylogenetic analysis showed that the poplar shared the most similarity with the PME gene family in arabidopsis.(3)In this study,we used a panel of monoclonal antibodies(JIM5,JIM7,LM5 and LM6)to map the distribution of pectic epitopes in different developmental stages of poplar vascular tissues.The monoclonal antibodies JIM5 and JIM7 are often used to identify the HGA of low and high methyl degrees,respectively.The monoclonal antibody LM5 is generated to recognize the(1?4)-?-d-galactan which presents in RG-I side chains,and LM6 reacts specifically with(1?5)-?-l-arabinan.The results indicate that JIM5 genes were marked less in locations where cell divisions were active because most JIM 5 genes were found in corners of differentiated xylem or phloem cells.With the acropetal development of stems,these highly methylated pectins were continuously demethylated by methyl esterase to produce low methyl ester pectin which were retained when the cells lost the division activity and gradually differentiated to specific cell types.In the secondary vascular tissues,there were nearly no JIM5 signals in the new tangential walls coming from the periclinal division in the cambium region.On the contrary,strong JIM5 signals could be detected in the cell walls of the secondary phloem,especially at the corners.For the xylem side,as the secondary wall was thickening,JIM5 signals in the primary wall weakened significantly.In the newly formed xylem cells without thickening secondary cell walls,JIM5 signals were merely detected in the tangential cell walls and part of radial cell walls,and there were only JIM5 signals in the corners between cells.LM5 always marked the continuous development from protoxylem to secondary xylem,and LM6 always marks from primary phloem to secondary phloem.(4)The change of pectic distribution take place when periclinal divisions appear within a procambial trace,so JIM5,LM5 and LM6 can be used as the markers to distinguish procambium from initiating layer.The difference is also present in their derivatives.The LM6 epitopes occurred in all cell walls of primary xylem,but absent from the tangential walls of secondary xylem cells.The LM5 epitopes occurred only in mature primary phloem,but in secondary phloem,its distribution kept constant.Constantly,LM5 epitopes occurred in cell walls of both primary and secondary xylem while LM6 epitopes occurred in cell walls of both primary and secondary phloem.These antibodies have been used in the studies of secondary vascular systems and provided evidence that the walls of phloem and xylem cells differ in their pectin composition even at a very early stage of commitment.(5)To further analyze the relationship between the different methyl degree of HGA and the expression of PME genes family,the tangential cryo-sectioning was used to separate poplar vascular tissue samples of different development stages.To obtain genome-wide insights on the transcriptome changes in the vascular tissue development,we used high-throughput RNA sequencing(RNA-seq)to characterize cDNA of vascular tissues from the six group samples.Finally,the PME gene was screened for regulate and control vascular tissues development.Three PMEs(Potri.014G117100.1,Potri.008G104800.1,Potri.003G076900.1)appear to be more highly expressed in the primary vascular tissuethan in the secondary vascular tissue,while the other three(Potri.004G128900.1,Potri.018G068400.1 and Potri.007G015700.1)appear to be express everywhere,but have higher expression level in the secondary vascular tissue.