Functional Analysis Of PtrFLA31/34 In Populus Trichocarpa

Fasciclin-like arabinogalactan proteins(FLAs)are a subclass of arabinogalactan proteins(AGPs).The number of mumbers of the FLA gene family varies.According to the previous research in our laboratory,46 FLA genes in the genome of Populus trichocarpa were found,and two numbers(PtrFLA31 and PtrFLA34)with high transcriptional level and specificlly expressed in xylem were selected.We constructed the cas9/gRNA double mutation vector of PtrFLA31/34,and it is speculated that this group of genes is involved in the secondary growth of populus.Therefore,in order to explore the function of PtrFLA31/34 genes,a series of confirmatory experiments were carried out on the young Ptrfla31/34 mutants,and the results showed that this group of genes does have a certain regulatory effect on the xylem development of the stem.This laid the foundation for the study of PtrFLA31 and PtrFLA34 gene functions,and provided the basis for the biological functions of other FLA genes in plants.The main research results are as follows:Phenotypic observation showed that compared with WT,the ptrfla31/34 mutant had a 13%increase in plant height,the length and width of stems slightly increased,and the length of leaves increased by 8%,which suggested that this group of genes could affect the vegetative growth of poplar.Cross section staining observations revealed that the cells in the xylem region of the same stem ptrfla31/34 mutants were larger than WT and had more xylem cell layers.The fiber and vessel cell sizes were measured by isolation.The results showed that the changes in the height and stem diameter of the ptrfla31/34 mutants were caused by the increase in xylem fiber and vessel cell size.It shows that knocking out the two genes PtrFLA31 and PtrFLA34 caused expanding of xylem cells,which did affect the development of stem xylem to a certain extent.The scanning electron microscopy results of the stems of young ptrfla31/34 mutants were consistent with the results of section staining.Compared with the WT,the mutant plants had more xylem cell layers in the same stem,and the thickness of the secondary cell wall in the transgenic line was not significantly thickened or thinned.The results of the xylem chemical components determination showed that compared with WT,the lignin content of ptrfla31/34 mutant increased by 8%and the cellulose content of ptrfla31/34 mutant decreased by 7%.This indicates that PtrFLA31 and PtrFLA34 may play a role in the synthesis of lignin and cellulose.These results further prove that PtrFLA31 and PtrFLA34 may affect the xylem development of the stem.It was found in the corresponding tensile response analysis of the young trees of the ptrfla31/34 mutant that compared with the WT,there were no significant changes in the xylem cells of the ptrfla31/34 mutant reaction wood and the tension wood.This indicates that PtrFLA31/34 may not respond to the formation of strains.