Abstract: Fasciclin-like arabinogalactan proteins(FLAs) play important roles in growth and development of plants. Based on bioinformatic analysis, we identified 46 FLAs in Populus trichocarpa. By phylogenetic analysis, Populus FLA family was classified into four Sub-groups(A-D) and PtrFLA31 and PtrFLA34 were in same small branch of the tree. By semi-quantitative RT-PCR analysis, PtrFLA31 and PtrFLA34 were specifically expressed in xylem on the high transcriptional level. In order to study their functions in wood formation, we knocked out PtrFLA31/34 double genes via Cas9/gRNA, and obtained three fla31/34 double mutants.