Function analysis of different Cas9 promoters on the efficiency of CRISPR/Cas9 system in soybean

The CRISPR/Cas9 system has been widely used in plants and animals as an efficient gene editing system.Several Cas9 promoters,such as RPS5A and YAO,have been reported to improve the efficiency of gene editing in CRISPR/Cas9 system.In soybean,the influence of different Cas9 promoters on the efficiency of the CRISPR/Cas9 gene-editing system has not been elucidated.In this study,six efficient promoters with known functions（p35S,pGmRPS5Ab,pGmRPS5Ac,pAtRPS5A,pGmYAO,and pZmUbiquitin） and one endogenous soybean promoter with unknown function（pGmHE） were selected to construct the CRISPR/Cas9 knockout vectors.The editing efficiency of the Cas9 promoters on the endogenous soybean genes GmSPA1a and GmEID1 was tested by Agrobacterium tumefaciens mediated hair roots system,which indicated that soybean endogenous promoter pGmRPS5Ab had the highest editing efficiency.The editing efficiency of pAtRPS5A,p35S,and pZmUbiquitin were higher than that of pGmYAO and pGmRPS5Ac.Further analysis of the sequencing maps of target sites s