CRISPR/Cas9-Mediated Genome Editing in Soybean Hairy Roots.
As a new technology for gene editing, the CRISPR (clustered regularly interspaced short palindromic repeat)/Cas (CRISPR-associated) system has been rapidly and widely used for genome engineering in various organisms. In the present study, we successfully applied type II CRISPR/Cas9 system to generate and estimate genome editing in the desired target genes in soybean (Glycine max (L.) Merrill.). The single-guide RNA (sgRNA) and Cas9 cassettes were assembled on one vector to improve transformation efficiency, and we designed a sgRNA that targeted a transgene (bar) and six sgRNAs that targeted different sites of two endogenous soybean genes (GmFEI2 and GmSHR). The targeted DNA mutations were detected in soybean hairy roots. The results demonstrated that this customized CRISPR/Cas9 system shared the same efficiency for both endogenous and exogenous genes in soybean hairy roots. We also performed experiments to detect the potential of CRISPR/Cas9 system to simultaneously edit two endogenous soybean genes using only one customized sgRNA. Overall, generating and detecting the CRISPR/Cas9-mediated genome modifications in target genes of soybean hairy roots could rapidly assess the efficiency of each target loci. The target sites with higher efficiencies can be used for regular soybean transformation. Furthermore, this method provides a powerful tool for root-specific functional genomics studies in soybean.
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China; National Center for Transgenic Research in Plants, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Yupeng
Y
Cai
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China; National Center for Transgenic Research in Plants, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Li
L
Chen
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China; National Center for Transgenic Research in Plants, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Xiujie
X
Liu
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Shi
S
Sun
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Cunxiang
C
Wu
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Bingjun
B
Jiang
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Tianfu
T
Han
Ministry of Agriculture Key Laboratory of Soybean Biology (Beijing), Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China; National Center for Transgenic Research in Plants, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, 100081, China.
Wensheng
W
Hou
D001483:Base Sequence; D064113:CRISPR-Cas Systems / Q000235:genetics*; D005818:Genetic Engineering; D005822:Genetic Vectors; D018745:Genome, Plant / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D018517:Plant Roots / Q000235:genetics* / Q000254:growth & development; D030821:Plants, Genetically Modified / Q000235:genetics* / Q000254:growth & development; D017393:RNA Editing / Q000235:genetics*; D012689:Sequence Homology, Nucleic Acid; D030262:Soybean Proteins / Q000235:genetics*; D013025:Glycine max / Q000235:genetics* / Q000254:growth & development
D030262:Soybean Proteins