Enhanced Rice Blast Resistance by CRISPR/Cas9-Targeted Mutagenesis of the ERF Transcription Factor Gene OsERF922.
Rice blast is one of the most destructive diseases affecting rice worldwide. The adoption of host resistance has proven to be the most economical and effective approach to control rice blast. In recent years, sequence-specific nucleases (SSNs) have been demonstrated to be powerful tools for the improvement of crops via gene-specific genome editing, and CRISPR/Cas9 is thought to be the most effective SSN. Here, we report the improvement of rice blast resistance by engineering a CRISPR/Cas9 SSN (C-ERF922) targeting the OsERF922 gene in rice. Twenty-one C-ERF922-induced mutant plants (42.0%) were identified from 50 T0 transgenic plants. Sanger sequencing revealed that these plants harbored various insertion or deletion (InDel) mutations at the target site. We showed that all of the C-ERF922-induced allele mutations were transmitted to subsequent generations. Mutant plants harboring the desired gene modification but not containing the transferred DNA were obtained by segregation in the T1 and T2 generations. Six T2 homozygous mutant lines were further examined for a blast resistance phenotype and agronomic traits, such as plant height, flag leaf length and width, number of productive panicles, panicle length, number of grains per panicle, seed setting percentage and thousand seed weight. The results revealed that the number of blast lesions formed following pathogen infection was significantly decreased in all 6 mutant lines compared with wild-type plants at both the seedling and tillering stages. Furthermore, there were no significant differences between any of the 6 T2 mutant lines and the wild-type plants with regard to the agronomic traits tested. We also simultaneously targeted multiple sites within OsERF922 by using Cas9/Multi-target-sgRNAs (C-ERF922S1S2 and C-ERF922S1S2S3) to obtain plants harboring mutations at two or three sites. Our results indicate that gene modification via CRISPR/Cas9 is a useful approach for enhancing blast resistance in rice.
College of Agriculture, Guangxi University, Nanning, China.|National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agriculture Sciences (CAAS), Beijing, China.
Fujun
0000-0003-4426-2750
F
Wang
National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agriculture Sciences (CAAS), Beijing, China.
Chunlian
C
Wang
College of Agriculture, Guangxi University, Nanning, China.
Piqing
P
Liu
National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agriculture Sciences (CAAS), Beijing, China.
Cailin
C
Lei
National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agriculture Sciences (CAAS), Beijing, China.
Wei
W
Hao
National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agriculture Sciences (CAAS), Beijing, China.
Ying
Y
Gao
State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, College of Life Sciences, South China Agricultural University, Guangzhou, China.
Yao-Guang
YG
Liu
National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI), Institute of Crop Science, Chinese Academy of Agriculture Sciences (CAAS), Beijing, China.
Kaijun
K
Zhao
D001483:Base Sequence; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics*; D017343:Genes, Plant; D016296:Mutagenesis; D012275:Oryza / Q000235:genetics / Q000382:microbiology*; D010935:Plant Diseases / Q000382:microbiology*; D030821:Plants, Genetically Modified; D012689:Sequence Homology, Nucleic Acid; D014157:Transcription Factors / Q000235:genetics*
D014157:Transcription Factors