A Simple Heat Treatment Increases SpCas9-Mediated Mutation Efficiency in Arabidopsis
| Autor: | Shunya Hirata, Hafizur Rhaman, Naoshi Yamanaka, Shuta Kurokawa, Tsuyoshi Aiso, Chisato Ishizaki, Mayuka Yamamoto, Shaikhul Islam, Hidetaka Kaya, Kappei Kobayashi |
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| Rok vydání: | 2021 |
| Předmět: |
Hot Temperature
Physiology Population Meristem Arabidopsis Plant Science medicine.disease_cause Genome Genome editing CRISPR-Associated Protein 9 medicine Arabidopsis thaliana education Promoter Regions Genetic Gene Genetics Gene Editing education.field_of_study Mutation biology food and beverages Cell Biology General Medicine biology.organism_classification Plants Genetically Modified CRISPR-Cas Systems Genome Plant |
| Zdroj: | Plantcell physiology. 62(11) |
| ISSN: | 1471-9053 |
| Popis: | The CRISPR/Cas9 system is now commonly employed for genome editing in various plants such as Arabidopsis, rice and tobacco. In general, in genome editing of the Arabidopsis genome, the SpCas9 and guide RNA genes are introduced into the genome by the floral dip method. Mutations induced in the target sequence by SpCas9 are confirmed after selecting transformants by screening the T1 seed population. The advantage of this method is that genome-edited plants can be isolated easily. However, mutation efficiency in Arabidopsis using SpCas9 is not as high as that achieved in rice and tobacco, which are subjected to a tissue culture step. In this study, we compared four promoters and found that the parsley UBIQITIN promoter is highly active in Arabidopsis meristem tissue. Furthermore, we examined whether a simple heat treatment could improve mutation efficiency in Arabidopsis. Just one heat treatment at 37°C for 24 h increased the mutation efficiency at all four target sites from 3 to 42%, 43 to 62%, 54 to 75% and 89 to 91%, without detectable off-target mutations. We recommend heat treatment of plate-grown plants at 37°C for 24 h as a simple method to increase the efficiency of CRISPR/Cas9-mediated mutagenesis in Arabidopsis. |
| Databáze: | OpenAIRE |
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