Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice
CRISPR-Cas9 is a common tool for gene editing, and appropriate sgRNAs are the key factor for successful editing. In this study, the effect of sgRNA length and number on editing efficiency was analyzed in rice using CYP81A6 as the target gene. A series of CRISPR-Cas9 plant expression vectors containi...
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| Format: | Article |
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KeAi Communications Co., Ltd.
2022-04-01
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| Series: | Crop Journal |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2214514121001537 |
| _version_ | 1811273063622246400 |
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| author | Xiaojing Liu Jiangtao Yang Yaya Song Xiaochun Zhang Xujing Wang Zhixing Wang |
| author_facet | Xiaojing Liu Jiangtao Yang Yaya Song Xiaochun Zhang Xujing Wang Zhixing Wang |
| author_sort | Xiaojing Liu |
| collection | DOAJ |
| description | CRISPR-Cas9 is a common tool for gene editing, and appropriate sgRNAs are the key factor for successful editing. In this study, the effect of sgRNA length and number on editing efficiency was analyzed in rice using CYP81A6 as the target gene. A series of CRISPR-Cas9 plant expression vectors containing single sgRNAs with different lengths (17, 18, 19, 20, 21, 22, 23 nt) or two sgRNAs were constructed and introduced into rice cultivar Zhonghua11 by Agrobacterium-mediated transformation. Analysis of the editing status of 1283 transgenic rice plants showed that 371 were successfully edited with base preference. Single A or T insertions were the most frequent among the six edited types. The editing efficiency of transgenic rice with two sgRNAs was higher than that with a single sgRNA. Editing efficiency and sgRNA length showed a normal distribution with 20 nt sgRNA (25%) being the most efficient. The editing efficiency decreased slightly with decreases of 1–2 bases (19 nt 20%, 18 nt 21%), but decreased significantly with a decrease of 3 bases (17 nt 4.5%). Editing efficiency was significantly reduced by adding 1 to 3 bases (21 nt 16.8%, 22 nt 13%, 23 nt 13%) to the sgRNA. These results provide data for successful gene editing or rice by CRISPR-Cas9. |
| first_indexed | 2024-04-12T22:52:17Z |
| format | Article |
| id | doaj.art-650d08106e3541da8eced7d0f0b8e533 |
| institution | Directory Open Access Journal |
| issn | 2214-5141 |
| language | English |
| last_indexed | 2024-04-12T22:52:17Z |
| publishDate | 2022-04-01 |
| publisher | KeAi Communications Co., Ltd. |
| record_format | Article |
| series | Crop Journal |
| spelling | doaj.art-650d08106e3541da8eced7d0f0b8e5332022-12-22T03:13:19ZengKeAi Communications Co., Ltd.Crop Journal2214-51412022-04-01102577581Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in riceXiaojing Liu0Jiangtao Yang1Yaya Song2Xiaochun Zhang3Xujing Wang4Zhixing Wang5Biotechnology Research Institute, Chinese Academy of Agricultural Sciences/MARA Key Laboratory on Safety Assessment (Molecular) of Agri-GMO, Beijing 100081, ChinaBiotechnology Research Institute, Chinese Academy of Agricultural Sciences/MARA Key Laboratory on Safety Assessment (Molecular) of Agri-GMO, Beijing 100081, ChinaBiotechnology Research Institute, Chinese Academy of Agricultural Sciences/MARA Key Laboratory on Safety Assessment (Molecular) of Agri-GMO, Beijing 100081, ChinaBiotechnology Research Institute, Chinese Academy of Agricultural Sciences/MARA Key Laboratory on Safety Assessment (Molecular) of Agri-GMO, Beijing 100081, ChinaCorresponding authors.; Biotechnology Research Institute, Chinese Academy of Agricultural Sciences/MARA Key Laboratory on Safety Assessment (Molecular) of Agri-GMO, Beijing 100081, ChinaCorresponding authors.; Biotechnology Research Institute, Chinese Academy of Agricultural Sciences/MARA Key Laboratory on Safety Assessment (Molecular) of Agri-GMO, Beijing 100081, ChinaCRISPR-Cas9 is a common tool for gene editing, and appropriate sgRNAs are the key factor for successful editing. In this study, the effect of sgRNA length and number on editing efficiency was analyzed in rice using CYP81A6 as the target gene. A series of CRISPR-Cas9 plant expression vectors containing single sgRNAs with different lengths (17, 18, 19, 20, 21, 22, 23 nt) or two sgRNAs were constructed and introduced into rice cultivar Zhonghua11 by Agrobacterium-mediated transformation. Analysis of the editing status of 1283 transgenic rice plants showed that 371 were successfully edited with base preference. Single A or T insertions were the most frequent among the six edited types. The editing efficiency of transgenic rice with two sgRNAs was higher than that with a single sgRNA. Editing efficiency and sgRNA length showed a normal distribution with 20 nt sgRNA (25%) being the most efficient. The editing efficiency decreased slightly with decreases of 1–2 bases (19 nt 20%, 18 nt 21%), but decreased significantly with a decrease of 3 bases (17 nt 4.5%). Editing efficiency was significantly reduced by adding 1 to 3 bases (21 nt 16.8%, 22 nt 13%, 23 nt 13%) to the sgRNA. These results provide data for successful gene editing or rice by CRISPR-Cas9.http://www.sciencedirect.com/science/article/pii/S2214514121001537CRISPR-Cas9sgRNA numbersgRNA lengthEditing efficiency |
| spellingShingle | Xiaojing Liu Jiangtao Yang Yaya Song Xiaochun Zhang Xujing Wang Zhixing Wang Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice Crop Journal CRISPR-Cas9 sgRNA number sgRNA length Editing efficiency |
| title | Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice |
| title_full | Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice |
| title_fullStr | Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice |
| title_full_unstemmed | Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice |
| title_short | Effects of sgRNA length and number on gene editing efficiency and predicted mutations generated in rice |
| title_sort | effects of sgrna length and number on gene editing efficiency and predicted mutations generated in rice |
| topic | CRISPR-Cas9 sgRNA number sgRNA length Editing efficiency |
| url | http://www.sciencedirect.com/science/article/pii/S2214514121001537 |
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