CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>)
Chinese cabbage has unintended bolting in early spring due to sudden climate change. In this study, late-bolting Chinese cabbage lines were developed via mutagenesis of the <i>BrLEAFY</i> (<i>BrLFY</i>) gene, a transcription factor that determines floral identity, using the c...
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MDPI AG
2022-12-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/24/1/541 |
| _version_ | 1797625632521191424 |
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| author | Yun-Hee Shin Young-Doo Park |
| author_facet | Yun-Hee Shin Young-Doo Park |
| author_sort | Yun-Hee Shin |
| collection | DOAJ |
| description | Chinese cabbage has unintended bolting in early spring due to sudden climate change. In this study, late-bolting Chinese cabbage lines were developed via mutagenesis of the <i>BrLEAFY</i> (<i>BrLFY</i>) gene, a transcription factor that determines floral identity, using the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) system. Double-strand break of the target region via gene editing based on nonhomologous end joining (NHEJ) was applied to acquire useful traits in plants. Based on the ‘CT001’ pseudomolecule, a single guide RNA (sgRNA) was designed and the gene-editing vector was constructed. <i>Agrobacterium</i>-mediated transformation was used to generate a Chinese cabbage line in which the sequence of the <i>BrLFY</i> paralogs was edited. In particular, single base inserted mutations occurred in the <i>BrLFY</i> paralogs of the LFY-7 and LFY-13 lines, and one copy of T-DNA was inserted into the intergenic region. The selected <i>LFY</i>-edited lines displayed continuous vegetative growth and late bolting compared to the control inbred line, ‘CT001’. Further, some <i>LFY</i>-edited lines showing late bolting were advanced to the next generation. The T-DNA-free E<sub>1</sub><i>LFY</i>-edited lines bolted later than the inbred line, ‘CT001’. Overall, CRISPR/Cas9-mediated mutagenesis of the <i>BrLFY</i> gene was found to delay the bolting time. Accordingly, CRISPR/Cas9 is considered an available method for the molecular breeding of crops. |
| first_indexed | 2024-03-11T09:59:03Z |
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| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-11T09:59:03Z |
| publishDate | 2022-12-01 |
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| series | International Journal of Molecular Sciences |
| spelling | doaj.art-90274fca191f49e398f56f816ea8a3542023-11-16T15:35:02ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672022-12-0124154110.3390/ijms24010541CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>)Yun-Hee Shin0Young-Doo Park1Department of Horticultural Biotechnology, Kyung Hee University, 1732 Deogyoung-daero, Giheung-gu, Yongin-si 17104, Gyeonggi-do, Republic of KoreaDepartment of Horticultural Biotechnology, Kyung Hee University, 1732 Deogyoung-daero, Giheung-gu, Yongin-si 17104, Gyeonggi-do, Republic of KoreaChinese cabbage has unintended bolting in early spring due to sudden climate change. In this study, late-bolting Chinese cabbage lines were developed via mutagenesis of the <i>BrLEAFY</i> (<i>BrLFY</i>) gene, a transcription factor that determines floral identity, using the clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (CRISPR/Cas9) system. Double-strand break of the target region via gene editing based on nonhomologous end joining (NHEJ) was applied to acquire useful traits in plants. Based on the ‘CT001’ pseudomolecule, a single guide RNA (sgRNA) was designed and the gene-editing vector was constructed. <i>Agrobacterium</i>-mediated transformation was used to generate a Chinese cabbage line in which the sequence of the <i>BrLFY</i> paralogs was edited. In particular, single base inserted mutations occurred in the <i>BrLFY</i> paralogs of the LFY-7 and LFY-13 lines, and one copy of T-DNA was inserted into the intergenic region. The selected <i>LFY</i>-edited lines displayed continuous vegetative growth and late bolting compared to the control inbred line, ‘CT001’. Further, some <i>LFY</i>-edited lines showing late bolting were advanced to the next generation. The T-DNA-free E<sub>1</sub><i>LFY</i>-edited lines bolted later than the inbred line, ‘CT001’. Overall, CRISPR/Cas9-mediated mutagenesis of the <i>BrLFY</i> gene was found to delay the bolting time. Accordingly, CRISPR/Cas9 is considered an available method for the molecular breeding of crops.https://www.mdpi.com/1422-0067/24/1/541<i>Brassica rapa</i>CRISPR/Cas9floral identitylate bolting<i>LEAFY</i> gene |
| spellingShingle | Yun-Hee Shin Young-Doo Park CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>) International Journal of Molecular Sciences <i>Brassica rapa</i> CRISPR/Cas9 floral identity late bolting <i>LEAFY</i> gene |
| title | CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>) |
| title_full | CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>) |
| title_fullStr | CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>) |
| title_full_unstemmed | CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>) |
| title_short | CRISPR/Cas9-Mediated Mutagenesis of <i>BrLEAFY</i> Delays the Bolting Time in Chinese Cabbage (<i>Brassica rapa</i> L. ssp. <i>pekinensis</i>) |
| title_sort | crispr cas9 mediated mutagenesis of i brleafy i delays the bolting time in chinese cabbage i brassica rapa i l ssp i pekinensis i |
| topic | <i>Brassica rapa</i> CRISPR/Cas9 floral identity late bolting <i>LEAFY</i> gene |
| url | https://www.mdpi.com/1422-0067/24/1/541 |
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