Multiplex CRISPR/Cas9 Mutagenesis of BrVRN1 Delays Flowering Time in Chinese Cabbage ( Brassica rapa L. ssp. pekinensis )

Joon Ki Hong, Eun Jung Suh, Sang Ryeol Park, Jihee Park and Yeon-Hee Lee
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Joon Ki Hong: Agricultural Biotechnology Department, National Institute of Agricultural Sciences, Rural Development Administration, 370 Nongsaengmyeong-ro, Jeonju 54874, Korea
Eun Jung Suh: Agricultural Biotechnology Department, National Institute of Agricultural Sciences, Rural Development Administration, 370 Nongsaengmyeong-ro, Jeonju 54874, Korea
Sang Ryeol Park: Agricultural Biotechnology Department, National Institute of Agricultural Sciences, Rural Development Administration, 370 Nongsaengmyeong-ro, Jeonju 54874, Korea
Jihee Park: Agricultural Biotechnology Department, National Institute of Agricultural Sciences, Rural Development Administration, 370 Nongsaengmyeong-ro, Jeonju 54874, Korea
Yeon-Hee Lee: Agricultural Biotechnology Department, National Institute of Agricultural Sciences, Rural Development Administration, 370 Nongsaengmyeong-ro, Jeonju 54874, Korea

Agriculture, 2021, vol. 11, issue 12, 1-15

Abstract: The VERNALIZATION1 ( VRN1 ) gene is a crucial transcriptional repressor involved in triggering the transition to flowering in response to prolonged cold. To develop Chinese cabbage ( Brassica rapa L. ssp. pekinensis ) plants with delayed flowering time, we designed a multiplex CRISPR/Cas9 platform that allows the co-expression of four sgRNAs targeting different regions of the endogenous BrVRN1 gene delivered via a single binary vector built using the Golden Gate cloning system. DNA sequencing analysis revealed site-directed mutations at two target sites: gRNA1 and gRNA2. T 1 mutant plants with a 1-bp insertion in BrVRN1 exhibited late flowering after the vernalization. Additionally, we identified ‘transgene-free’ BrVRN1 mutant plants without any transgenic elements from the GE 1 (gene-editing 1) and GE 2 generations. All GE 2 mutant plants contained successful edits in two out of three BrVRN1 orthologs and displayed delayed flowering time. In GE 2 mutant plants, the floral repressor gene FLC1 was expressed during vernalization; but the floral integrator gene FT was not expressed after vernalization. Taken together, our data indicate that the BrVRN1 genes act as negative regulators of FLC1 expression during vernalization in Chinese cabbage, raising the possibility that the ‘transgene-free’ mutants of BrVRN1 developed in this study may serve as useful genetic resources for crop improvement with respect to flowering time regulation.

Keywords: Brassica rapa; BrVRN1; CRISPR/Cas9; late flowering; transgene-free; vernalization (search for similar items in EconPapers)
JEL-codes: Q1 Q10 Q11 Q12 Q13 Q14 Q15 Q16 Q17 Q18 (search for similar items in EconPapers)
Date: 2021
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Citations: View citations in EconPapers (1)

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