Abstract
Key message
CRISPR/Cas9 system can precisely edit genomic sequence and effectively create knockout mutations in T0 generation watermelon plants.
Abstract
Genome editing offers great advantage to reveal gene function and generate agronomically important mutations to crops. Recently, RNA-guided genome editing system using the type II clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) has been applied to several plant species, achieving successful targeted mutagenesis. Here, we report the genome of watermelon, an important fruit crop, can also be precisely edited by CRISPR/Cas9 system. ClPDS, phytoene desaturase in watermelon, was selected as the target gene because its mutant bears evident albino phenotype. CRISPR/Cas9 system performed genome editing, such as insertions or deletions at the expected position, in transfected watermelon protoplast cells. More importantly, all transgenic watermelon plants harbored ClPDS mutations and showed clear or mosaic albino phenotype, indicating that CRISPR/Cas9 system has technically 100% of genome editing efficiency in transgenic watermelon lines. Furthermore, there were very likely no off-target mutations, indicated by examining regions that were highly homologous to sgRNA sequences. Our results show that CRISPR/Cas9 system is a powerful tool to effectively create knockout mutations in watermelon.
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Acknowledgements
We thank Dr. Shuhua Yang for technical help. We thank Dr. Qijun Chen for providing the CRIPSR/Cas9 genome targeting vector. This work was supported by grants from National Natural Science Foundation of China (31361140355, 31401893, 31272184, 31471785 and 31301738), Beijing Ministry of Science and Technology of China (2014BAD01B09), Beijing Municipal Science and Technology Commission of China (Z161100004916081, 6141001, 2014000021223TD03), Beijing Scholar Program (BSP026), Ministry of Agriculture of China (CARS-26) and BAAFS Innovation Team (JNKYT301601).
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Communicated by N. Stewart.
S. Tian and L. Jiang contributed equally to this work.
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Tian, S., Jiang, L., Gao, Q. et al. Efficient CRISPR/Cas9-based gene knockout in watermelon. Plant Cell Rep 36, 399–406 (2017). https://doi.org/10.1007/s00299-016-2089-5
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DOI: https://doi.org/10.1007/s00299-016-2089-5