Affiliation:
1. Institute of Crop, Wuhan Academy of Agricultural Sciences, Wuhan 430065, China
2. Research Center of Hami Melon, Xinjiang Academy of Agricultural Sciences, Urumqi 830091, China
3. Hainan Sanya Crops Breeding Trial Center, Xinjiang Academy of Agricultural Sciences, Sanya 572019, China
Abstract
Melon (Cucumis melo L.) is a protected crop in China with high economic value. Agrobacterium-mediated genetic transformation is a powerful tool to improve agronomic traits and obtain elite germplasm. However, current transformation protocols in melons are inefficient and highly genotype-dependent. To improve transformation in melon, we tested different infiltration methods for Agrobacterium-mediated transformation. Among these methods, micro-brushing and sonication for 20 s, followed by vacuum infiltration at −1.0 kPa for 90 s, resulted in the strongest green fluorescent protein signal and increased the proportion of infected explants. We transformed melon with developmental regulatory genes AtGRF5, AtPLT5, AtBBM, AtWUS, AtWOX5, and AtWIND1 from Arabidopsis and estimated regeneration frequencies as the number of regenerating shoots/total number of inoculated explants in the selection medium. The overexpression of AtGRF5 and AtPLT5 in melon resulted in transformation efficiencies of 42.3% and 33% in ZHF and 45.6% and 32.9% in Z12, respectively, which were significantly higher than those of the control. AtGRF5 and AtPLT5 expression cassettes were added to CRISPR/Cas9 genome-editing vectors to obtain transgenic phytoene desaturase CmPDS knockout mutants. Using AtGRF5 or AtPLT5, multi-allelic mutations were observed at CmPDS target sites in recalcitrant melon genotypes. This strategy enables genotype-flexible transformation and promotes precise genome modification technologies in melons.
Funder
Hubei Key Research Project of CRISPR Gene-Editing Technology in Fruit Vegetable Crops
Wuhan Major Project of Key Technologies in Biological Breeding and New Variety Cultivation
Wuhan applied basic frontier Project
Tianshan Innovation Team Plan Project in The People’s Government of Xinjiang Uygur Autonomous Region of China
Subject
Genetics (clinical),Genetics
Cited by
4 articles.
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