CRISPR/Cas9 has been widely used for precise and efficient genome editing in a number of organisms, notably in major crops, for gene function characterization and trait improvement. However, the application of this genome editing tool is very limited in Brassica oleracea, especially in broccoli (Brassica oleracea var. italica). Here, we report that the genome of broccoli, a worldwide important Brassicaceae vegetable, can also be precisely edited by a CRISPR/Cas9 system with multiple single-guide RNA-expressing cassettes. The gene BoGL5, previously identified as a candidate for the glossy green mutant, was chosen as the target gene. We generated 14 T0 transgenic plants, among which 14.3% and 50% produced mutations at the expected position in targeted regions 1 and 2, respectively. Two of them with a 100% mutation rate showed a glossy green phenotype and dramatically reduced cuticular wax load. These results indicate that CRISPR/Cas9 can be used as a promising technique for gene functional verification and trait improvement in broccoli.

CRISPR/Cas9 已广泛用于许多生物体的精确和高效的基因组编辑，特别是在主要作物中，用于基因功能表征和性状改良。然而，这种基因组编辑工具在Brassica oleracea中的应用非常有限，尤其是在西兰花 ( Brassica oleracea var. italica ) 中。在这里，我们报道了西兰花的基因组，这是一种世界上重要的十字花科植物，也可以通过具有多个单向导 RNA 表达盒的 CRISPR/Cas9 系统进行精确编辑。BoGL5基因，先前被确定为有光泽的绿色突变体的候选者，被选为目标基因。我们生成了 14 株 T0 转基因植物，其中 14.3% 和 50% 分别在目标区域 1 和 2 的预期位置产生了突变。其中两个具有 100% 突变率的小鼠表现出有光泽的绿色表型，并显着减少了表皮蜡质负荷。这些结果表明，CRISPR/Cas9 可作为一种很有前途的技术用于西兰花的基因功能验证和性状改良。