葡萄是我国主要的栽培果树之一，具有重要经济价值。优质高抗是葡萄未来育种的主要方向。传统杂交育种方式在聚合多个优异性状时效率低、时间长、成本高。作为新一代基因编辑工具，CRISPR/Cas9在葡萄育种中逐渐开始应用；然而使用拟南芥AtU3或AtU6启动子启动sgRNA表达的CRISPR/Cas9体系导致葡萄基因编辑效率较低，同时单一性状的基因编辑也制约了品种改良的效率。因此，本项目主要研究内容为：1）鉴定并克隆葡萄自身U3/U6启动子，建立启动子库，优化葡萄基因编辑体系；2）利用葡萄U3/U6启动子建立葡萄CRISPR/Cas9多基因编辑体系；3）利用构建的多基因编辑体系对葡萄果实重要品质性状糖、酸，以及抗寒等性状进行多基因编辑，快速富集葡萄糖、酸和抗寒性状，获得优质高抗的育种新材料。通过本项目的实施能够促进葡萄基因编辑的理论研究，提升葡萄育种过程中多性状聚合效率，进而加快未来葡萄育种进程。

As one of the mainly cultivated fruit crops in China, grape is economically important. Development of new cultivars with high qualities and high-multi resistance is the principle objective of grape breeding in future, but it is time-consuming, costly and inefficient to combine different desirable traits into a new cultivar through conventional breeding approach. The CRISPR/Cas9 technology, which emerges as a new and powerful genome editing tool, is gradually introduced to the filed of grape breeding these years. However, the use of Arabidopsis AtU3 or AtU6 promoters to drive the expression of sgRNA resulted in a low editing efficiency in grape. In addition, genome editing involving in single crop trait is not suitable to combine several desirable traits simultaneously considering the low efficiency. Hence, the aims of this project are (1) identifying the grape U3 and U6 promoters, which can be used to improve the genome editing efficiency in grape after experimental verification, (2) developing grape U3/U6 promoters-based CRISPR/Cas9 system that can be employed for multiplex genome editing, and (3) editing the target genes involved in sugar accumulation, organic acid synthesis, and cold resistance in grape, expecting to create new cultivar with improved grape traits rapidly. Moreover, the obtained grape cultivar with high qualities and improved cold resistance could serve as material for grape breeding. The implementation of the project would promote the theoretical study of genome editing in grape, which in turn would greatly accelerate the process of in future grape breeding by increasing the efficiency of gathering multiple elite traits of grape.