羊毛的细度与密度存在显著负相关，研究羊毛毛囊在皮肤上的空间排布规律对于羊毛细度育种具有重要意义。DKK2 的转基因过表达可降低毛囊密度，由此推测DKK2 表达量的下降会引起细毛羊毛囊密度的增加，从而降低羊毛细度。为验证这一假说，本项目拟利用最新基因打靶技术Cas9 切刻酶系统，针对敖汉细毛羊DKK2 基因，以pX335 载体为骨架，设计并构建多对特异性向导 RNA 表达载体，通过与SSA 双荧光素酶报告系统共转染敖汉细毛羊胎儿成纤维细胞，筛选出效率较高的DNA靶标。利用优选的特异性向导 RNA 表达载体对敖汉细毛羊胎儿成纤维细胞实施DKK2 基因敲除，旨在通过体细胞核移植的途径获得基因修饰动物。

A highly negative correlatiom exsits between wool fiber diameter and wool follicle density. The study of the spacing pattern of wool follicle.is highly relevant for fine wool sheep breeding. Transgenic DKK2 overexpression reduces hair follicle density. Hence we speculated that interfollicular distances would decrease if the production of DKK2 was reduced, then the wool diameter would become lower. In order to test this hypothesis, we plan to apply Cas9 nickase system for DKK2 gene targeting in Aohan fine wool sheep. Using pX335 plasmid as a backbone, several pairs of sgRNA will be designed and constructed. By the help of SSA report system, the best sgRNA pairs will be picked up and used for gene knock-out experiment using sheep fetal fibroblast cells. The positive cell clones will be used to generate genetic modified animals through nuclear transfer.