UCH-L1基因敲除小鼠的饲养、繁殖及鉴定

FEEDING,BREEDING,AND IDENTIFYING UCH-L1 GENE KNOCKOUT MICE

目的饲养、繁殖和鉴定泛素羧基末端水解酶L1(UCH-L1)基因敲除小鼠,获得UCH-L1基因敲除纯合子小鼠,为深入研究UCH-L1的生物学功能奠定基础。方法从美国购进的UCH-L1基因敲除小鼠经适应性饲养1周后,将1只雌性杂合子和1只雄性杂合子小鼠合笼饲养并繁殖,剪取子代小鼠尾尖部0.5 cm组织,提取基因组DNA,PCR方法鉴定其基因型。雄性杂合子与雌性杂合子交配生产后代中,野生型和UCH-L1基因敲除纯合子小鼠用于后续的生物学功能研究;杂合子小鼠用于繁殖。观察UCH-L1基因敲除纯合子小鼠的表型变化,并利用免疫印迹法验证UCH-L1基因敲除的效果。结果小鼠成功繁殖后,利用杂合子交配共生产370只小鼠,其中UCH-L1+/+占34.1%、UCH-L1+/-占50.5%、UCH-L1-/-占15.4%,成功获得UCH-L1基因敲除纯合子小鼠,并建立了饲养、繁殖和鉴定方法。结论采用雌性和雄性UCH-L1基因敲除杂合子小鼠交配,不仅可以获得UCH-L1基因敲除纯合子小鼠,而且有利于长期饲养与繁殖。

Objective To obtain UCH-L1 gene knockout homozygous mice by feeding, breeding, and identifiying ubiquitin carboxyl-terminal hydrolase L1 （ UCH-L1 ） knockout mice, so as to lay the foundation for fur- ther study of the biological function of UCH-L1. Methods Seven UCH-L1 gene knockout mice purchased from the United States were fed for one week to adapt new circumstance, then one female heterozygous mouse and one male heterozygous mice were put into a cage for mating and breeding. The tissue of tail end （0.5 cm in length） of the offspring was clipped to extract the genomic DNA, and PCR method was used for identification of the genotype. The offspring of wild-type（ UCH-L1＋/-） ,and UCH-L1 gene knockout homozygous（ UCH-L1-/-） mice were used for sub- sequent biological function study; the heterozygous （UCH-L1 ＋/-） mice for breeding. The phenotypic changes of UCH-L1 gene knockout homozygous mice were observed, and th Western blotting. Results Mice purchased from abroad was succe e UCH-L1 gene knockout effect was verified by ssfully reproduced. A total of 370 filial generation mice were reproduced;and genotypic identification revealed that the percentage of UCH-L1 ~/~ mice was 34.1%, of UCH-L1 ~/- mice 50.5% ,and of UCH-L1 / mice 15.4%. UCH-L1 gene knockout homozygous mice were obtained successfully. The methods for feeding, breeding, and identification of UCH-L1 gene knockout mice were established. Conclusion The usage of male knockout homozygous mice, but and female UCH-L1 heterozygous mice for mating, not only can get UCH-L1 gene also can conduce to the long-term feeding and breeding