Damak S, Jay N P, Barrell G K, Bullock D W
Centre for Molecular Biology, Lincoln University, Canterbury, New Zealand.
Biotechnology (N Y). 1996 Feb;14(2):181-4. doi: 10.1038/nbt0296-181.
To establish the feasibility of overexpressing foreign genes in the wool follicle, transgenic sheep were produced by pronuclear microinjection of a DNA construct consisting of a mouse ultrahigh-sulfur keratin promoter linked to the bacterial chloramphenicol acetyl transferase (CAT) gene. Four of 31 lambs born were transgenic. The overall efficiency of transgenesis was 1.1% of zygotes injected and transferred. Two transgenic rams were mated to nontransgenic ewes, and both transmitted the gene to their offspring in Mendelian fashion. CAT expression was found in the skin of one G0 ram and in 9 out of 26 transgenic G1 progeny. Two G1 lambs were sacrificed to study tissue specificity. Both had high levels of expression in skin but One had high expression in spleen and kidney with lower levels of expression in lung; the other had low expression in spleen, lung, and muscle. In situ hybridization demonstrated that transgene expression in the skin was confined to the keratogenous zone of the wool follicle cortex. Expression of CAT activity in skin was correlated with diet-induced or seasonal changes in the rate of wool growth. This keratin promoter appears useful for overexpressing factors in the wool follicle that might influence wool production or properties.
为了确定在毛囊中过表达外源基因的可行性，通过原核显微注射一种由与细菌氯霉素乙酰转移酶（CAT）基因相连的小鼠超高硫角蛋白启动子组成的DNA构建体来生产转基因绵羊。出生的31只羔羊中有4只是转基因的。转基因的总体效率为注射并移植的受精卵的1.1%。两只转基因公羊与非转基因母羊交配，二者均以孟德尔方式将基因传递给后代。在一只G0公羊的皮肤以及26只转基因G1后代中的9只的皮肤中发现了CAT表达。处死了两只G1羔羊以研究组织特异性。二者在皮肤中均有高水平表达，但一只在脾脏和肾脏中有高表达，在肺中的表达水平较低；另一只在脾脏、肺和肌肉中的表达较低。原位杂交表明，皮肤中的转基因表达局限于毛囊皮质的生角质区。皮肤中CAT活性的表达与饮食诱导的或羊毛生长速率的季节性变化相关。这种角蛋白启动子似乎有助于在毛囊中过表达可能影响羊毛生产或特性的因子。
