关键词: 肥大软骨细胞, Cre重组酶, 转基因小鼠, 组织特异性

Abstract:

Hypertrophic chondrocytes, which are the terminally differentiated form of chondrocytes, play a key role in endochondral ossification. In order to investigate the functions of hypertrophic chondrocytes during bone development, we generated a new transgenic line expressing Cre recombinase under the control of a 8.2 kb mouse type X collagen gene promoter (Col10a1(8.2)-Cre). Microinjection was employed to introduce the 11.5 kb transgenic fragment into 328 oocytes, from which 51 progenies were obtained. Three mice carrying the transgene in genome were identified by PCR genotyping. PCR detected expression of Col10a1(8.2)-Cre transgene within tissues containing hypertrophic chondrocytes. To examine the activity and specificity of Cre recombinase in vivo, transgenic line was crossed with ROSA26 report line. As indicated by LacZ staining, ROSA26; Col10a1(8.2)-Cre double transgenic mice showed efficient expression of Cre recombinase within hypertrophic chondrocytes. In situ hybridization analyses further confirmed the transcription of Col10a1(8.2)-Cre transgene within the upper zone of hypertrophy, indicating a better activity and specificity in contrast to the previously constructed Col10a1(1.0)-Cre transgenic line. These results showed that this Col10a1(8.2)-Cre transgenic line could be used as a powerful tool to achieve conditional gene knockout in hypertrophic chondrocytes