Expression of Cre recombinase in alveolar epithelial cells of the AQP2-Cre transgenic mini-pigs.
OBJECTIVE
Optimal use of Cre mediated recombination in conditional animal models depends on well characterized Cre driver lines. Unfortunately, some Cre driver lines exhibit unexpected expression patterns hindering their utility in Cre/loxP systems. Thus, systematic assessment of new Cre lines is essential for generating useful Cre driver lines for future studies.


METHODS
Here, we describe a Cre Transgenic (Tg) mini-pig line in which the expression of Cre is directed by a 3-kb 5' fragment of the kidney-specific aquaporin 2 (AQP2); however, the AQP2-Cre Tg mini-pig line exhibits expression of Cre in alveolar epithelial cells (AECs) instead of collecting duct cells. The specificity of the AQP2-Cre plasmid was validated in vitro, and indicating that the AQP2-Cre was specifically expressed in the transfected LLC-PK1 cells. Absolute quantitative real-time PCR (qRT-PCR) and inverse PCR were performed to determine the copy numbers and integration sites of the AQP2-Cre transgene. Relative qRT-PCR was performed to evaluate variation in Cre expression levels over time.


RESULTS
Our data indicated that this AQP2-Cre Tg mini-pig line exhibits stable expression of Cre recombinase over time and in subsequent generations, even though the AQP2-Cre transgene was segregated and reduced in subsequent generations.


CONCLUSIONS
Combined with our previous studies of the activity of this Cre, we conclude that this Cre Tg mini-pig line will provide a reliable tool for generating lung-specific gene targeting mini-pig models, thereby allowing the investigation of gene functions in lung development and studying the molecular mechanisms of human lung disease.
College of Animal Science, Jilin University, Changchun, China.
Weiwei
W
Luo
Zhanjun
Z
Li
Peng
P
Li
Yongye
Y
Huang
Yang
Y
Han
Chaogang
C
Yao
Zhongzhan
Z
Zhang
Honggang
H
Yan
Daxin
D
Pang
Hongsheng
H
Ouyang
Li
L
Li
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics* / Q000378:metabolism; D051399:Aquaporin 2 / Q000235:genetics* / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D005260:Female; D019426:Integrases / Q000235:genetics* / Q000378:metabolism; D008297:Male; D023421:Models, Animal; D011650:Pulmonary Alveoli / Q000378:metabolism*; D013552:Swine; D013556:Swine, Miniature / Q000235:genetics* / Q000378:metabolism
D051399:Aquaporin 2; C045073:Cre recombinase; D019426:Integrases