Northern blot analysis of Cre-expressing sub-lines. (a) Six sub-lines carrying the CAG-Cre-IRES-Hyg cassette were isolated from each of three double-mutant lox lines: 71/66-P, 71/JTZ-P, and 71/KR3-P. Ten micrograms of total RNA were subjected to electrophoresis and hybridization with a Cre probe. The two highest Cre-expressing lines selected from each of the six sub-lines are indicated by a circle. (b) Selected sub-lines were passaged 11 times, and RNAs from 1st, 7th, and 11th passage were prepared. Four micrograms of total RNAs were subjected to electrophoresis and hybridization with a Cre probe. The expression levels of Cre were maintained during passages.