Analysis of Mx1-Cre, LSL-Kras^G12D, 5A3^fl/+, and Mx1-Cre, Nf1^fl/fl, 5A3^fl/+ compound mutant mice. (A) Comparison of the white blood count (A left, P = .692) and hemoglobin (A right, P = 0.120) of Mx1-Cre, LSL-Kras^G12D, 5A3^+/+ (+/+), Mx1-Cre, LSL-Kras^G12D,5A3^fl/+(fl/+), and control Mx1-Cre negative littermates (control) at death after treatment with pIpC at weaning. (B) Kaplan-Meier survival curve of Mx1-Cre–negative (control; n = 9), Mx1-Cre, LSL-Kras^G12D, 5A3^+/+ (+/+, n = 8), and Mx1-Cre, LSL-Kras^G12D5A3^fl/+ (fl/+, n = 7) mice (P = .136). pIpC treatment at 21 days is indicated with arrow. (C) Southern blot analysis of genomic DNA from the bone marrow (BM) of a Mx1-Cre, LSL-Kras^G12D, 5A3^fl/+ mouse showing the presence of the deleted allele. (D) Kras^G12D expression and deletion of the 5A3 region in the individual CFU-GM colonies. (Top) Bone marrow cells from a Mx1-Cre, LSL-Kras^G12D5A3^fl/+ mouse were plated in methylcellulose medium without added cytokines. CFU-GM colonies that formed after 7 days were picked and lysed, and the presence of the deleted 5A3 allele was confirmed by PCR. Asterisks indicate the presence of the recombination PCR product. (Bottom) Cre-mediated recombination of the inhibitory LSL cassette 5′ of the Kras^G12D gene was verified in the same cells (*) by PCR. The 285-bp product is indicative of the wt Kras locus, whereas the 315-bp product is specific for excised LSL-Kras^G12D allele. The unrearranged LSL-Kras^G12D allele does not amplify. PCR conditions were confirmed with peripheral blood DNA from Mx1-Cre, LSL-Kras^G12D5A3^fl/+ (K) and wt (WT) mice. (E) Kaplan-Meier survival curve of Mx1-Cre, Nf1^fl/fl, 5A3^+/+ (+/+, n = 7) and Mx1-Cre, Nf1^fl/fl, 5A3^fl/+ (fl/+, n = 9) mice. P = .403. (F) Southern blot of genomic DNA from the peripheral blood of a Mx1-Cre, Nf1^fl/fl, 5A3^fl/+ mouse showing the presence of the deleted allele.