Fig. 2. Comparison of keratin expression profiles in gallbladders of different mouse keratin genotypes. (A) Gallbladders from six mice of each indicated genotype were isolated and pooled followed by high salt extraction of the relatively insoluble proteins (primarily keratins) as described in Materials and methods. The extracts were analyzed by SDS-PAGE followed by visualization of the protein bands by Coomassie Blue staining (upper panel). Arrowhead indicates position of desmin. Duplicate gels were transferred to membranes followed by blotting with the indicated anti-keratin antibodies (lower panel). (B) Gallbladders were isolated from K8-null (–/–), K8 heterozygous (+/–), or K8 WT (+/+) mice (2 mice/genotype). The gallbladders from each genotype were pooled then used to prepare total tissue homogenates that were analyzed by immunoblotting using antibodies specific to K8, K18, K19 and actin. (C) Keratin expression at the protein and mRNA levels was analyzed. (a) Gallbladders were isolated from three K19 WT and three K19-null mice, followed by analyses of equal amount of total lysate protein samples using SDS-PAGE and immunoblotting using the indicated keratin-specific antibodies. (b) Total RNA was isolated from K19-null (striped bars) or K19 WT (open bars) gallbladders (pooled, n=3 for each genotype) followed by RT-PCR and real time PCR analysis. Note that both K18 and K20 are up-regulated at the protein and mRNA levels in K19-null gallbladders.

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