Fig. 2. Phenotype of Lig 1 gene targeted mice and expression from the targeted alleles. (A) Morphology of wild-type (left) and Lig 1^{-(#12)/-(#12)} (right) embryos at E16.5. Note that the mutant embryos are smaller, anaemic and lack the erythropoiesis clearly evident in the wild-type foetal liver (^*). (B) Northern analysis of Lig 1 transcripts. RNA (30 µg) extracted from primary embryonic fibroblasts (wild-type, heterozygous and homozygous for the Lig 1^-(#12) and Lig 1^-(#53) targeted alleles) was probed with a 2.1 kb fragment of mouse Lig 1 cDNA (Bentley et al., 1996). The position of the 3.2 kb wild-type Lig 1 mRNA is indicated. Note the larger (4.1 kb) transcript from the Lig 1^-(#12) allele, whereas a transcript is undetectable from the original Lig 1^-(#53) allele by northern analysis. Equivalent RNA loadings were confirmed by ethidium bromide staining of the gel prior to transfer. (C) Western analysis of DNA ligase I in wild-type and homozygous Lig 1^-(#53) mutant embryos. Protein extracted from E13.5 embryos was probed with the TL5 rabbit polyclonal antibody raised against purified bovine DNA ligase I. This antibody predominantly recognises epitopes in the N-terminus of the protein. 100 µg of protein was loaded in each lane, but in addition to pure wild-type and mutant extracts, mutant extracts were also spiked with the percentages indicated of the wild-type sample. The position of the 102 kDa wild-type DNA ligase I protein is indicated by the arrow. The mobility of M_r markers (kDa) on the same gel is shown.