Fig. 1.

Knock-in mice expressing nonfarnesylated versions of lamin B1 and lamin B2. (A) Gene-targeting strategy to create the Lmnb1^CS allele, in which the cysteine (Cys⁵⁸⁵) of the CaaX motif of Lmnb1 is replaced with a serine (Ser⁵⁸⁵). A new restriction endonuclease site, SacI, was introduced into adjacent sequences to facilitate genotyping. Exons are depicted as black boxes; the 3′ UTR is white; loxP sites are represented by black arrowheads. Primer locations for 5′ (black) and 3′ (gray) long-range PCR reactions are indicated with arrows. (B) Gene-targeting strategy to create the Lmnb2^CS allele, in which the cysteine (Cys⁵⁹³) of the CaaX motif of Lmnb2 is replaced with a serine (Ser⁵⁹³). A SalI site was introduced in adjacent sequences. (C) The 5′ long-range PCR with the Lmnb1^CS allele yields an 8.2-kb fragment (lanes 2 and 3). The identity of the PCR product was confirmed by SacI digestion; the SacI fragments (taking into account the newly introduced SacI site) were 2.4, 2.2, 2.0, 1.1, and 0.6 kb in length (lanes 4 and 5). No PCR product was amplified from wild-type DNA (lane 6). (D) The 3′ long-range PCR with the Lmnb1^CS allele yields a 4.5-kb fragment (lanes 2 and 3). No PCR product was amplified from wild-type DNA (lane 4). (E) With the Lmnb2^CS allele, the 5′ long-range PCR yields an 8.7-kb fragment. The identity of the PCR product was verified by digestion with SalI (with the newly introduced SalI site, the expected products were 5.8 and 2.9 kb) (lanes 4 and 5). No PCR product was obtained with wild-type DNA (lane 6). (F) The 3′ long-range PCR with the Lmnb2^CS allele yields a 5.2-kb fragment (lanes 2, 4, 5, 7, and 8). No PCR product was obtained with wild-type DNA (lanes 3, 6, and 9).