Construction of PGRP-S-/- mice. (A) Genomic organization of PGRP-S+/+ and PGRP-S-/- loci with 3 PGRP-S exons, genomic clones, targeting vector, and replacement of exon 1 with NEO cassette in PGRP-S-/- mice. Relevant restriction sites, primers, and screening probes are also shown. (B) Genotypes of PGRP-S-/-, PGRP-S+/+, and PGRP-S+/- mice determined by typing of genomic DNA by PCR using either PGRP-specific or NEO cassette–specific sense primer and intron-specific (S1) antisense primer. DNA from homozygous PGRP-S-/- mice yielded only 320-bp NEO PCR fragment and no PGRP fragment, demonstrating the replacement of PGRP-S exon 1 with NEO cassette in both chromosomes. DNA from wild-type PGRP-S+/+ mice yielded only 283-bp PGRP PCR fragment and no NEO fragment, whereas DNA from heterozygous PGRP-S+/- mice yielded both PGRP and NEO PCR fragments. (C) Phenotypes of PGRP-S-/- and PGRP-S+/+ mice determined by Western blot: lysates of 2 × 106 bone marrow cells/lane were separated by SDS-PAGE, and PGRP-S protein was detected by anti–PGRP-S antibodies on Western blots in lysates from PGRP-S+/+ but not from PGRP-S-/- mice.
