A targeting vector was designed to replace exon 2, including the initiation codon, of the programmed cell death 1 ligand 2 (Pdcd1lg2) gene with an enhanced green fluorescent protein (EGFP) and a loxP-flanked neomycin resistance (neo) cassette. The construct was electroporated into (129X1/SvJ x 129S1/Sv)F1-Kitl+-derived R1 embryonic stem (ES) cells, and chimeras were made by ES cell aggregation. The donating investigator reported that chimeric mice were bred to CMV-Cre mice on a BALB/c background - with the intention of removing the neo cassette (genotyping later determined it retained the neo cassette). The B7-DC KO colony was backcrossed at least nine generations to BALB/c mice (and the CMV-Cre was removed). Following this, the donating investigator reported they mice were backcrossed for at least 12 generations to C57BL/6NCrl (see SNP note below), and then sent to The Jackson Laboratory. Upon arrival, mice were bred to C57BL/6NJ (Stock No. 005304) for at least one generation to establish the colony. Genotyping performed at The Jackson Laboratory determined the B7-DC KO allele retained the neo cassette from the original construct.
A 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the rederived living colony at The Jackson Laboratory Repository. While 1 of the 27 markers throughout the genome suggested a C57BL/6 genetic background, 3 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating. These data suggest the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6J ; C57BL/6N genetic background.
