A targeting vector containing neomycin resistance and herpes simplex virus thymidine kinase genes was used to disrupt exons 1 and 2, which encode the signal peptide and the first 39 amino acids. The construct was electroporated into 129S2/SvPas-derived D3 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6 blastocysts. The donating investigator reported that the resulting male chimeric animals were crossed to C57BL/6 female mice, and then backcrossed to C57BL/6J mice for 6 generations using a speed congenic protocol (see SNP note below).
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 the 27 markers throughout the genome suggested a C57BL/6 genetic background, all 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 C57BL/6N genetic background.
