A targeting vector was designed to replace the endogenous intein cleavage-cholesterol attachment site (Shh processing site) in exon 3 of the targeted gene with a Green Fluorescent Protein (GFP) followed by a new intein cleavage-cholesterol attachment site (Shh processing site), as well as insert a frt-flanked PGK-Neo cassette in intron 2. The donating investigator reports that this construct was microinjected into 129X1/SvJ-derived AV3 embryonic stem (ES) cells, correctly targeted ES cells were injected into recipient blastocysts, and chimeric mice were bred to C57BL/6 inbred mice to generate Shh::gfp mice. These Shh::gfp mice were reported to have been bred with C57BL/6 mice for a small number generations prior to sending to The Jackson Laboratory Repository in 2008 (see SNP notes below). Upon arrival, these mice were bred to C57BL/6J inbred mice (Stock No. 000664) to establish the colony. Over the next two years, the colony was backcrossed to C57BL/6J and/or wildtype mice from the colony for several additional generations. By March 2011, the colony was found to be ~96% C57BL/6 genetic background by SNP analysis (see SNP notes below).
In 2009, a 27 SNP (single nucleotide polymorphism) panel analysis performed on the rederived living colony at The Jackson Laboratory Repository. This revealed 16 markers (across several different chromosomes) that were not C57BL/6 allele-type. These data suggest the strain was sent to The Jackson Laboratory Repository with multiple sources of genetic background contamination (more than 129 genetic background).
In 2011, a 32 SNP (single nucleotide polymorphism) panel analysis, with markers covering all 19 chromosomes and the X chromosome, was performed on the colony at The Jackson Laboratory Repository. This revealed one marker on chromosome 1 (~89.1 Mbp) that was not fixed for C57BL/6 allele-type (e.g.: still segregating for 129X1 allele-type).
