A loxP site flanked targeting vector containing a loxP site and FRT site flanked PGK-neo cassette. This selection cassette was inserted downstream of exon 8 of the targeted gene, and another loxP site was inserted upstream of exon 8. This construct was electroporated into 129P2/OlaHsd derived E14 embryonic stem (ES) cells. The resulting chimeric mice were crossed to FLP recombinase expressing transgenic mice (on a 129X1/SvJ x B6SJLF1 mixed background) to remove the selection cassette. Correctly targeted ES cells were injected into recipient blastocysts. The donating investigator reports that mice were then backcrossed to 129X1/SvJ (obtained from RCC in Fullinsdorf, Switzerland) for more than 40 generations (see SNP results below) prior to sending albino males to The Jackson Laboratory Repository in 2011. Upon arrival, males were bred with 129X1/SvJ females (Stock No. 000691) for one generation to establish the live colony. Mice homozygous for the mutation were then bred together to maintain the live colony.
A 27 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, was performed on the F3 generation living colony at The Jackson Laboratory Repository. This revealed 5 of 27 markers, one each on chromosomes 2, 7, 11, 15 and 17, that were not fixed for 129 allele-type (e.g., still segregating for C57BL/6;SJL allele-type markers). These data suggest the mice were backcrossed to 129X1/SvJ for fewer generations than reported prior to arrival at The Jackson Laboratory Repository, and the mice are most likely now a mix of 129 (~80%) and C57BL/6;SJL (~20%).
