Sperm-typing of four C57BL/6JxCAST/EiJ(HFM1-KI/KI) mice and four C57BL/6JxCAST/EiJ(HFM1-WT/WT) mice

A mutant Hfm1 allele (Hfm1-KI) was introduced in the zygote of a cross between two F1 mice deriving from hybridisations between two Mus musculus domesticus strains: strain C57BL/6J, hereafter called B6 and strain DBA/2J, hereafter called DBA2. The resulting founder mice were thus heterozygous for the Hfm1 gene (Hfm1-WT/KI) and their genetic backgrounds were composed of 50% DBA2 and 50% B6 genomes. Further crosses with other B6 and Mus musculus castaneus (strain CAST/EiJ, hereafter called CAST) mice resulted in individuals carrying either two mutant alleles for Hfm1 (Hfm1-KI/KI), two WT alleles (Hfm1-WT/WT) or one allele of each (Hfm1-WT/KI).Of these, four hfm1 homozygous mutant (28347, 28353, 28364 and 28367) and two WT (28355, 28359, 28371 and 28377) male mice were selected for further analysis: their sperm DNA was extracted and sonicated to produce fragments of a mean size of 450 bp.After collecting sperm DNA from the two individuals, we performed capture-based DNA sequencing using an assay targeting 890 recombination hotspots and 500 control loci (that do not correspond to known recombination hotspots but that display similar genomic characteristics in terms of GC-content, SNP density, sequence quality and TE content).For the efficiency of DNA capture (i.e. hybridisation-based targeted-DNA enrichment) to be identical in both haplotypes, two baits were designed for each of the 1,390 selected regions: one corresponding to the CAST haplotype and one to the B6 haplotype.Libraries were then sequenced by an Illumina device using a 250-bp paired-end protocol and reads were mapped to both the GRCm38/mm10 version of the B6 genome (ftp://ftp-mouse.sanger.ac.uk/ref/) and the CAST/EiJ draft genome (ftp://ftp-mouse.sanger.ac.uk/REL-1509-Assembly/) using BWA-MEM with default parameters and marking shorter split hits as secondary.