Procedure for chromosome mapping. (A), Method. Typically 30 mutant animals (homozygous Bristol DNA surrounding the mutation) and 30 wild-type animals (heterozygous Bristol/Hawaiian or homozygous Hawaiian DNA) are lysed in 20 μL lysis buffer. The lysate is then added to a PCR mix lacking primers, and the mix is aliquoted into every other row of a 96 well plate. Primers are added by pin replication from a master plate. Because the 8-channel pipette loads every other lane of the gel, each mutant reaction is placed next to its control. DNA ladder is typically placed in lanes 17 and 34. (B), Results from homozygous N2 Bristol and CB4856 Hawaiian genotypes. 50 Bristol adults and 50 Hawaiian adults were lysed in 20 μL lysis buffer, and used for the DNA template for the 48 PCR reactions covering all six chromosomes. Note that pure Bristol and Hawaiian DNA was used for each PCR reaction in the gel shown. When mapping a recessive mutant in the Bristol background against the Hawaiian strain, unlinked SNPs will display a 50-50 mix of Bristol bands and Hawaiian bands in both mutant and non-mutant lanes. Linked SNPs will display an enrichment of Bristol bands in the mutant lane, approaching 100% Bristol for tight linkage. The non-mutant lane will display a 2/3 to 1/3 enrichment of Hawaiian compared to Bristol DNA.
Davis et al. BMC Genomics 2005 6:118 doi:10.1186/1471-2164-6-118