Genetic position of SNPs used for mapping. The genetic position of each SNP was obtained from Wormbase Release 143.

Interval mapping. Individual recombinants are singled from heterozygotes and the animal, or a representative sample of their progeny, are placed in wells of a 96-well PCR plate and lysed. The plate may also contain three control wells, with Bristol, Hawaiian, and a 50-50 mix of animals. DNAs from the lysed animals are pin-replicated into a PCR master mix containing primers for the desired SNP. The plates are processed for PCR amplification, digested with DraI and samples run on an agarose gel.

SNP mapping two synthetic unc-31 suppressor mutations. A, EG5296 ox300; unc-31(e928) interval mapping data. B, EG5297 dpy-5(e61); ox305; unc-31(e928) interval mapping data. The two suppressor mutations were separated from KY5029 and the suppressing activities were separately mapped to chromosome I and chromosome II, respectively. Each row illustrates the results from a single recombinant animal, and each colored box represents the genotype of a recombinant at the indicated SNP. Bristol is represented by blue, heterozygotes by purple, Hawaiian by red, and PCR failures by gray. F12B6 is wormbase allele snp_F12B6[1], F28H1 is wormbase allele snp_F28H1[1], T10D4 is a polymorphism identified by the St. Louis SNP consortium[5] (see Methods).

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.

SNP mapping dpy-5(e61). A, chromosome mapping. Each pair of lanes shows results from the SNP at the indicated genetic map position, using either the Dumpy (D) or the wild-type (+) template. Linkage is visible as an increase in the proportion of Bristol N2 DNA in Dumpy lanes compared to the wild-type lanes, and is visible on LGI from -12.2 to 15.5. B and C, interval mapping. Each column in B is an individual Dpy recombinant, assayed for the three SNPs W03D8 (top row), D1007 (middle row), and B0205 (bottom row). Most (31/47) recombinants show Bristol DNA at all three SNPs. This indicates that these recombinants were homozygous Bristol at these loci, as expected for tightly linked markers. Sixteen animals show half Bristol and half Hawaiian DNA at one or more loci, indicating that they have one chromosome that is recombinant in this interval. Columns marked "a" are recombinant in the W03D8-D1007 interval, those marked "b" in the D1007-dpy-5 interval, and those marked "c" in the dpy-5-B0205 interval. These data are summarized in C, which depicts the three recombinant genotypes using blue for Bristol DNA and red for Hawaiian DNA. One recombinant, marked with an asterisk, is homozygous Hawaiian at two SNP loci and heterozygous at the third, and is thus very unlikely to be homozygous Bristol at the dpy-5 gene (see Results for explanation).