Quantitative trait locus (QTL) studies, in which random marker loci are used to estimate the location and average effects of loci influencing complex phenotypes, are becoming more feasible as techniques for identifying variation in marker loci are improving. Scoring simple sequence repeat polymorphisms (SSRPs) with the polymerase chain reaction has become the method of choice for identifying marker loci in QTL investigations, because of the hypervariability of these loci and the high repeatability of the technique (Dietrich et al., 1992; Routman and Cheverud, in press). Variation in SSR loci (also known as microsatellites) is usually found by running radioactive PCR products on denaturing polyacrylamide gels and scoring length variants by autoradiography. This methodology is very sensitive for detecting small size differences among alleles.
For logistic reasons, radioactive labeling of PCR products was not possible for our QTL study of mouse growth. In this paper we describe some techniques for scoring SSRPs by agarose electrophoresis and ethidium bromide staining. These techniques have the advantages of (1) allowing the scoring of large numbers of individuals quickly, (2) eliminating the use of radioactivity, and (3) being extremely inexpensive. We also describe some solutions to problems that arose during the screening of 534 individuals for a large number of SSR loci for a QTL study of growth in mice.
Total cellular DNA was extracted from spleen with the following protocol. A small piece of spleen (<27 mm 3) was placed into a microcentrifuge tube with 650 gl of 4.7 M guanidinium isothiocyanate and macerated with a ground-glass pestle. This suspension was placed at 60~ for anywhere from 1 h to overnight, resulting in complete digestion of the tissue. Proteins were precipitated by adding 65-70 gl of 5 M NaC1, vortexing, and incubating on ice for