The genetic structure of 49 natural populations of four species (P. attwateri, P. boylii, P. pectoralis, and P. polius) of the Peromyscus boylii species group was analyzed through application of chromosomal and electrophoretic techniques. Chromosomal variation within and among populations of the boylii species group was analyzed from 178 specimens. Electrophoretic techniques were utilized for the demonstration of variation in enzymes and other proteins encoded by structural loci and applied to the study of the evolution of the boylii species group by estimation of levels of genetic heterozygosity within populations, estimation of degree of genetic similarity between conspecific populations and between species, and determination of patterns of geographic variation in allelic frequencies and levels of heterozygosity. Six distinct chromosomal patterns were observed among the populations of the four species of the boylii species group. All specimens had a diploid number of 48 and the major difference in chromosomal morphology was in the number of pairs of large to medium biarmed autosomes. Little or no chromosomal variation was observed in three species (attwateri, pectoralis and polius), but considerable chromosomal variation occurred among populations of P. boylii. Generally, the chromosomal variation in P. bylii was between allopatric populations, with each chromosomal pattern limited to a recognized subspecies. Polymorphism was observed in two populations. The polymorphism observed in P. polius was the result of pericentric inversion involving the smallest pair of metacentric autosomes. The polymorphism observed in P. bolii cileus was interpreted at the result of gene flow between P. boylii rowleyi and P. boylii spicilegus. In addition to chromosomal evidence, analysis of electrophoretic data demonstrated and suggested effective gene flow between the chromosomal forms of P. boylii. Electrophoretically demonstrable variation was analyzed in 11 proteins encoded by 17 autosomal loci. Of the 17 structural loci, 11 were polymorphic in one or more populations . No more than five loci were observed to be polymorphic within a single population, with a mean number of polymorphic loci of 2.26. The levels of genetic variability, as measured by the proportion of loci in a heterozygous state in the average individual of a population, for populations of the boylii species group were within the range reported for other rodents. Populations of P. attwateri exhibited levels of heterozygosity considerably lower than those reported for other species of Peromyscus. The low level of genetic variability in P. attwateri was probably the result of founder effect of the original population which was isolated on the Edwards Plateau during the late Pleistocene and subsequent genetic drift which allowed fixation of genes within this isolated population. Geographic variation in levels of heterozygosity observed among populations of P. attwateri and P. boylii indicates a north to south cline of increasing heterozygosity with the most variable populations occurring in the southern portion of the range of each species. A wide range of the levels of heterozygosity and genetic similarity was observed among populations of P. pectoralis and is the result of genetic contribution from three Pleistocene refugia. Analysis of paired combinations of populations of P. boyli indicated that the various chromosomal forms of P. boylii. form a conspecific unit. Populations of P. attwateri were genetically distinct from other members of this group. Examination of specific loci for allelic homology suggested that P. attwateri arose from genetic divergence of a population of P. boylii isolated on the Edwards Plateau during the late Pleistocene.