GLABROUS1 (GL1) belongs to the large family of MYB transcription factors and is known to play a central role in trichome initiation. We studied trichome distribution and the molecular variation of GL1 in 28 A. thaliana accessions. Trichome density on rosette leaves was highly variable among those accessions. On the molecular level, we detected substantial sequence variation in a 3-kb fragment which included the complete coding region of the GL1 locus (pi = 0.01). Phylogenetic analysis of GL1 indicates the presence of two diverged clades among 28 accessions. Using ANOVA, we show that the phenotypic variation in trichome density cannot be explained by the sequence divergence between the two phylogenetic lineages. Sequence analysis of wild-type Arabidopsis thaliana and Arabidopsis lyrata accessions indicates that all amino acid substitutions are located outside of the conserved helix-turn-helix DNA-binding domains R2 and R3. Using plants of A. thaliana and A. lyrata with either naturally occurring or ethyl methane sulfonate--induced glabrous phenotypes, we demonstrate that the last 14 C-terminal amino acids of the GL1 gene have no major impact on the initiation of trichomes.