Abstract The nature and extent of mutational pleiotropy remain largely unknown, despite the central role that pleiotropy plays in many areas of biology, including human disease, agricultural production, and evolution. Here, we investigate the variation in 11,604 gene expression traits among 41 mutation accumulation (MA) lines of Drosophila serrata. We first confirmed that these expression phenotypes were heritable, detecting genetic variation in 96% of them in an outbred, natural population of D. serrata. Among the MA lines, 3385 (29%) of expression traits were variable, with a mean mutational heritability of 0.0005. In most traits, variation was generated by mutations of relatively small phenotypic effect; putative mutations with effects of greater than one phenotypic standard deviation were observed for only 8% of traits. With most (71%) traits unaffected by any mutation, our data provide no support for universal pleiotropy. We further characterized mutational pleiotropy in the 3385 variable traits, using sets of 5, randomly assigned, traits. Covariance among traits chosen at random with respect to their biological function is expected only if pleiotropy is extensive. Taking an analytical approach in which the variance unique to each trait in the random 5-trait sets was partitioned from variance shared among traits, we detected significant (at 5% false discovery rate) mutational covariance in 21% of sets. This frequency of statistically supported covariance implied that at least some mutations must pleiotropically affect a substantial number of traits (>70; 0.6% of all measured traits).