We cloned an Achaearanea tepidariorum (Chelicerata, Arachnida) gene related to Drosophila twist (twi), which encodes a basic helix-loop-helix transcription factor required to specify mesoderm fate in the Drosophila embryo. The cloned spider gene was designated At.twist (At.twi). We examined its expression by whole-mount in situ hybridization. At.twi transcripts were first detected in cells located at the polar and equatorial areas of the spherical embryo when the cumulus reached the equator. As the extra-embryonic area expanded, more cells expressed At.twi transcripts. The At.twi-expressing cells became distributed nearly uniformly in the embryonic area. At these stages, some At.twi-expressing cells were found in the surface epithelial cell layer, but other At.twi-expressing cells were at slightly deeper positions from the surface. When the embryo was transformed into a germ band, all At.twi-expressing cells were situated just beneath the surface ectoderm, where they became metamerically arranged. Although little expression was observed in the caudal lobe of the elongating germ band, new stripes of At.twi expression appeared beneath the ectoderm in accordance with the posterior growth. These observations suggested that the cells expressing At.twi were most likely mesoderm. We propose that At.twi can be used as a molecular marker for analyzing mesoderm development in the spider embryo. Moreover, comparison of the expression patterns of twi and At.twi revealed divergent aspects of mesoderm development in the fly and spider. In addition, we cloned an Achaearanea gene related to snail, which is another mesoderm-determining gene in Drosophila, and showed that its expression was restricted to the ectoderm with no indication for a role in mesoderm development.