Differential expression between mature zone of the olfactory organ and the dactyl of the walking legs, a taste organ. Homarus americanus

A cDNA microarray was constructed and used to assess mRNA abundance differences between two of the lobster’s major chemosensory structures, the mature zone of the olfactory organ and the dactyl of the walking legs, a taste organ. The 115 differences detected again emphasized the abundance of neurons in the olfactory organ, especially a cluster of mRNAs encoding cytoskeletal-associated proteins and cell adhesion molecules such as 14-3-3zeta, actins, tubulins, trophonin, Fax, Yel077cp, Suppressor of profilin 2, and gelsolin. Keywords: Differential expression Overall design: We designed a cDNA microarray that consisted of five subarrays to cover all of the cDNA species in our library. We generated two of the sub-arrays, each with 1,152 clones spotted in duplicate, giving 2,304 spots per sub-array. A common set of 96 clones, consisting of positive and negative control sequences, was spotted on each sub-array. Each cloned cDNA insert was amplified by PCR and spotted on nylon membranes using an Affymetrix 417 Arrayer according to protocols supplied by Dr. Kevin Becker (Tanaka et al., 2000; Barrett et al., 2001). Arrays were hybridized against 33P-labeled cDNA produced by oligo-dT priming of reverse transcription using the RadPrime DNA labeling kit (Invitrogen). Hybridization was done in quadruplicate at 50°C in 50 ml tubes in a hybridization oven as described in Barrett et al. (2001).