title: Lobster2
status: Public on Dec 08 2005
contact.city: Lexington
contact.first: Timothy
contact.last: McClintock
contact.email: mcclint@uky.edu
contact.state: KY
contact.address: 800 Rose Street
contact.department: Department of Physiology
contact.country: USA
contact.institute: University of Kentucky
organism: Homarus americanus
description: Clones spotted in duplicate. Duplicates have the same numeric part of ID in the table (for example 2225 and 2225b).
manufacture_protocol: cDNA microarray that consisted of five subarrays was  designed. We generated two of the sub-arrays, each with 1,152 clones spotted in duplicate. 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). Washing wash done twice in 2x SSC and 0.1% SDS for 10 min and twice in 1x SSC and 0.1%SDS for 15 min at room temperature. Signals from hybridized arrays were quantified using a STORM phosphorimaging device running ImageQuant software (Molecular Dynamics, Sunnyvale, CA). ImageQuant,was used to segment each spot and calculate the volume signal intensity for each.