genotype/variation: Logjam tissue: head to thorax other: loj 00898/loj 04026 mutants and control samples containing equal numbers of loj 00898/+ and loj 04026/+ gender: Female age: 4 days
Treatment protocol
Females of the genotypes loj 00898/loj 04026, loj 00898/+, and loj 04026/+ were collected as virgins within 1-2 h of the beginning of the 12h lighted portion of the cycle, kept in glass vials in groups of 20 or fewer flies, and aspirated singly into new vials on day 3 after collection. On day 4, a single wild-type male was aspirated into a vial containing a single female and observed for mating.
Growth protocol
Drosophila melanogaster stocks and cross progeny were maintained at 25ºC on a 12h light-dark cycle using a standard cornmeal, sugar, agar and yeast culture medium.
Extracted molecule
total RNA
Extraction protocol
Standard Trizol (Invitrogen) extraction. Only females that mated within 30 min for 18-30 min were collected for subsequent RNA extractions. Female abdomens were dissected away from the head/thorax 3 hours after the end of the mating period and each of the two tissue types (abdomen or head with thorax) was quick-frozen in Trizol reagent (Invitrogen). Total RNA from abdomen or head/thorax tissue was extracted in Trizol using the manufacturer's protocol. Twelve female tissue samples were collected for each of 6 independent RNA extractions for loj 00898/loj 04026. Our control samples consisted of 6 loj 00898/+ and 6 loj 04026/+ samples combined together for each of 6 independent RNA extractions.
Label
biotin
Label protocol
Standard Affymetrix
Hybridization protocol
Standard Affymetrix
Scan protocol
Standard Affymetrix
Description
Abdomen or Head/thorax tissue samples from females of the following genotypes: loj 00898/loj 04026, loj 00898/+, and loj 04026/+
Data processing
Data were analyzed 3 different ways:
VALUE [dCHIP_PMMM]: All 6 chips were analyzed simultaneously in dCHIP, which uses a model-based algorithm to estimate expression values. The normalization procedure implemented in dCHIP normalizes all arrays in the analysis against the array with median intensity. This array was analyzed using the PM-MM procedure under default parameters.
VALUE2 [dCHIP_PMonly]: All 6 chips were analyzed simultaneously in dCHIP, which uses a model-based algorithm to estimate expression values. The normalization procedure implemented in dCHIP normalizes all arrays in the analysis against the array with median intensity. This array was analyzed using the PM_only procedure under default parameters.
VALUE3 [GCOS]: Expression values were computed using the recommended settings in GCOS version 1.2. All probesets were scaled using a TGT value of 500, and no normalization was applied (i.e., the normalization factor was 1.0).
