Drosophila melanogaster
ovaries transformed into  testes, separated from fatbody
strain
product of a cross
age
late third instar
dev_stage
Late third instar larval stage: immobilized, foregut midgut clear, anterior spiracle non-everted
sex
male
tissue
gonads
biomaterial_provider
Brian Oliver Lab
genotype
w[1118]/ w[1118]; dsx[1]/dsx[D]
growth_protocol
We grew flies on Fly food A (LabExpress Ann Arbor, MI) at 25 C in constant light and 60 % relative humidity.
phenotype
sex transformed
sample_type
RNA
store_cond
-80 oC
treatment
mutant
sample_role
treatment mutant
temperature_regimen
25 oC
target_gene_induced
dsx FBgn0000504
target_gene_inhibited/mutated
dsx FBgn0000504
details
We grew flies on Fly food A (LabExpress Ann Arbor, MI) at 25 C in constant light and 60 % relative humidity. We dissected the third instar larvae (L3) just after they were immobilized and prior to anterior spiracle extension; just prior to pupal development. Additionally, staging was aided by staining gut contents using 0.5 mg/ml Sulforhodamine B sodium salt (Sigma-Aldrich, Cat. No. S1402-25G. St. Louis, MO) yeast paste fed to third instar larvae. We used larvae with clear foreguts, and full midguts/hindguts. We dissected larvae in PBS pH 7.4 (Gibco Life Technologies, Carlsbad, CA) at 22 C. We collected larval tranformed testes without fatbody by enzymatically removing fatbody by treating with 0.075 mg/ml Papain (Sigma-Aldrich, St. Louis, MO) and 0.075 mg/ml Collagenase enzymes (Sigma-Aldrich, St. Louis, MO) in PBS in 1.5ml eppendorf tube for 2-3 min at 22o C by gently flicking the tube with a forefinger. We decanted floating fatbody cells and washed 2X with PBS. Sample (N = 200) was collected in 350 l RLT buffer (Qiagen, Cat. No. 79216. Valencia, CA), vortexed quickly and frozen in a dry ice ethanol bath and stored at -80 C until total RNA extraction using the RNeasy Mini kit (Qiagen, Cat. No. 74104. Valencia, CA) following the manufacturers protocol. RNA was quantified using Quant-iT RiboGreen quantification kit (Life Technologies, Cat. No. R11490. Carlsbad, CA). RNA was stored in -80 C.
BioSampleModel
Model organism or animal