|
| Status |
Public on Sep 01, 2012 |
| Title |
larval Kir-LNvs at CT-3, biological rep3 |
| Sample type |
RNA |
| |
|
| Source name |
Drosophila larval Kir-LNvs at CT-3
|
| Organism |
Drosophila melanogaster |
| Characteristics |
tissue: FACs-purified LNvs
genotype: yw; Pdf-Gal4/UAS-Kir; Pdf-RFP
developmental stage: L3 larval stage
|
| Treatment protocol |
Dissected brains (L3 larvae) were transferred to Schneider’s Insect Medium (Sigma) in non-stick tubes (Neptune) and were kept on ice throughout. Prior to dissociation, brains were washed with ice-cold PBS buffer. For dissociation, we followed the method of Wegener et al: brains transferred to a 50:50 mix of 1X Collagenase (Sigma): 1X Dispase II (Roche) and incubated for 2hr at 25°C. After 2hr, the dissociation solution was replaced with Schneider’s medium with 10% FBS (Fetal Bovine Serum), brains were triturated 100x with a pipette, and strained through a 35mm nylon mesh filter. Cells in Schneider’s medium / 10% FBS were kept on ice for transport FACS.
|
| Growth protocol |
All larval genotypes raised at 25C in 12:12hr Light/Dark cycle.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The 0.01%-most RFP+ cells (150 - 300 cells) were isolated by FACS, and sorted directly into Arcturus PicoPure RNA extraction buffer. mRNA was amplified using the NuGen WT-Ovation™ Pico System to yield ~ 5ug biotin labeled, fragmented, single-stranded cDNA
|
| Label |
biotin
|
| Label protocol |
Affymetrix protocol
|
| |
|
| Hybridization protocol |
Following fragmentation, 5 ug of cDNA were hybridized for 20 hr at 45C on GeneChip Drosophila Genome Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.
|
| Scan protocol |
GeneChips were scanned using the Hewlett-Packard GeneArray Scanner G2500A.
|
| Description |
Gene expression data from Kir-expressing LNvs at CT-3
|
| Data processing |
Raw hybridization intensities from each CEL Affymetrix file (Drosophila 2.0 GeneChip) were analyzed using gcrma algorithm in Matlab. Intensities were quantile-normalized across all replicates and experiments to yield a single expression measure for each probeset on the chip.
|
| |
|
| Submission date |
Feb 28, 2011 |
| Last update date |
Aug 28, 2018 |
| Contact name |
Marc Ruben |
| Organization name |
New York University
|
| Department |
Biology
|
| Lab |
Justin Blau
|
| Street address |
100 Washington Square East
|
| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10003 |
| Country |
USA |
| |
|
| Platform ID |
GPL1322 |
| Series (1) |
| GSE27565 |
Expression data from electrically-altered larval Drosophila LNvs |
|
| Relations |
| Reanalyzed by |
GSE119084 |
