|
| Status |
Public on Oct 01, 2007 |
| Title |
larval_gut_gastric_caeca_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
Gastric caeca of 4th. instar An. gambiae larvae
|
| Organism |
Anopheles gambiae |
| Characteristics |
Strain:G3
Larval instar: 4th.
|
| Treatment protocol |
For each biological repeat, 1,000 fourth-instar larvae were anesthetized on ice and dissected in RNAse-free 70% ETOH. Their guts were extracted and divided into gastric caeca (GC), anterior midgut (AM), posterior midgut (PM) and hindgut / Malpighian tubes (HG/MT). Three biological repeats were performed in order to obtain statistical significance.
|
| Growth protocol |
Eggs of the G3 strain of An. gambiae (originally collected in the Gambia in 1975) (Romans et al., 1998) were obtained from the Centers for Disease Control and Prevention (CDC; Atlanta, Georgia). Mosquito larvae were reared in distilled water at 28oC, 12h: 12h light: darkness cycle, and provided with a mixture of baker’s yeast and ground Tetramin® flakes as food.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNAeasy-Mini Kit™ (Qiagen®, Chatsworth, CA, USA) following the manufacturer’s recommendations
|
| Label |
biotin
|
| Label protocol |
cDNA was synthesized from 5ug of total RNA using the One-Cycle Eukaryotic Target Labeling reagents (Affymetrix, Inc.). The cDNA was used as a template for in vitro transcription (IVT) in the presence of T7 RNA Polymerase and a biotinytlated nucleotide/ribonucleotide mix for cRNA amplification and biotin labeling using the IVT labeling kit (Affymetrix, Inc).
|
| |
|
| Hybridization protocol |
The biotin-labeled cRNA was fragmented and hybridized for 16 hours to the Affymetrix GeneChip® Plasmodium/Anopheles Genome Array according to the manufactures’ protocols. The arrays were then washed and stained with streptavidin-phycoerythrin on an Affymetrix Fluidics Station 450
|
| Scan protocol |
Hybridized chips were scanned on a GeneChip Scanner 3000 (Affymetrix, Inc) using default values to generate signal intensities
|
| Description |
Sample represents pooled material from 1,000 dissected individual guts
|
| Data processing |
The GeneSifter™ (VizX Labs, Seattle, WA, USA) software package was used to perform all statistical analyses. Array data was background-adjusted, normalized, and log2-transformed by applying the robust multiarray average (RMA) algorithm (Irizarry et al., 2003).
|
| |
|
| Submission date |
Feb 27, 2007 |
| Last update date |
May 22, 2007 |
| Contact name |
Marco V. Neira |
| E-mail(s) |
mneira@whitney.ufl.edu
|
| Phone |
904-461-4037
|
| Fax |
904-461-4052
|
| Organization name |
University of Florida
|
| Department |
The Whitney Laboratory
|
| Lab |
Mosquito group/Paul Linser's Lab.
|
| Street address |
9505 Oceanshore Blvd.
|
| City |
St. Augustine |
| State/province |
FL |
| ZIP/Postal code |
32080 |
| Country |
USA |
| |
|
| Platform ID |
GPL1321 |
| Series (1) |
| GSE7149 |
A Microarray-based Analysis of Transcriptional Compartmentalization in the Alimentary Canal of Anopheles gambiae |
|
