NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM415879 Query DataSets for GSM415879
Status Public on Jun 12, 2009
Title Test array design TA4_2520025_array 1_PMT gain 400
Sample type RNA
 
Source name Pooled RNA isolated from adult, juvenile and cocoon earthworms
Organism Eisenia fetida
Characteristics tissue: whole body or cocoon
age: varied
Treatment protocol Adults bearing clitelum, juveniles and cocoons were pulled out of the culture immediately before being used for RNA extraction.
Growth protocol Earthworms were kept in moistened sphagnum peat (pH 6.5–7.5, moisture content 50%) and fed ad libitum on a diet of with Magic Worm Food (Carolina Biological Supply).
Extracted molecule total RNA
Extraction protocol RNA was prepared using the RNeasy mini kit (QIAGEN, Valencia, CA) following the manufacturer's recommendations. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
Label Cy3
Label protocol Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
 
Hybridization protocol 1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to the custom-designed Agilent test array for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
Scan protocol Slides were scanned immediately after washing on GenePix 4200AL scanner at PMT gain levels 400 and 500 using default setting (Scan resolution 5um, One channel-Green, Laser power 100%).
Description Testing 244K 60-mer or 40-mer probes designed using eArray
Data processing The scanned images were analyzed with GenePix 5 (Molecular Devices, Sunnyvale, CA) using default parameters and Agilient-provided GAL grid file 020025_D_20080421_gal to obtain signal intensities. The background-subtracted median signal intensity (SI) was reported. A threshold SI (mean + 2 × standard deviation of the lowest three spike-in RNA concentrations) was set for each array. Any spot with a SI smaller than the threshold was flagged as absent, whereas others were considered present and thus probes deposited on these spots were regarded as positive probes. Negative control spots were also checked to make sure their SI did not exceed those of the lowest spike-in RNAs.
 
Submission date Jun 11, 2009
Last update date Jun 11, 2009
Contact name Ping Gong
E-mail(s) ping.gong@us.army.mil
Phone (601) 634-3521
Organization name US Army ERDC
Department Environmental Laboratory
Lab Environmental Genomics and Genetics
Street address 3909 Halls Ferry Road
City Vicksburg
State/province MS
ZIP/Postal code 39180
Country USA
 
Platform ID GPL8683
Series (2)
GSE16550 Using pooled earthworm RNA to test 244K probes on TA-4 test array design 2520025 antisense 2
GSE16551 Using pooled earthworm RNA to test 244K probes on TA-1, TA-2, TA-3, TA-4 test arrays

Data table header descriptions
ID_REF
VALUE Background-subtracted median signal intensity

Data table
ID_REF VALUE
242842 12
242518 16
241984 2
242254 16
241744 17
241279 3
240770 3
240624 8
240453 7
239200 6
238856 5
238322 18
238260 22
238618 -5
237784 -10
236175 11
236147 18
234627 14
234931 3
234433 12

Total number of rows: 243504

Table truncated, full table size 2273 Kbytes.




Supplementary file Size Download File type/resource
GSM415879_252002510001_PMT400_gpr.txt.gz 9.1 Mb (ftp)(http) TXT
Processed data included within Sample table

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap