1. Using ArrayStar with a public dataset
Jean-Yves Sgro

2. ArrayStar Part of Lasergene Genomics Suite for analysis:
Global gene expression
Variation across groups
Optional Qseq module to analyze RNA-Seq, ChIP-Seq, CNV, and miRNA data.

3. Public repositories
2 cross referenced world databases for microarray and Next Gen data:
USA: Gene Omnibus (GEO) (
Europe (UK): ArrayExpress (EMBL/EBI) (

4. Search example: rhinovirus

6. Today: GEO series GSE27973
Or short URL version: (see page 14)
1.usa.gov/WiUkmP

7. Today: GEO series GSE27973

8. 4 groups
Our analysis today with 2 groups: find the difference between medium alone and RV16 alone

10. Sample files
2 types:
CEL
CHP
We want the CEL (raw data) files

11. Essential: rename files
RENAME GSM CEL medium_Donor_1.CEL
RENAME GSM CEL RV16_Donor_1.CEL
RENAME GSM CEL CSE_Donor_1.CEL
RENAME GSM CEL RV16+CSE_Donor_1.CEL
RENAME GSM CEL medium_Donor_2.CEL
RENAME GSM CEL RV16_Donor_2.CEL
RENAME GSM CEL CSE_Donor_2.CEL
RENAME GSM CEL RV16+CSE_Donor_2.CEL
RENAME GSM CEL medium_Donor_3.CEL
RENAME GSM CEL RV16_Donor_3.CEL
RENAME GSM CEL CSE_Donor_3.CEL
RENAME GSM CEL RV16+CSE_Donor_3.CEL
RENAME GSM CEL medium_Donor_4.CEL
RENAME GSM CEL RV16_Donor_4.CEL
RENAME GSM CEL CSE_Donor_4.CEL
RENAME GSM CEL RV16+CSE_Donor_4.CEL
Files are ALREADY RENAMED and on the Windows Desktop

12. Replicate names: automatic

13. Grouped automatically

14. Register with Affymetrix
Affymetrix.com

15. From home / outside Biochem department
Step 1:
VPN connection
Step 2:
Lasergene software: ArrayStar
See also page 8
dept-ra-cssc.vpn.wisc.edu

16. High Density Oligonucleotide Affymetrix Arrays

17. http://www. affymetrix
The next six slides will provide a simple graphical explanation of the interaction of molecules in the GeneChip® microarray at a single feature.
They will help explain how this interaction leads to a fluorescing feature that illustrates a gene is expressed in a cell.
Most of the molecules in this example are simplified, such as the probes being shown as 7 bases long instead if 25 base pairs.
This slide shows the dimensions of a typical chip and the connection from chip to feature to probe

18. When the tagged (red ball) RNA (in purple) is added to the array, the RNAs “look” for their complimentary match

19. Match | no match This illustrates what happens when there is no match
The RNA essentially bounces off the probes
Even one base pair is enough to prevent the match

20. Match | no match
The visualization occurs because the fluorescent molecules are washed over the entire array and stick to a RNA fragment tagged with the Biotin
When hit with a laser in the scanner, any place the fluorescent molecule sticks, it will glow and be picked up by the computer

21. Credit: slide IBC course 2004

22. Credit: slide IBC course 2004

23. CEL and CDF files
Image analysis of each scanned Affy array produces a CEL file containing the mean intensity (and other statistics) for each square probe cell
To do anything with this file you also need the CDF (Chip Description File) which specifies the probe and the probe set that each cell belongs to
CDF information is provided by Affymetrix (download)
Credit: slide IBC course 2004

24. RMA Histogram of log(PM) with fitted model
Robust Multichip Average (RMA)
PM data on log2 scale:
raw and fitted model
Histogram of log(PM)
with fitted model
Credit: slide IBC course 2004

25. RMA normalization
Force the empirical distribution of probe intensities to be the same for every chip in an experiment
The common distribution is obtained by averaging each quantile across chips
Credit: slide IBC course 2004