Microarray analysis of cytoplasmic versus whole cell RNA reveals a considerable number of missed and false positive mRNAs.
With no known exceptions, every published microarray study to determine differential mRNA levels in eukaryotes used RNA extracted from whole cells. It is assumed that the use of whole cell RNA in microarray gene expression analysis provides a legitimate profile of steady-state mRNA. Standard labeling methods and the prevailing dogma that mRNA resides almost exclusively in the cytoplasm has led to the long-standing belief that the nuclear RNA contribution is negligible. We report that unadulterated cytoplasmic RNA uncovers differentially expressed mRNAs that otherwise would not have been detected when using whole cell RNA and that the inclusion of nuclear RNA has a large impact on whole cell gene expression microarray results by distorting the mRNA profile to the extent that a substantial number of false positives are generated. We conclude that to produce a valid profile of the steady-state mRNA population, the nuclear component must be excluded, and to arrive at a more realistic view of a cell's gene expression profile, the nuclear and cytoplasmic RNA fractions should be analyzed separately.
Trask
Heidi W
HW
Norris Cotton Cancer Center, Dartmouth Hitchcock Medical Center, Lebanon, New Hampshire 03756, USA.
Cowper-Sal-lari
Richard
R
Sartor
Maureen A
MA
Gui
Jiang
J
Heath
Catherine V
CV
Renuka
Janhavi
J
Higgins
Azara-Jane
AJ
Andrews
Peter
P
Korc
Murray
M
Moore
Jason H
JH
Tomlinson
Craig R
CR
D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D020869:Gene Expression Profiling; D006801:Humans; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics*
D012333:RNA, Messenger
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