Improved microarray gene expression profiling of virus-infected cells after removal of viral RNA.
BACKGROUND
Sensitivity and accuracy are key points when using microarrays to detect alterations in gene expression under different conditions. Critical to the acquisition of reliable results is the preparation of the RNA. In the field of virology, when analyzing the host cell's reaction to infection, the often high representation of viral RNA (vRNA) within total RNA preparations from infected cells is likely to interfere with microarray analysis. Yet, this effect has not been investigated despite the many reports that describe gene expression profiling of virus-infected cells using microarrays.


RESULTS
In this study we used coronaviruses as a model to show that vRNA indeed interferes with microarray analysis, decreasing both sensitivity and accuracy. We also demonstrate that the removal of vRNA from total RNA samples, by means of virus-specific oligonucleotide capturing, significantly reduced the number of false-positive hits and increased the sensitivity of the method as tested on different array platforms.


CONCLUSIONS
We therefore recommend the specific removal of vRNA, or of any other abundant 'contaminating' RNAs, from total RNA samples to improve the quality and reliability of microarray analyses.
Raaben
Matthijs
M
Virology Division, Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL Utrecht, The Netherlands. m.raaben@uu.nl
Whitley
Penn
P
Bouwmeester
Diane
D
Setterquist
Robert A
RA
Rottier
Peter J M
PJ
de Haan
Cornelis A M
CA
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D018352:Coronavirus Infections / Q000235:genetics* / Q000821:virology*; D020869:Gene Expression Profiling / Q000379:methods* / Q000706:statistics & numerical data; D006524:Hepatitis, Viral, Animal / Q000235:genetics* / Q000821:virology*; D051379:Mice; D006517:Murine hepatitis virus / Q000235:genetics* / Q000472:pathogenicity; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods* / Q000706:statistics & numerical data; D012367:RNA, Viral / Q000235:genetics* / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
D012367:RNA, Viral