TY - JOUR
T1 - Accurate and reproducible gene expression profiles from laser capture microdissection, transcript amplification, and high density oligonucleotide microarray analysis
AU - Luzzi, Veronica
AU - Mahadevappa, Mamatha
AU - Raja, Rajiv
AU - Warrington, Janet A.
AU - Watson, Mark A.
N1 - Funding Information:
Supported in part by grant number 016-99 from the Mary Kay Ash Charitable Foundation (M.A.W.) and the Alvin J. Sikman Cancer Center.
PY - 2003/2
Y1 - 2003/2
N2 - Gene expression profiling using high density oligonucleotide arrays is a powerful method to generate an unbiased survey of a cell's transcriptional landscape. Increasingly complex biological questions require that this approach be applicable to the small numbers of cells that are obtained from sources such as laser capture microdissection (LCM) of solid tissues. In this report, we demonstrate that two rounds of transcript amplification can generate accurate and reproducible gene expression profiles using high density oligonucleotide microarrays, starting with as little as 10 ng of total RNA. Biased amplification of the 3′ end of transcripts does not have a major impact on the overall transcript profile due to the 3′ bias of probe sets incorporated in the array design. Furthermore, greater than 95% of all genes detected demonstrate less than a twofold difference in expression when independent tissue dissections of identical cell populations are compared. The accuracy and technical reproducibility of the method suggests that expression profiling using transcript amplification and high density oligonucleotide microarrays can be used on a routine basis.
AB - Gene expression profiling using high density oligonucleotide arrays is a powerful method to generate an unbiased survey of a cell's transcriptional landscape. Increasingly complex biological questions require that this approach be applicable to the small numbers of cells that are obtained from sources such as laser capture microdissection (LCM) of solid tissues. In this report, we demonstrate that two rounds of transcript amplification can generate accurate and reproducible gene expression profiles using high density oligonucleotide microarrays, starting with as little as 10 ng of total RNA. Biased amplification of the 3′ end of transcripts does not have a major impact on the overall transcript profile due to the 3′ bias of probe sets incorporated in the array design. Furthermore, greater than 95% of all genes detected demonstrate less than a twofold difference in expression when independent tissue dissections of identical cell populations are compared. The accuracy and technical reproducibility of the method suggests that expression profiling using transcript amplification and high density oligonucleotide microarrays can be used on a routine basis.
UR - http://www.scopus.com/inward/record.url?scp=0037320528&partnerID=8YFLogxK
U2 - 10.1016/S1525-1578(10)60445-X
DO - 10.1016/S1525-1578(10)60445-X
M3 - Article
C2 - 12552074
AN - SCOPUS:0037320528
SN - 1525-1578
VL - 5
SP - 9
EP - 14
JO - Journal of Molecular Diagnostics
JF - Journal of Molecular Diagnostics
IS - 1
ER -