Optimized Protocol for Linear RNA Amplification and Application to Expression Profiling of Human Renal Biopsies

Andreas Scherer, Andreas Krause, J.R. Walker, S.E. Sutton, Daniel Seron, Friedrich Raulf, Michael P. Cooke

Research output: Contribution to journalArticleScientificpeer-review

Abstract

Gene expression analysis using highdensity
cDNA or oligonucleotide arrays is a
rapidly emerging tool for transcriptomics,
the analysis of the transcriptional state of a
cell or organ. One of the limitations of current
methodologies is the requirement of a
relatively large amount of total or
polyadenylated RNA as starting material.
Standard array hybridization protocols require
5–15 μg labeled RNA. To obtain these
quantities from small amounts of starting
RNA material, RNA can be amplified in a
linear fashion. Here we introduce an optimized
protocol for rapid and easy-to-use
amplification of as little as 1 ng total RNA.
Our analysis shows that this method is linear
and highly reproducible and that it preserves
similarities as well as dissimilarities
between normal and disease-related samples.
We applied this technique to the RNA
expression profiling of human renal allograft
biopsies with normal histology and
compared them to the profiles of renal biopsies
with histological evidence of chronic
transplant nephropathy or chronic rejection.
Among others, complement component C1r
was found to be significantly up-regulated in
chronic rejection and chronic transplant
nephropathy biopsies compared to normal
samples, while fructose-1,6-biphosphatase
showed lower-than-normal expression.
Original languageEnglish
JournalBioTechniques
Volume34
Pages (from-to)546-556
ISSN0736-6205
Publication statusPublished - 2003
MoE publication typeA1 Journal article-refereed

Cite this

Scherer, A., Krause, A., Walker, J. R., Sutton, S. E., Seron, D., Raulf, F., & Cooke, M. P. (2003). Optimized Protocol for Linear RNA Amplification and Application to Expression Profiling of Human Renal Biopsies. BioTechniques, 34, 546-556.