Transcriptome amplification methods in gene expression profiling.
The increasing use of microarray expression profiling to study the molecular biology of cancer and the cellular physiology of difficult-to-isolate cell types has led to a need for methods that accurately and precisely amplify small quantities of RNA. The purpose of this review is to provide an overview of the existing methods for transcriptome amplification and to define the parameters for comparing different amplification methods. The authors propose a standardized protocol for the assessment and evaluation of amplification methods, focusing on a new whole-transcriptome amplification kit, which amplifies total RNA into cDNA fragments. Reproducibility and reliability of the method were analyzed and discussed using both quantitative real-time PCR and a high-density oligonucleotide microarray platform.
Peano
Clelia
C
Institute for Biomedical Technologies, National Research Council, Milan, Italy. clelia.peano@itb.cnr.it
Severgnini
Marco
M
Cifola
Ingrid
I
De Bellis
Gianluca
G
Battaglia
Cristina
C
D018076:DNA, Complementary / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D020022:Genetic Predisposition to Disease; D006801:Humans; D008954:Models, Biological; D025202:Molecular Diagnostic Techniques; D021141:Nucleic Acid Amplification Techniques; D020411:Oligonucleotide Array Sequence Analysis; D012313:RNA / Q000378:metabolism; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
D018076:DNA, Complementary; D012313:RNA