Isolation of rare transcripts by representational difference analysis.
Representational difference analysis (RDA) is a powerful technique for cloning the differences between genomes, and has recently been adapted for cloning differentially expressed genes. RDA, like other PCR-based differential screening methods, is prone to the production of false positives. We have identified a major source of false positives in RDA of cDNA and have introduced improvements which minimise their production. These modifications also significantly increase sensitivity, allowing for the isolation of rare differential transcripts from nanogram amounts of mRNA.
O'Neill
M J
MJ
Department of Paediatrics and Centre for Hormone Research, The University of Melbourne, Royal Children's Hospital, Melbourne, Victoria 3052, Australia. oneill@cryptic.rch.unimelb.edu.au
Sinclair
A H
AH
D000818:Animals; D001483:Base Sequence; D001923:Brain Chemistry; D002642:Chick Embryo; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry; D004351:Drug Resistance / Q000235:genetics; D005189:False Positive Reactions; D005821:Genetic Techniques; D008969:Molecular Sequence Data; D009355:Neomycin; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
D018076:DNA, Complementary; D012333:RNA, Messenger; D009355:Neomycin