Screening differentially expressed cDNA clones obtained by differential display using amplified RNA.
The major obstacle of differential display is not the technique itself but rather the post-differential display issueof discriminating between false positives and the truly differentially expressed mRNAs. This process is arduous and requires large amounts of RNA. We present and validate a method which allows one to screen putative positives from differential display analysis using only micrograms of total RNA. More importantly, we demonstrate that cDNA probes generated from amplified RNA are representative of the starting mRNA population and can be used for differential screening of mRNA species at a detectable limit of sensitivity of>/=1/40 000.
Poirier
G M
GM
R.W. Johnson Pharmaceutical Research Institute, San Diego, CA 92121, USA.
Pyati
J
J
Wan
J S
JS
Erlander
M G
MG
D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000096:biosynthesis / Q000302:isolation & purification*; D005189:False Positive Reactions; D005784:Gene Amplification; D006367:HeLa Cells; D006801:Humans; D012313:RNA / Q000235:genetics*
D018076:DNA, Complementary; D012313:RNA