AJP - Lung Cellular and Molecular Physiology, Vol 269, Issue 3 273-L284, Copyright © 1995 by American Physiological Society
Differential display RT-PCR for identifying novel gene expression in the lung
M. E. Sunday
Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.
The unique identity of each cell is the result of differential gene
expression. A new strategy for differential cDNA screening introduced by
Liang and Pardee utilizes anchored oligo-dT primers and random 5'
oligonucleotide 10-mers to carry out polymerase chain reaction (PCR) on
reverse-transcribed RNA from different cell populations. The amplified
cDNAs are displayed on a standard sequencing gel as 100-500 bands per lane
on the resulting autoradiograms and comparisons are drawn between the
different cell populations. The major advantages of this method over
previous differential screening approaches are its high sensitivity, the
small amounts of tissue required, and the ability to carry out rapid mRNA
analyses using total RNA and to test multiple tissues in parallel. Its
limitations are the need for many primer combinations for adequate
representation of mRNAs and the large number of bands displayed. A
screening strategy should include multiple positive and negative control
samples and Northern blots to identify cDNAs differentially expressed. This
approach should facilitate the identification of many novel genes expressed
in a variety of physiological and pathological conditions.
