COGENE Linearity Tests
We have conducted a series of control experiments to validate the linearity of
amplification in our micro-cDNA method. These experiments consisted of
constructing a conventional cDNA library from ~5ug of Hela cell mRNA (with no
amplification steps) and a micro cDNA library from ~10ng of Hela cell mRNA
(using our amplification scheme). We then evaluated by colony lifts and
hybridizations what the frequency of clones was in each type of library. In
addition, we seeded three synthetic run-off transcripts at known concentrations
into the RNA that was used to construct the micro-cDNA library (for details on
these controls see http://hg.wustl.edu/lovett/projects/intlctrl.html).
Conventional vs. micromethod comparison: all numbers are the average of
three separate experiments conducted on 1,000,000 colonies from each library;
Ce tag RNAs were only seeded into the micromethod starting RNA and were
present in a ten-fold dilution series.
| Probe |
Conventional |
Micromethod |
|---|
| Actin |
2058 |
1240 |
| GAPDH |
3931 |
4296 |
| EF 1 Gamma |
275 |
169 |
| Ferritin |
2000 |
1576 |
| Leptin Receptor |
86 |
44 |
| Mucin |
1584 |
1905 |
| CE tag 1 |
-- |
2525 |
| CE tag 2 |
-- |
366 |
| CE tag 3 |
-- |
47 |
|
Last updated Feb. 28, 2002