Computer-aided analysis of DNA curves on transverse gradient gels.
Transverse pore gradient polyacrylamide gel electrophoresis of DNA restriction fragments was used to generate gel patterns describing migration distance as a function of gel concentration (Ferguson curves). These Ferguson curves were digitized, traced and analyzed with the aid of a personal computer. The traced curves were plotted semi-logarithmically and the plots were subjected to least-squares linear regression analysis to yield values of the slope (KR) and the intercept at %T = 0 (YO). These values are highly precise since they are based on approx. 100 measurements per curve. The computerized method reduces the errors due to manual measurements of migration distances and is time and labor saving. The method is still limited to intra-experimental comparison of Ferguson curves, since it does not as yet comprise a determination of gel concentration. At present, curve tracing remains semi-automated, requiring manual intervention when Ferguson curves cross or approach one another. Potentially, the importance of the computerized analysis of transverse pore gradient gels lies in the rapid quantitative interpretation of Ferguson curves for detection of anomalously migrating DNA species. Potentially, that application provides a more sensitive and informative mode of detection than either the mere visual observation of crossing Ferguson curves or of a shift in mobility at a single gel concentration.
Wheeler
D
D
Section on Macromolecular Analysis, National Institute of Child Health and Human Development, NIH, Bethesda, MD 20892.
Orban
L
L
Garner
M M
MM
Chrambach
A
A
D002021:Buffers; D016001:Confidence Intervals; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D007091:Image Processing, Computer-Assisted; D015203:Reproducibility of Results; D013194:Staining and Labeling
D002021:Buffers; D004247:DNA