ABSTRACT
We describe an original apparatus for the study of the dynamics of single stranded DNA migration. Four detectors based on laser-induced fluorescence (LIF) are equidistantly placed on one migration lane, allowing repeated measurements of the same DNA band at different positions along migration. This article presents the characteristics and performances of this system and focuses on the data analysis, showing how the multiple detection scheme enables the study of band broadening and band resolution during a migration run. Our results suggest the existence of anomalous (nonthermal) diffusion of DNA molecules during the electrophoretic process.
Subject(s)
Bacteriophage M13/genetics , DNA, Single-Stranded/chemistry , DNA, Viral/chemistry , Electrophoresis, Polyacrylamide Gel/instrumentation , Electrophoresis, Polyacrylamide Gel/methods , Nucleic Acid Denaturation , Time FactorsABSTRACT
An original apparatus based on laser-induced fluorescence detection is presented. One lane migration combined to four equidistant detection points allows the study of the dynamics of DNA bands during electrophoresis. We focus this article on the study of the mobility of DNA sequencing fragments as a function of temperature; mobility is determined in 4% T, 5% C and 4.3% T, 5% C cross-linked polyacrylamide gels at an electric field of 45 V/cm [T=(g acrylamide+g N,N'-methylenebisacrylamide)/100 ml solution; C=g N,N'-methylenebisacrylamide/% T]. Activation energy has been investigated under these experimental conditions with a temperature varying from 25 to 50 degrees C. The activation energy for migration through the cross-linked polyacrylamide gel decreases with fragment length under our experimental conditions and it varies along the migration.
