Recovery of functional DNA inserts by electroendosmotic elution during gel electrophoresis.
In contrast to all previous preparative electrophoresis apparatus which used a pump, electroendosmotic elution uses bound electrical charges at the end of the separating gel to generate a buffer flow. The electroendosmotic flow increased with increasing currents and decreasing buffer concentrations: its exact characteristics for the built apparatus were determined. The electroendosmotic device was able to separate two DNA fragments differing in size by only 5% with a recovery over 95%. As demonstrated in practical examples of recovery and uses of DNA inserts, up to 10 micrograms of DNA per band can be loaded at a time. The recovered DNA can be used directly for nick-translation, ligation... without further treatment. The performances of the method are expected to improve still further if the charge density and pores of the electroendosmotic medium can be "made-to-order" to provide a better flow profile of the eluting buffer.
Tan
H V
HV
INSERM U15, Paris, France.
Kitzis
A
A
Berthollet
T
T
Hamard
G
G
Beldjord
C
C
Benarous
R
R
D002021:Buffers; D004247:DNA / Q000302:isolation & purification*; D004262:DNA Restriction Enzymes; D004586:Electrophoresis / Q000295:instrumentation*; D004587:Electrophoresis, Agar Gel / Q000295:instrumentation*; D009995:Osmosis
D002021:Buffers; D004247:DNA; D004262:DNA Restriction Enzymes