INFLUENCE OF THE AGAROSE MATRIX IN PULSED-FIELD ELECTROPHORESIS

Properties of agarose potentially relevant to PFGE (pulsed-field gel e lectrophoresis) are reviewed, and some new information is presented. A garose polymers appear to have molecular weights in the range of 100 0 00 to 200 000 Da,but this is not tightly related to the effective gel strenght. Agarose has some residual charge, and hence exhibits electro endosmosis,(EEO). It is possible to markedly increase the speed of sep aration of DNA molecules by using agarose of low EEO, especially in lo w ionic strenght, non-borate buffers. This increase is especially noti ceable in the relatively long experiments required for separation of l arge DNAs. It is also possible to increase the range of separation in a single run by use of step gradients of agarose concentration, which allows visualization of yeast chromosomes and lambda-phage restriction fragments in the same lane. Because of the strong influence of concen tration on separation, it may be useful for investigators to control w ater content and related variables. Our lack of knowledge of the detai led microstructure of gels may be barrier to complete understanding of PFGE.