ELECTROPHORESIS OF DNA IN NOVEL THERMOREVERSIBLE MATRICES

We demonstrate the feasibility of temperature-sensitive polymers as no vel matrices for capillary and slab electrophoresis of DNA. These matr ices combine the ease-of-use of agarose with resolution properties of polyacrylamide. Two classes of matrices ate used: (i) aqueous suspensi ons of gel microspheres and (ii) solutions containing uncross-linked t emperature-sensitive polymers. When heated, the viscosity of these sol utions drops dramatically because of the phase transition behavior of these polymers, and therefore these formulations are easy to pour or l oad. Results are presented for separation of double-stranded DNA fragm ents (< 2000 bp) in capillary, tube, and slab electrophoresis. Prelimi nary results are also presented for separation of single-stranded sequ encing fragments by capillary electrophoresis. In the tube format, goo d resolution was obtained for Phi X174/HaeIII fragments. In the slab f ormat, excellent resolution of double-stranded DNA was obtained, inclu ding simultaneous separation of a 10 bp ladder up to 150 base pairs. I n the capillary format, pBR322/MspI fragments were completely resolved , and single-base resolution of sequencing fragments was obtained up t o 150 bases. We believe that temperature-sensitive polymers represent a new and promising class of electrophoretic media.