Dominik Haudenschild 7-28-93 at CBRC

Casting gel: use FMC Catalog # 50920 ProSieve Gel System

-Melt gel in microwave at 50% power

-Cool to 50-55'C in water bath

-Pour resolving gel solution into horizontal electrophoresis

chamber until it is about 2-3mm thick

-Allow to cool at room temperature for 30 minutes

-Cut 15mm strip of resolving gel to make room for stacking gel

-Melt and cool stacking gel as before

-Pour stacking gel, then quickly insert combs about 10mm from

interface between stacking and resolving gels

-Allow to cool 15 minutes at room temperature, then put in 4'C

for 30 minutes

-Flood electrophoresis unit with regular SDS-PAGE running

buffer

-Very carefully remove combs, do not allow stacking and

resolving gels to separate

-Put protein in normal SDS-PAGE sample buffer, boil for 5 min.

-Load samples and run at 100 to 150 V, be very careful not to

melt agarose gel. Keep on Ice or do in cold room