thus, for example, DNA samples separated in a gel can be cut out of the gel, returned
to solution and recovered.
Owing to the poor elasticity of agarose gels and the consequent problems of removing
them from small tubes, the gel rod system sometimes used for acrylamide gels is not
used. Horizontal slab gels are invariably used for isoelectric focussing or immunoelectro-
phoresis in agarose. Horizontal gels are also used routinely for DNA and RNA gels
(Section 10.4), although vertical systems have been used by some workers.10.2.2 Polyacrylamide gels
Electrophoresis in acrylamide gels is frequently referred to as PAGE, being an abbre-
viation forpolyacrylamidegelelectrophoresis.
Cross-linked polyacrylamide gels are formed from the polymerisation of acrylamide
monomer in the presence of smaller amounts ofN,N^0 -methylene-bisacrylamide (nor-
mally referred to as ‘bis’-acrylamide) (Fig. 10.5). Note that bisacrylamide is essentially
two acrylamide molecules linked by a methylene group, and is used as a cross-linking
agent. Acrylamide monomer is polymerised in a head-to-tail fashion into long chains
and occasionally a bis-acrylamide molecule is built into the growing chain, thus
introducing a second site for chain extension. Proceeding in this way a cross-linked
matrix of fairly well-defined structure is formed (Fig. 10.5). The polymerisation of
acrylamide is an example of free-radical catalysis, and is initiated by the addition of
ammonium persulphate and the base N,N,N^0 ,N^0 -tetramethylenediamine (TEMED).
TEMED catalyses the decomposition of the persulphate ion to give a free radical
(i.e. a molecule with an unpaired electron):S 2 O^28 þe!SO^24 þSO 4 CH 2 = CHCONH 2
AcrylamideCH 2 (NHCOHC = CH 2 ) 2
N,N,N′,N′-methylenebisacrylamideCH 2 CH
CO
NHCH 2
NH
COCHCH
CO
NHCH 2
NH
COCHCH 2 CH CH 2
CO
NH 2NH 2
COCH 2 CH CH 2
nnCH 2+
Free radical
catalystFig. 10.5The formation of a polyacrylamide gel from acrylamide and bis-acrylamide.405 10.2 Support media