Hoefer HE33 - Manual (Page 19)

• p11
Notes, buffers, and volumes
Agarose gel electrophoresis notes
Agarose gel electrophoresis can be used to sepa-
rate DNA fragments as small as 0.1 kb or less.
Polyacrylamide gels are usually used for frag-
ments smaller than 1 kb.
DNA mobility
Suggested agarose concentration for separating
fragments of various sizes is given in Table 2
below. Other factors affecting separation results
include the running buffer, voltage setting,
temperature, conformation, and the presence of
ethidium bromide. Special agaroses are available
that can extend resolution ranges.
A common standard is a Hind III digest of
lambda phage, which gives eight fragments rang-
ing in size from 0.1 to 23 kb pairs. For good
resolution, run 45 minutes on a 10-cm long, 1%
agarose gel in 0.5X TBE gel at 150 V.
Table 2:
Agarose concentrations for separating DNA
fragments of various sizes
effective range of resolution
agarose (%) of linear DNA fragments* (kb)
0.5 1.0–30
0.7 0.8–12
1.0 0.5–10
1.2 0.4–7
1.5 0.2–3
*Current Protocols in Molecular Biology, p 2.5.2 (1993)