Biomedical Engineering Reference
In-Depth Information
Fig.
3
CPEC of a 258 bp gene. The image shows gel electrophoresis analysis of
the CPEC reaction product after 1, 2, and 5 cycles (lanes 1-3) together with a
full-length plasmid (lane 4) and 1 kb DNA ladder (lane M, New England Biolabs).
The assembled full-length plasmid is 2,644 bp shown as the
upper band
of the
two prominent bands close to each other. The
lower band
of the two is the empty
vector of 2,386 bp that has not been incorporated within reaction cycles
Fig.
4
Agarose gel examination of the CPEC product for a 258 bp gene library
after 5 thermal cycles (lane 2) together with linear vector (lane 1) and 1 kb DNA
ladder (lane M, New England Biolabs), respectively
2. Run the CPEC reaction using the following conditions
(
see
Note 7
) (Fig.
4
):
Cycle number
Denature
Anneal
Extend
1
98 °C, 30 s
2-6
98 °C, 10 s
(
Tm
+ 3) °C, 30 s (
see
Note 5
)
72 °C, 15 s/kb (
see
Note 6
)
7
72 °C, 5 min
