Chemistry Reference
In-Depth Information
shows the sequences of pcPNA additives (as well as those of DNA substrate near the
scission sites).
7.6.2.1
Invasion of Two pcPNAs into Double-stranded DNA
The invasion complex was formed simply by incubating the linearized DNA with
pcPNA
(1)
and pcPNA
(2)
, as clearly evidenced by a gel shift assay. These two pcPNA
additives are complementary with C1826-A1840 in the upper strand of the linearized
DNA and A1821-G1835 in its lower strand, respectively [Figure 7.9(a)]. Accordingly,
when these pcPNA additives invade the linearized DNA, a gap-like structure is formed
in each strand [T1821-T1825 of the upper strand and G1836-T1840 in the lower strand:
underlined bases in Figure 7.9(a)]. These gap-like (single-stranded) sites are the scis-
sion targets.
Figure 7.10 Agarose gel electrophoresis patterns for Ce(
IV
)/EDTA-induced
site-selective hydrolysis of double-stranded DNA by Ce(
IV
)/EDTA and pcPNA
additives. Bands were detected by staining with GelStar. (a) Site-selective
hydrolysis of linearized PBR322 using pcPNAs. Lane 1, control; lane 2, Ce(
IV
)/
EDTA only; lane 3, pcPNA
(1)
/pcPNA
(2)
+Ce(
IV
)/EDTA; lane 4, pcPNA
(3)
/
pcPNA
(4)
+Ce(
IV
)/EDTA; M, 1000 base pair ladder. [linearized PBR322] = 4 n
M
,
[each of pcPNAs] = 100 n
M
,[Ce(
IV
)/EDTA] = 20
M
, [NaCl] = 100m
M
,atpH7.0
(5.0m
M
Hepes buffer) and 37
8
C. (b) Direct site-selective hydrolysis of su-
percoiled PBR322. Lane 1, control; lane 2, pcPNA
(1)
/pcPNA
(2)
+Ce(
IV
)/EDTA;
lane 3, EcoRI digests of the products in lane 2.
l
