SUPPLEMENTARY DATA
Equilibrious Strand Exchange Promoted by DNA Conformational
Switching
Zhiguo Wu1, Xiao Xie1, Puzhen Li1, Jiayi Zhao1, Lili Huang1 and Xiang Zhou*1, 2
1
College of Chemistry and Molecular Sciences, Key Laboratory of Biomedical Polymers of Ministry of
Education, Wuhan University, Wuhan, Hubei, 430072, P. R. of China.
2
State Key Laboratory of Natural and Biomimetic Drugs, Peking University, Beijing, P. R. of China.
Table S1. The DNA sequences with and without c-myc (mu-c-myc).
Primers
Sequence (5’-3’)
label
GCATCTACGTATCGTAGATGCATT
24 bp
free
AATGCATCTACGATACGTAGATGC
24 nt
AATGCATCTACGATACGTAGATGC
5’-Cy5
TGGGGAGGGTGGGGAGGGTGGGGT
c-myc 24 bp
free
ACCCCACCCTCCCCACCCTCCCCA
c-myc 24 nt
G4-24 nt
ACCCCACCCTCCCCACCCTCCCCA
5’-Cy5
TGGGGAGGGTGGGGAGGGTGGGGT
5’-Cy5
TGCTTACGCTGTGCATCTACGTATCGTAGATGCATTTGAGGAT
AGGCT
48 bp
free
AGCCTATCCTCAAATGCATCTACGATACGTAGATGCACAGCGT
AAGCA
48 nt
AGCCTATCCTCAAATGCATCTACGATACGTAGATGCACAGCGT
AAGCA
5’-Cy5
TGCTTACGCTGTTGGGGAGGGTGGGGAGGGTGGGGTTGAGG
ATAGGCT
c-myc 48 bp
free
AGCCTATCCTCAACCCCACCCTCCCCACCCTCCCCAACAGCGT
AAGCA
c-myc 48 nt
AGCCTATCCTCAACCCCACCCTCCCCACCCTCCCCAACAGCGT
AAGCA
5’-Cy5
G4-48 nt
TGCTTACGCTGTTGGGGAGGGTGGGGAGGGTGGGGTTGAGG
ATAGGCT
5’-Cy5
ACGTATCTACTTTGCTTACGCTGTGCATCTACGTATCGTAGATG
CATTTGAGGATAGGCTCGTTCAACTGTA
72 bp
free
TACAGTTGAACGAGCCTATCCTCAAATGCATCTACGATACGTA
GATGCACAGCGTAAGCAAAGTAGATACGT
72 nt
TACAGTTGAACGAGCCTATCCTCAAATGCATCTACGATACGTA
GATGCACAGCGTAAGCAAAGTAGATACGT
5’-Cy5
ACGTATCTACTTTGCTTACGCTGTTGGGGAGGGTGGGGAGGG
TGGGGTTGAGGATAGGCTCGTTCAACTGTA
c-myc 72 bp
free
TACAGTTGAACGAGCCTATCCTCAACCCCACCCTCCCCACCCT
CCCCAACAGCGTAAGCAAAGTAGATACGT
c-myc 72 nt
G4-72 nt
TACAGTTGAACGAGCCTATCCTCAACCCCACCCTCCCCACCCT
CCCCAACAGCGTAAGCAAAGTAGATACGT
ACGTATCTACTTTGCTTACGCTGTTGGGGAGGGTGGGGAGGG
TGGGGTTGAGGATAGGCTCGTTCAACTGTA
5’-Cy5
5’-Cy5
TGGGGAGGGTGGAGAGGGTGGGGT
mu-c-myc 24bp
free
ACCCCACCCTCTCCACCCTCCCCA
mu-c-myc 24 nt
ACCCCACCCTCTCCACCCTCCCCA
5’-Cy5
TGCTTACGCTGTTGGGGAGGGTGGAGAGGGTGGGGTTGAGG
ATAGGCT
mu-c-myc 48 bp
free
AGCCTATCCTCAACCCCACCCTCTCCACCCTCCCCAACAGCGT
AAGCA
mu-c-myc 48 nt
AGCCTATCCTCAACCCCACCCTCTCCACCCTCCCCAACAGCGT
AAGCA
5’-Cy5
ACGTATCTACTTTGCTTACGCTGTTGGGGAGGGTGGAGAGGG
TGGGGTTGAGGATAGGCTCGTTCAACTGTA
mu-c-myc 72 bp
free
TACAGTTGAACGAGCCTATCCTCAACCCCACCCTCTCCACCCT
CCCCAACAGCGTAAGCAAAGTAGATACGT
mu-c-myc 72 nt
TACAGTTGAACGAGCCTATCCTCAACCCCACCCTCTCCACCCT
5’-Cy5
CCCCAACAGCGTAAGCAAAGTAGATACGT
ACGTATCTACTTTGGGGAGGGTGGGGAGGGTGGGGTTGGGG
AGGGTGGGGAGGGTGGGGTCGTTCAACTGTA
2c-myc 72 bp
free
TACAGTTGAACGACCCCACCCTCCCCACCCTCCCCAACCCCAC
CCTCCCCACCCTCCCCAAAGTAGATACGT
2c-myc 72 nt
TACAGTTGAACGACCCCACCCTCCCCACCCTCCCCAACCCCAC
CCTCCCCACCCTCCCCAAAGTAGATACGT
5’-Cy5
Figure S1. The PAGE analysis of the SERs of dsDNAs and homologous ssDNAs. TE buffer was used
instead of sodium phosphate buffer. The invading strands contained the complementary strand of
c-myc. Lane 1: SERs of random sequences; lane 2: SERs of c-myc containing sequences; lane 3:
dsDNAs markers of random sequences; lane 4: dsDNAs markers of c-myc sequences; lane 5: ssDNAs
markers of random sequences; lane 6: ssDNAs markers of c-myc sequences. The SERs were carried
out in the presence of 200 mM KCl, 20 mM TE, pH 7.4. The incubation temperature was 37 oC, and the
incubation times of 24 bp and 24 nt, 48 bp and 48 nt, 72 bp and 72 nt, 2c-myc 72 bp and 72nt were 3
h, 30 h, 60 h, 60 h, respectively.
Figure S2. The CD spectrum of c-myc and mu-c-myc sequences (single strands) at the condition of:
200 mM KCl, 20 mM sodium phosphate, pH 7.4. 5 μM ssDNA for c-myc and mu-c-myc.
Figure S3. The CD spectra of the duplexes containing c-myc (were in red, c-myc 24 bp, A; c-myc 48
bp, B; c-myc 72 bp, C) and not containing c-myc (were in black, random 24 bp, A; random 48 bp, B;
random 72 bp, C). In the right panel, the CD spectra of a, c-myc 24 bp – random 24 bp; b, c-myc 48
bp – random 48 bp; c, c-myc 72 bp – random 72 bp were obtained. The CD spectra were obtained at
the same conditions to the SERs (200 mM KCl, 20 mM sodium phosphate, pH 7.4). The dsDNAs
concentrations were 5 μM, 2 μM and 2 μM for 24 bp, 48 bp and 72 bp, respectively.
Figure S4. In left panel, the CD spectra of mu-c-myc 24 bp and random 24 bp, mu-c-myc 48 bp and
random 48 bp, mu-c-myc 72 bp and random 72 bp were recorded. In right panel, the difference values
of mu-c-myc sequences and random sequences were obtained. The CD spectra were performed at 5 μM,
2 μM and 2 μM for 24 bp, 48 bp and 72 bp, respectively, under the condition of 200 mM KCl, 20 mM
sodium phosphate, pH 7.4.
Figure S5. The c-myc promoted SER kenetics. The SERs DNAs were adjusted to 20 μM dsDNAs (bp)
and 20 μM ssDNAs (nt), buffered by 200 mM KCl, 20 mM sodium phosphate, pH 7.4. Then the SERs
were carried out by gradiently increasing the incubating time from 0 to 7 h, 33 h and 72 h for A, B, and
C, respectively. The SERs were stopped by cooling to 4 oC, and then bromophenol blue was added as
the loading buffer. 20% PAGE gel containing 200 mM KCl, 20 mM TBE (pH 7.4) was employed to
electrophorese the DNA samples. All the c-myc contained sequences (green) and random sequences
(yellow) were placed together with the PAGE gel results. The Cy5-labeled complementary strands of
c-myc contained sequences were red, and the Cy5-labeled complementary strands of random
sequences were yellow.
Figure S6. The Tm values of the random sequence ssDNAs. The normalized Tm experiments were
carried out at 10 μM, 5 μM, 2 μM single strand for 24 nt and 24 nt com, 48 nt and 48 nt com, 72 nt and
72 nt com, respectively. In the same buffer as SERs, 200 mM KCl, 20 mM sodium phosphate, pH 7.4.
The system temperatures were increased gradually from 25 oC to 90 oC, and the ultraviolet absorbance
at 260 nm were monitored to calculate the Tm values for each ssDNA.
Figure S7. The denaturing PAGE gel electrophoresis of Cy5 labeled ssDNAs. 6 M urea and 40%
formamide were added into 20% PAGE gel to denature the single stranded DNAs. The experiment was
carried out in the buffer containing 20 mM KCl, 20 mM TBE, pH 7.4. Lane 1: random 24 nt com; lane 2:
c-myc 24 nt com; lane 3: random 48 nt com; lane 4: c-myc 48 nt com; lane 5: random 72 nt com; lane
6: c-myc 72 nt com. The concentration of ssDNAs was 2μM.
Figure S8. The Tm values of the dsDNAs with or without c-myc. The normalized Tm experiments were
carried out at 5 μM, 2 μM, 2 μM double strand for 24 bp (A), 48 bp (B), 72 bp (C) lengths DNA,
respectively. In the same buffer as SERs, 200 mM KCl, 20 mM sodium phosphate, pH 7.4. The system
temperatures were increased gradually from 25 oC to 90 oC, and the ultraviolet absorbances at 260 nm
were monitored to calculate the Tm values for each duplex.
Figure S9. The UV-melting temperature of all the double stranded DNAs used in this paper, recorded
at 260 nm. The Tm experiments were performed 5 μM, 2 μM and 2 μM for 24 bp, 48 bp and 72 bp,
respectively, under the condition of 20 mM KCl, 20 mM NaCl, 20 mM TE, pH 7.4.
Figure S10. The SER of 2c-myc sequence as the control. A, the PAGE analysis of the SER, lane 1: SER
of random 72 bp and 72 nt; lane 2: SER of c-myc 72 bp and 72 nt; lane 3: SER of 2c-myc 72 bp and 72
nt; lane 4: dsDAN marker of random sequence; lane 5: dsDNA marker of c-myc sequence; lane 6:
dsDNA marker of 2c-myc sequence; lane 7: ssDNA marker of random sequence; lane 8: ssDNA marker
of c-myc sequence; lane 9: ssDNA marker of 2c-myc sequence. These SERs were carried out at 37 oC,
60 h, in the buffer of 200 mM KCl, 20 mM sodium phosphate, pH 7.4. B, the histogram of SERs product
concentrations extracted from A, the random sequence, c-myc sequence and 2c-myc sequence.
Figure S11. The SERs of dsDNAs and ssDNAs with mutated c-myc (mu-c-myc) and with the random
sequences. Lane 1: the random sequences; lane 2: the mu-c-myc contained sequences; lane 3:
dsDNAs with random sequence; lane 4: dsDNAs with the mu-c-myc sequence; lane 5: ssDNAs with
random sequence; lane 6: ssDNAs with mu-c-myc sequence. The SERs were carried out under the
condition of 200 mM KCl, 20 mM sodium phosphate, pH 7.4. The incubation times of 24 bp and 24 nt,
48 bp and 48 nt, 72 bp and 72 nt were 3 h, 30 h, 60 h, respectively. And the incubation was performed
at 37 oC.
Figure S12. The SERs containing single-base mismatch system. Lane 1: random sequences without
c-myc; lane 2: c-myc contained sequences; lane 3: the sequences containing 1 mismatch in the double
strands, and the Cy5-labeled sequences were the fully complementary strands of the mu-c-myc
contained strands; lane 4: dsDNAs with random sequence; lane 5: dsDNAs with mu-c-myc sequence;
lane 6: ssDNAs with random sequence; lane 7: ssDNAs with mu-c-myc sequence. The SERs were
performed under the same conditions to Figure 2 in main text. The single-base mismatch sequences
were presented under the PAGE gel.
Figure S13. The random sequences and c-myc containing sequences formed double stranded DNA
duplexes under the SERs conditions. Lane 1: DNA markers; lane 2: random 24 bp; lane 3: c-myc 24 bp;
lane 4: random 48 bp; lane 5: c-myc 48 bp; lane 6: random 72 bp; lane 7: c-myc 72 bp; lane 8: 2c-myc
72 bp. The DNAs were incubated at 37 oC, 200 mM KCl, 20 mM sodium phosphate, pH 7.4. Then native
PAGE was employed to analysis the DNA samples, stained by EB, and the image was collected by UV
imager.
Figure S14. The SERs between dsDNAs and Cy5-labeled ssDNAs mediated by Rec A in the absence of
ATP. Lane 1: random 24 bp, nt; lane 2: c-myc 24 bp, nt; lane 3: random 48 bp, nt; lane 4: c-myc 48
bp, nt; lane 5: random 72 bp, nt; lane 6: c-myc 72 bp, nt. All the SER conditions were identical with the
experiment in Figure 6 except ATP, and no SER product was observed.
Figure S15. The Tm values of the double stranded DNAs in the presence of 40% PEG200. The
normalized Tm experiments were carried out at 5 μM, 2 μM, 2 μM dsDNAs for 24 bp and c-myc 24 bp,
48 bp and c-myc 48 bp, 72 bp and c-myc 72 bp, respectively. The buffer contains 5 mM KCl, 5 mM TE,
and 40% PEG200 as the crowding agent pH 7.4. The system temperatures were increased gradually
from 25 oC to 90 oC, and the ultraviolet absorbance at 260 nm were monitored to calculate the Tm
values for each dsDNA.