Figure 1.
Structure drawing of single-stranded circular syn-G4/m-G4 genome (5577 bp) and sites of restriction endonucleases.
New NcoI, KpnI and EcoRI sites marked in red slash were added into the m-G4 genome. Three specific PCR fragments were shown in yellow.
Figure 2.
Close up images of the plaques (A, syn-G4; B, m-G4) and the bacterial lawns without plaques (C, control).
The plaques appeared small, clear and round. Some merged or ran together to form a mass. The appearance of the plaques of both syn-G4 and m-G4 showed no difference in A and B (A, syn-G4; B, m-G4).
Figure 3.
Synthetic G4 morphology by transmission electron microscope (Arrow).
Magnification: 800,000×. Bar: 100 nm.
Figure 4.
Agarose gel showing the PCR results obtained when specific primers were used. Lane 1, D2000 marker; Lane 2, 3 and 4, PCR amplification of the three specific products 603 bp, 443 bp and 645 bp, respectively, of syn-G4 (Location shown in Fragment 3, 1 and 2 in yellow in Fig. 1, respectively); Lane 4, 5 and 6, PCR amplification of the three specific products 603 bp, 443 bp and 645 bp, respectively, of m-G4 (Location shown in Fragment 3, 1 and 2 in yellow in Fig. 1, respectively).
Figure 5.
Digestion of restriction endonuclease.
Lane 1, D2000marker. Agarose gels (Lane 2, 4, 6) showing the products (shown in Fragment 2 in Fig. 1) of syn-G4 digested by restriction endonucleases and generations of restriction endonuclease digestion fragments of 645 bp (EcoRI), 125 bp+520 bp (KpnI) and 645 bp (NcoI), respectively. The 3 modified restriction endonuclease sites of m-G4 were confirmed by agarose gels and different digestion fragments of 518 bp+127 bp (EcoRI), 125 bp+423 bp+97 bp (KpnI), and 209 bp+436 bp (NcoI) (Lane 3, 5 and 7 respectively), were observed.
Table 1.
Primers for specific PCR.