Discussion
Experiment 1 showed that desired hybridizations occurred in the combinations of st1-l1, l1-j2, l1-j3 and j3-j3. In all cases, there were great peaks at higher temperature. This means that the hybridizations of these strands would have enough robustness to maintain structure. The combination of st1~st4 can make X-motif DNA hydrogel. Above sets of hybridization implied that swelling and shrinking DNA hydrogel could stable thermodynamically.
Figure D-1. st1-l1 hybridization
st1-l1 hybridization has a role to bridge adjacent X-motifs.
Figure D-2. l1-j2, j3 hybridization
l1-j2 hybridization has a role to open l1 loop for swelling. l1-j3 hybridization also open loop, but can handle with next shrinking.
Figure D-3. j3-j3' hybridization
j3-j3' hybridization has a role to remove j3 from DNA hydrogel and recover l1 loop.
Experiment 2 achieved to make DNA beads. The size of DNA beads was uniform, around 40 µm and this contributed for measurement of hydrogel area. However, DNA spider-web could not be made. Some hypothesis why hydrogel was not formed are suggested, effect of bubbles and manipulation mistakes, but understandable reasons were not clear. Further investigation is needed.
Experiment 3 showed DNA hydrogel swelled. The rates correlated to [j2], and maximum area is around 1.6~1.8 times. From experiment 1, desired hybridization occurred, so in theory, maximum area should be around 3.42 times(1.85**2, size of the DNA hydrogel mesh becomes 1.85 times). Compared with this calculation, observed area was much smaller. It is thought that even if all of j2 hybridized with l1, mesh size does not necessarily 1.85 times. DNA is not so rigid and tolerant for some distortion. Otherwise, j2 could not enter the core of DNA hydrogel, hybridized with l1 just around surface. Longer time scale experiment may give more swelled DNA hydrogel.
Experiment 4 showed DNA hydrogel shrunk. However, the shrunk size was not the same as that before swelling. One reason for this is thought that j3' could remove only surface j3 and multiplex, j3-j3', could not leave from DNA hydrogel net. This also can be improved by longer time scale experiment.
Figure D-4. Interaction area of DNA hydrogel
It is thought that the core of DNA hydrogel(gray) is unaccessible with strands in solution just around surface(yellow) interact with strands. If so, once the hydrogel swelled, unaccessible area become larger. As a result, after the gel shrank, the size of gel becomes slightly larger.
Conclusion
Through this project, we aimed to develop swelling and shrinking spider-web DNA hydrogel which acts in millimeter scale. We checked designed strands, st1~4, l1, j2, j3 and j3’ representing desired hybridizations in meltcurve test and also showed DNA hydrogel swelling 1.6~1.8 times in maximum and shrinking 0.85 times in minimum by DNA hydrogel beads test.
We are succeed to generate swelling and shrinking DNA hydrogel. Further research can realize swelling and shrinking spider-web DNA hydrogel. This research opened a door for millimeter-scale moving DNA hydrogel.