Review Reports

Round 1

Reviewer 1 Report

Gao et al. present a way to manipulate the properties of DNA hydrogels using DNA tiles, different linker lengths and nanomaterials (CNTs and AuNPs).

The paper is scientifically sound and could be of interest to the DNA hydrogel community. I recommend publication after a few concerns/questions have been addressed:

1) L79: I don’t think describing the bond as dative is correct. Indeed I do not believe that the true nature of the bonding is truly known (see e.g. https://www.nature.com/articles/nchem.1352.pdf?origin=ppub)

2) L83: Why did the authors decide for the polyA method of preparing DNA-AuNP conjugates? With this method the loading should be reasonably low (cf. : https://pubs.acs.org/doi/pdf/10.1021/acs.chemrev.8b00733) and particles are less stable. Was the loading efficiency calculated?/Was it important for the application? Would higher DNA loading result in a higher degree of cross-linking and thus in an altered property of the resulting hydrogel?

3) The authors only used one size of AuNPs. Would they expect different sizes to result in different properties of the hydrogel?

4) Figure 5 would benefit from more description inside the figure (e.g. a header above the left and right panels for hydrogel and conjugates and a label or schematic for each type (i.e. SWCNT vs MWCNT vs AuNPs).

5) Also, Figures 5b and c are not explicitly mentioned in the manuscript at all.

6) Figure 5c) Why is the red graph for cycle 1 the only one that passes through the origin?

7) The authors claim that "...the trend of slight decreases in current over successive pulses..." This is not visible in the graphs. Maybe an inset would help here.

Author Response

Response to Reviewer 1:

1) L79: I don’t think describing the bond as dative is correct. Indeed I do not believe that the true nature of the bonding is truly known.

            We appreciate the comment and have modified the sentence accordingly.

2) L83: Why did the authors decide for the polyA method of preparing DNA-AuNP conjugates? With this method the loading should be reasonably low and particles are less stable. Was the loading efficiency calculated? Was it important for the application? Would higher DNA loading result in a higher degree of cross-linking and thus in an altered property of the resulting hydrogel?

            We chose polyadenine as the anchoring group because we want to use unmodified DNA (i.e. nonthiolated) to adsorb onto AuNP surfaces, and polyadenine has shown great ability to become efficiently physisorbed on gold. An A13 tail has especially shown strong and consistent adsorption since adenine displays strong interaction with gold. Please refer to our reference 39 for discussion of loading efficiency and comparison with other types of anchors. For our purposes, it is not necessary to achieve the highest loading capacity as long as the DNA crosslinking can efficiently assemble percolating nanocircuits with AuNP here.

3) The authors only used one size of AuNPs. Would they expect different sizes to result in different properties of the hydrogel?

            It would be interesting to explore the influence of AuNP of different sizes. We see that as a future step for this line of study.

4) Figure 5 would benefit from more description inside the figure (e.g. a header above the left and right panels for hydrogel and conjugates and a label or schematic for each type (i.e. SWCNT vs MWCNT vs AuNPs). 

            Thank you. We have modified Figure 5 accordingly.

5) Also, Figures 5b and c are not explicitly mentioned in the manuscript at all.

            We apologize for the oversight. We have added explicit reference to both of those figure panels.

6) Figure 5c) Why is the red graph for cycle 1 the only one that passes through the origin?

            We believe that the hollow loop structure seen in Figure 5c is due to some capacitance effect within the gold-containing composite, but our confidence level in this interpretation is not yet high enough to make such a statement in the text. We have therefore, let the data speak for itself at this point, and we hope that community discussion of the results will assist us in defining further experiments, collecting data, and further understanding the observed behavior.

7) The authors claim that "...the trend of slight decreases in current over successive pulses..." This is not visible in the graphs. Maybe an inset would help here.

            We thank the reviewer for pointing this sentence out to us. It was actually present for describing another figure panel that has since be deleted. Therefore, the sentence was no longer appropriate in the text, and we have deleted it.

Reviewer 2 Report

This manuscript utilises a bottom-up DNA self-assembly approach to construct functional nanocomposite hydrogels. The design and construction of multifunctional self-assembled materials have significant fundamental and technological importance and therefore this work will be of interest to the researchers in the field. 

The authors have used various physicochemical characterisation techniques to study the DNA based nanocomposites hydrogels and data is presented in a convincing manner. However, there are few queries 

1. Though the authors have presented a schematic representation for DNA based nanocomposite hydrogels, from the readers' point of view, it would be a good idea to present photographic images of the control gel, DNA-CNT and DNA-Au hydrogels.
2. For rheological studies, the authors have taken 30 uL of hydrogel sample - it is not clear how such small quantities were measured. If micropipettes were used then it suggests that the hydrogels have been subjected to shear forces prior to the rheological studies, which will have a significant influence on the final properties. 
3. DNA stabilised CNT nanostructures/dispersions are well known. To confirm the DNA is associated with the CNT - TEM studies, Figure 2 (d to h), have the authors considered to perform EDX analysis to show the presence of Phosphorous? 
4. The authors have used DNA sequences to programme the hydrogels - in this view, it would be useful to know the physical properties of the control and nanocomposite hydrogels, for instance - the effect of temperature that is would the hydrogels disassemble due to heat-induced denaturation/melting. Is the gelation processes reversible with cooling and re-hybridization of the DNA network. 
5. Similarly, what would be the effects of the temperature on the optical properties of DNA-Au hydrogels.

Other comments - SI information was not accessible! 

In summary, the work present in the manuscript provides a new approach for the fabrication of functional DNA based nanocomposite hydrogels. Therefore, the manuscript can be recommended for publication provided that the address above mentioned queries in the revised manuscript. 

 

 

Author Response

Response to Reviewer 2:

1. Though the authors have presented a schematic representation for DNA based nanocomposite hydrogels, from the readers' point of view, it would be a good idea to present photographic images of the control gel, DNA-CNT and DNA-Au hydrogels.

            The optical images of hydrogels can be found at Figure S2 in the supplementary materials, which is submitted in a separate file.

 

2. For rheological studies, the authors have taken 30 uL of hydrogel sample - it is not clear how such small quantities were measured. If micropipettes were used then it suggests that the hydrogels have been subjected to shear forces prior to the rheological studies, which will have a significant influence on the final properties.

            This is a very good point, and to clarify how we avoided any shear problem we have added the follow sentence in the Methods section: “We prepared DNA hydrogel samples by pipetting the components onto parafilm; following gel formation, samples were transferred to the rheometer plate without pipetting, to avoid shearing the gels.”

 

3. DNA stabilised CNT nanostructures/dispersions are well known. To confirm the DNA is associated with the CNT - TEM studies, Figure 2 (d to h), have the authors considered to perform EDX analysis to show the presence of Phosphorous?

            The reviewer’s suggestion is a good one, and one that we will follow during the next iteration of these experiments. Unfortunately, we do not have time to run EDX quickly enough at this time to add it as characterization for the present report.

 

4. The authors have used DNA sequences to programme the hydrogels - in this view, it would be useful to know the physical properties of the control and nanocomposite hydrogels, for instance - the effect of temperature that is would the hydrogels disassemble due to heat-induced denaturation/melting. Is the gelation processes reversible with cooling and re-hybridization of the DNA network.
5. Similarly, what would be the effects of the temperature on the optical properties of DNA-Au hydrogels.

            The reviewer is correct that the temperature dependence of the physical and optical properties of the hydrogels would be interesting to study. The simplest DNA based hydrogels have been shown to melt and reform reversibly, and we would expect similar behavior from our composites. However, with different associations occurring at somewhat different temperatures, the composites might form weaker structures following a melt/reanneal cycle. One would also expect the optical properties to change in the AuNP-containing gels, if the nanoparticles shift into or out of plasmonic coupling distance. For the most part, we have been interested in the properties of as-formed gels, however, examination of melting and reformation would be an interesting follow-up line of investigation for a future study. We thank the reviewer for these suggestions.