Deoxyribonucleic acid (DNA) hydrogels, a three-dimensional (3D) network made from DNA chains, have attracted great attention because of its molecular programmability, excellent biocompatibility and wide biomedical applications. Construction of hydrogel incorporating genetic function is still a challenge because of the limitations in available preparation methods. Herein, we develop a polymerase chain reaction (PCR) based strategy to construct gene integrated hydrogel to mimic the biofunction of nucleus zone. DNA primers were chemically modified by methacrylamide, which were used as modular primers in PCR to hybridize with template plasmid DNA, yielding methacrylamide functionalized gene (Acry-gene). Afterwards, Acry-gene was chemically cross-linked and compressed via free radical polymerization of terminal group methacrylamide to form a three-dimensional gene network, namely gene hydrogel. The gene hydrogel retained the genetic function and expressed protein successfully in a cell free protein expression system. This work provides a general approach for the construction of biofunctional gene hydrogel which mimics bioprocesses, showing great potential in biomedicine and biomimetic fields.
the National Natural Science Foundation of China(21621004,21575101,21622404)
This work was supported by the National Natural Science Foundation of China (21621004, 21575101, 21622404).
The authors declare that they have no conflict of interest.
These authors contributed equally to this work.
The supporting information is available online at
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Figure 1
Agarose gel electrophoresis analysis of PCR produced Acry-eGFP and the effect of concentration of APS on the free radical polymerization of Acry-eGFP. (a) Schematic diagram of the preparation of polymerized-gene. (b) Gel electrophoresis image of Acry-eGFP with
Scheme 1
Scheme of the preparation of gene hydrogel and protein expression in a CFPS system. Acry-primer sequences hybrided with plasmid DNA template and were extended by polymerase to synthesize Acry-gene via PCR process. Acry-gene was afterwards cross-linked via free radical polymerization of methacrylamide groups to form a 3D gene network, namely gene hydrogel. The gene hydrogel preserved the biological function of DNA and expressed protein in CFPS system (color online).
Figure 2
Formation and rheological property
Figure 3
SEM and fluorescence microscopy of gene hydrogel. (a, b) The morphology of Hydrogel-1235 (the arrow indicates nanofibers); (c) the fluorescence microscope image of the Hydrogel-1235 (stained with SYBR Green I); (d, e) the morphology of Hydrogel-2393; (f) the fluorescence microscope image of the Hydrogel-2393 (stained with SYBR Green I) (color online).
Figure 4
Protein expression of gene hydrogel in a CFPS system. (a) Schematic illustration of protein expression in a gene hydrogel containing CFPS system; (b) time course of the eGFP expressed from Acry-eGFP Hydrogel-1235 in a CFPS system; (c) time course of the eGFP expressed from Acry-eGFP Hydrogel-2393 in a CFPS system (color online).
Name | Sequence | Use |
Primer 1 | TAACCGTATTACCGCCTTTGAGT | Forward primerfor PCR |
Primer 2 | GCGGGCCTCTTCGCTATTA | Reverse primer for PCR |
Primer 3 | GCGGGCCTCTTCGCTATTA | Reverse primer for PCR |
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