Sequence‐specific nucleic acid mobility using a reversible block copolymer gel matrix and DNA amphiphiles (lipid‐DNA) in capillary and microfluidic electrophoretic separations

Abstract

Reversible noncovalent but sequence-dependent attachment of DNA to gels is shown to allow programmable mobility processing of DNA populations. The covalent attachment of DNA oligomers to polyacrylamide gels using acrydite-modified oligonucleotides has enabled sequence-specific mobility assays for DNA in gel electrophoresis: sequences binding to the immobilized DNA are delayed in their migration. Such a system has been used for example to construct complex DNA filters facilitating DNA computations. However, these gels are formed irreversibly and the choice of immobilized sequences is made once off during fabrication. In this work, we demonstrate the reversible self-assembly of gels combined with amphiphilic DNA molecules, which exhibit hydrophobic hydrocarbon chains attached to the nucleobase. This amphiphilic DNA, which we term lipid-DNA, is synthesized in advance and is blended into a block copolymer gel to induce sequence-dependent DNA retention during electrophoresis. Furthermore, we demonstrate and characterize the programmable mobility shift of matching DNA in such reversible gels both in thin films and microchannels using microelectrode arrays. Such sequence selective separation may be employed to select nucleic acid sequences of similar length from a mixture via local electronics, a basic functionality that can be employed in novel electronic chemical cell designs and other DNA information-processing systems.

Electronic pH switching of DNA triplex reactions

“…22,62 Whereas UV and CD spectroscopic methods allowed us to study equilibria between DNA strands, we also investigated the disproportionation of molecular complexes during strand separation, since reveals kinetic information, because transport of DNA can be used as an essential component of triplex based DNA replication schemes. 49 We intentionally designed our triplex oligonucleotides ss Y24 as well as ss R24 with an overall structure similar to the previously reported one used to investigate triplex-based DNA replication. 40 The hairpin DNA template ds H24 enables simpler (bimolecular) binding equilibria, clearer control of triplex labelling as well as better thermal stability of the ds template 63 (see also ESI Fig.…”

“…Fast separation as well as online monitoring of DNA was achieved in Pluronic® thermoresponsive gel matrix, 48 which as a reversible gel also allows further integration of this fractional analysis of DNA complexes with microuidic channels. 49 Following the Materials and methods section, the main results of the paper are presented, relegating supporting and background experimental data to the ESI. † Firstly, a spectroscopic analysis and comparison of the intermolecular Y:R:Y with the hairpin/single stranded H:Y triplex complexes are presented.…”

Separation of single‐base sequential isomers of single‐stranded DNA by capillary electrophoresis and its application in the discrimination of single‐base DNA mutations

“…Past studies employing CE for DNA separation mainly focused on size‐based techniques using molecular‐sieving media such as gel or polymer matrixes . Recently, CE separation of sequential DNA isomers having single‐base substitutions and the same chain length has attracted the attention of researchers because it aids in the identification of gene mutations . However, DNA separation remains a highly challenging task because of a number of technical difficulties.…”

Optomagnetic detection of DNA triplex nanoswitches

“…1 The predictable and pH-controlled base pairing makes triplex-forming sequences very useful for programming chemical reactions, 2,3 nanomachines, 4,5 nanostructures, [6][7][8] and hydro-gels. 9,10 Switching of pHresponsive DNA composites can be employed for controlled release of targets in delivery systems 11,12 and in on/off regulation of the nanoscale devices upon cyclic alternation of pH values. 13 Further, triplex-forming sequences of DNA are known to play a significant role in genetic regulation.…”

“…17 For the latter, the assay time can be reduced to 10-15 min using capillary gel electrophoresis. 3,9 Palindromic homopurine-homopyrimidine tracts have been shown to cause a pHdependent structural transition of a plasmid DNA in vivo and have been observed during the initial replication of tumor viruses. 20 Therefore, dosedependent detection of polypurine tracts (PPT) in human and viral genomes can contribute to molecular diagnostics of genetic biomarkers such as the conserved PPT region of HIV-1.…”