Phase separation and liquid crystallization of complementary sequences in mixtures of nanoDNA oligomers

Abstract

Using optical microscopy, we have studied the phase behavior of mixtures of 12-to 22-bp-long nanoDNA oligomers. The mixtures are chosen such that only a fraction of the sample is composed of mutually complementary sequences, and hence the solutions are effectively mixtures of single-stranded and double-stranded (duplex) oligomers. When the concentrations are large enough, such mixtures phase-separate via the nucleation of duplex-rich liquid crystalline domains from an isotropic background rich in single strands. We find that the phase separation is approximately complete, thus corresponding to a spontaneous purification of duplexes from the single-strand oligos. We interpret this behavior as the combined result of the energy gain from the end-to-end stacking of duplexes and of depletion-type attractive interactions favoring the segregation of the more rigid duplexes from the flexible single strands. This form of spontaneous partitioning of complementary nDNA offers a route to purification of short duplex oligomers and, if in the presence of ligation, could provide a mode of positive feedback for the preferential synthesis of longer complementary oligomers, a mechanism of possible relevance in prebiotic environments.condensation ͉ nucleation ͉ depletion ͉ prebiotic ͉ PEG M olecular crowding of cellular interior is known to play a role in the spontaneous organization of biological macromolecules (1). Several biochemical and physiological processes are found to be influenced by strong packing constraints (2, 3), and complex ordered arrangements, such as liquid-crystalline mesophases, have been shown to arise from highly packed biomolecules (4). Of particular interest is the ordering of highly concentrated DNA in cell nuclei, which can lead to a variety of mesophases in vivo (5). However, whether such ordering caused by packing constraints had represented an evolutionary advantage remains an open question.Concentrated solutions of fully hybridized nanoDNA (nDNA) exhibit various liquid crystalline forms of supramolecular ordering, promoted by end-to-end adhesion of the paired bases at the terminals of the duplexes. This behavior has been reported recently for a wide set of self-complementary (SC) 6-to 20-bp nDNA sequences and for mutually complementary sequences in the same length range (6). Here we explore the phase behavior of mixtures of nDNA in which only some of the sequences are complementary, thus able to pair in double strands (DSs), and part of the sequences are not, and thus always remain in the solution as single strands (SSs). We have found that in concentrated mixtures of duplex and SS nDNA, the system phaseseparates into duplex-rich liquid crystal (LC) domains coexisting with a duplex-poor isotropic phase, leading to the physical segregation of 4-to 6-nm-long complementary chains from noncomplementary ones. This phase separation is a collective effect of the duplex and SS nDNA because the duplexes alone would not form LCs in such solutions, their concentration being well below that required for LC forma...

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