Activation of DNA strand exchange by cationic comb-type copolymers: effect of cationic moieties of the copolymers

Choi, Sung Won; Kano, Arihiro; Maruyama, Atsushi

doi:10.1093/nar/gkm1035

Cited by 38 publications

(27 citation statements)

References 40 publications

(55 reference statements)

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“…In future studies, it will also be interesting to test thermostable RNA ligases (Blondal et al 2003(Blondal et al , 2005 as well as the influence of additives such as PEG-8000 (Miyoshi and Sugimoto 2008) or compounds such as cationic comb-type copolymers (CCC), both of which enhance the dynamic of RNA folding (Kim et al 2003;Choi et al 2007). It is possible that these conditions lead to an increased rate of interconversion between secondary structure and thereby allow for more product accumulation, given that adapters are always used in excess over input RNAs.…”

Section: Discussionmentioning

confidence: 99%

RNA-ligase-dependent biases in miRNA representation in deep-sequenced small RNA cDNA libraries

Hafner¹,

Renwick²,

Brown³

et al. 2011

RNA

307

367

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Sequencing of small RNA cDNA libraries is an important tool for the discovery of new RNAs and the analysis of their mutational status as well as expression changes across samples. It requires multiple enzyme-catalyzed steps, including sequential oligonucleotide adapter ligations to the 39 and 59 ends of the small RNAs, reverse transcription (RT), and PCR. We assessed biases in representation of miRNAs relative to their input concentration, using a pool of 770 synthetic miRNAs and 45 calibrator oligoribonucleotides, and tested the influence of Rnl1 and two variants of Rnl2, Rnl2(1-249) and Rnl2(1-249)K227Q, for 39-adapter ligation. The use of the Rnl2 variants for adapter ligations yielded substantially fewer side products compared with Rnl1; however, the benefits of using Rnl2 remained largely obscured by additional biases in the 59-adapter ligation step; RT and PCR steps did not have a significant impact on read frequencies. Intramolecular secondary structures of miRNA and/or miRNA/39-adapter products contributed to these biases, which were highly reproducible under defined experimental conditions. We used the synthetic miRNA cocktail to derive correction factors for approximation of the absolute levels of individual miRNAs in biological samples. Finally, we evaluated the influence of 59-terminal 5-nt barcode extensions for a set of 20 barcoded 39 adapters and observed similar biases in miRNA read distribution, thereby enabling cost-saving multiplex analysis for large-scale miRNA profiling.

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Section: Discussionmentioning

confidence: 99%

RNA-ligase-dependent biases in miRNA representation in deep-sequenced small RNA cDNA libraries

Hafner¹,

Renwick²,

Brown³

et al. 2011

RNA

307

367

View full text Add to dashboard Cite

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“…[24][25][26] We previously showed that a cationic comb-type copolymer with guanidino groups accelerates SER relative to the unmodified copolymer. 27 Guanidine modification did not significantly alter cationic properties of the copolymer but did change the hydrogen-bonding capacity. The guanidinium ion is also known to have chaotropic effects that destabilize dsDNA, which should accelerate SER.…”

Section: Introductionmentioning

confidence: 92%

DNA strand exchange reaction activated by cationic comb-type copolymers having ureido groups

2014

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Strand exchange reactions (SERs) of nucleic acids are essential for genetic recombination and often require nucleic acid chaperone activity. A variety of proteins induce strand exchange reactions in vivo. We have reported that cationic comb-type copolymers composed of a polycation backbone and hydrophilic graft chains greatly accelerate the SER in vitro and exhibit nucleic acid chaperone like activity. To elucidate structure/function relationships involved in the SER accelerating activity of the copolymer, we modified the copolymer with ureido groups. The melting temperature of dsDNA in the presence of the copolymers with ureido groups decreased with increasing ureido content, suggesting that the ureido groups have a destabilizing effect on the DNA duplex. Copolymers modified with ureido groups (about 50 mol%) accelerated a SER more strongly than unmodified copolymers. Given our previous observation that modification of the copolymer with guanidino groups resulted in enhanced chaperone-like activity, we propose that incorporation of chaotropic functional groups into the copolymer structure results in an increase in nucleic acid chaperone-like activity.

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“…Binding of DNA to the copolymer was assessed by fluorescence correlation spectroscopy [18,36,37] using an Olympus MF20 single-molecular fluorescence detection system (Olympus, Tokyo Japan). All samples for the fluorescence …”

Section: Dna Binding To Copolymer As Determined By Fluorescence Corrementioning

confidence: 99%

“…To further understand the effect of the copolymer, binding of copolymer to either ssDNA or tetramolecular quadruplex DNA was analyzed by fluorescence correlation spectroscopy [18,36,37]. Since a diffusion time calculated by the autocorrelation analysis of the fluorescence fluctuation is proportional to cubic root of molecular mass, the complex formation of oligonucleotides with the copolymer leads to an increased in the diffusion time.…”

Section: Binding Assay Of the Copolymer To Ssdna And Tetramolecular Qmentioning

confidence: 99%

DNA assembly and re-assembly activated by cationic comb-type copolymer

et al. 2011

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Guanine-rich oligonucleotides, such as TG(4)T and TG(5)T, assemble into a tetramolecular quadruplexes with layers of G-quartets stabilized by coordination to monovalent cations. Association rates of the quadruplexes are extremely slow, likely owing to electrostatic repulsion among the four strands. We have shown that comb-type copolymers with a polycation backbone and abundant hydrophilic graft chains form water-soluble polyelectrolyte complexes with DNA and promote DNA hybridization. Here, we report the effect of cationic comb-type copolymers on the kinetics of tetramolecular quadruplex formation. The copolymer significantly increased the association rate of tetramolecular quadruplexes without altering kinetic effects of metal cations in quadruplex formation. Dissociation rates of the quadruplexes were also accelerated by the copolymer suggesting that the copolymer has chaperone-like activity that reduces the energy barriers associated with dissociation and re-assembly of base pairs. This hypothesis was further supported by the observation that the copolymer activated the strand exchange reaction between the quadruplex and a constituting single-stranded.

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Activation of DNA strand exchange by cationic comb-type copolymers: effect of cationic moieties of the copolymers

Cited by 38 publications

References 40 publications

RNA-ligase-dependent biases in miRNA representation in deep-sequenced small RNA cDNA libraries

RNA-ligase-dependent biases in miRNA representation in deep-sequenced small RNA cDNA libraries

DNA strand exchange reaction activated by cationic comb-type copolymers having ureido groups

DNA assembly and re-assembly activated by cationic comb-type copolymer

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