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Template length-dependent amplification bias in rolling circle amplification

Bastian Joffroy, Yavuz O. Uca, Domen PreŇ°ern, Jonathan .P.K. Doye and Thorsten L. Schmidt

Abstract

DNA amplification is essential for fields ranging from medicine and biology to material sciences and nanotechnology. The polymerase chain reaction (PCR) is the most common amplification method, but is ill-suited for some applications that require high yields, single-stranded DNA or for bioassays and sensors that require bias free, precise quantification. For some of those cases, rolling circle amplification (RCA) can be an attractive alternative. Here we describe an RCA bias that depends on the length of the circular template and exhibits a periodicity corresponding to the helical repeat of B-form DNA. Coarse grained molecular dynamics simulations suggest that different amplification efficiencies come from a varying fraying probability of the last two base pairs of the downstream template. We conclude that an increased strain-promoted fraying probability can increase the polymerization rate compared to a relaxed template.