Recombinase polymerase amplification (RPA) is an isothermal DNA amplification method with broad applications as a point-of-care test and in molecular biology techniques. Currently, most of the applications are focused on target-specific amplification. Because RPA has the advantage of amplifying DNA under isothermal conditions, we utilized RPA as a DNA library amplification tool. In this study, we used a sheared genomic DNA library and an oligonucleotide (oligo) library for the comparison of polymerase chain reaction and RPA. For the sheared DNA library, we observed biased amplification after RPA was conducted. Thus, to amplify the size-variable DNA library uniformly, we introduced a linear amplification strategy with RPA and successfully improved the uniformity. On the other hand, using the same-sized oligo library, we confirmed that RPA amplified this library uniformly without modification of the protocol. These results demonstrate that RPA can be applied not only to amplify a specific target as previously demonstrated but also to amplify a complex DNA library composed of a large number of different DNA molecules.
Bibliographical noteFunding Information:
J.L. and D.B. conceived the project. J.L. performed all experiments. J.L. and S.H. analyzed the sequencing data. J.L. wrote the manuscript. D.B. supervised the research. This work was supported by (i) the Mid-career Researcher Program (NRF-2018R1A2A1A05079172) from the National Research Foundation of Korea (NRF), funded by the Ministry of Science, ICT & Planning, (ii) the Bio & Medical Technology Development Program of the NRF, funded by the Korean government (MSIT; NRF-2016M3A9B6948494), and (iii) the Bio & Medical Technology Development Program of the NRF, funded by the Korean government (MSIT; NRF-2018M3A9H3024850), and (iv) the Korea Health Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (HI18C2282). The authors declare no competing financial interest.
© 2019 American Chemical Society.
All Science Journal Classification (ASJC) codes
- Chemical Engineering(all)