TY - JOUR
T1 - Liquid drop of DNA libraries reveals total genome information
AU - Terekhov, Stanislav S.
AU - Eliseev, Igor E.
AU - Ovchinnikova, Leyla A.
AU - Kabilov, Marsel R.
AU - Prjibelski, Andrey D.
AU - Tupikin, Alexey E.
AU - Smirnov, Ivan V.
AU - Belogurov, Alexey A.
AU - Severinov, Konstantin V.
AU - Lomakin, Yakov A.
AU - Altman, Sidney
AU - Gabibov, Alexander G.
N1 - Funding Information:
ACKNOWLEDGMENTS. We thank Mr. Gregory Leleytner for his contribution to the computational design of the DNA libraries. We also thank St. Petersburg University for providing computational resources. This study was supported by Russian Science Foundation Grant 17-74-30019; Y.A.L. received personal financial support from The Russian Foundation for Basic Research Grant НР 17-04-01233 A; and Grant 18-29-08054 (to S.S.T. and I.V.S.).
Publisher Copyright:
© 2020 National Academy of Sciences. All rights reserved.
PY - 2020/11/3
Y1 - 2020/11/3
N2 - Conventional “bulk” PCR often yields inefficient and nonuniform amplification of complex templates in DNA libraries, introducing unwanted biases. Amplification of single DNA molecules encapsulated in a myriad of emulsion droplets (emulsion PCR, ePCR) allows the mitigation of this problem. Different ePCR regimes were experimentally analyzed to identify the most robust techniques for enhanced amplification of DNA libraries. A phenomenological mathematical model that forms an essential basis for optimal use of ePCR for library amplification was developed. A detailed description by high-throughput sequencing of amplified DNA-encoded libraries highlights the principal advantages of ePCR over bulk PCR. ePCR outperforms PCR, reduces gross DNA errors, and provides a more uniform distribution of the amplified sequences. The quasi single-molecule amplification achieved via ePCR represents the fundamental requirement in case of complex DNA templates being prone to diversity degeneration and provides a way to preserve the quality of DNA libraries.
AB - Conventional “bulk” PCR often yields inefficient and nonuniform amplification of complex templates in DNA libraries, introducing unwanted biases. Amplification of single DNA molecules encapsulated in a myriad of emulsion droplets (emulsion PCR, ePCR) allows the mitigation of this problem. Different ePCR regimes were experimentally analyzed to identify the most robust techniques for enhanced amplification of DNA libraries. A phenomenological mathematical model that forms an essential basis for optimal use of ePCR for library amplification was developed. A detailed description by high-throughput sequencing of amplified DNA-encoded libraries highlights the principal advantages of ePCR over bulk PCR. ePCR outperforms PCR, reduces gross DNA errors, and provides a more uniform distribution of the amplified sequences. The quasi single-molecule amplification achieved via ePCR represents the fundamental requirement in case of complex DNA templates being prone to diversity degeneration and provides a way to preserve the quality of DNA libraries.
KW - Diversity degeneration
KW - Emulsion PCR modeling
KW - Quasi single-molecule amplification
KW - Template mispairing
KW - Uniform distribution of amplicons
UR - http://www.scopus.com/inward/record.url?scp=85095668840&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85095668840&partnerID=8YFLogxK
U2 - 10.1073/pnas.2017138117
DO - 10.1073/pnas.2017138117
M3 - Article
C2 - 33087570
AN - SCOPUS:85095668840
SN - 0027-8424
VL - 117
SP - 27300
EP - 27306
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 44
ER -