DropSynth 2.0: high-fidelity multiplexed gene synthesis in emulsions
作者： Angus M Sidore , Calin Plesa , Joyce A Samson , Nathan B Lubock , Sriram Kosuri
DOI: 10.1093/NAR/GKAA600
关键词: Extramural 、 Biological system 、 Oligonucleotide 、 Gene synthesis 、 Multiplexing 、 Fidelity 、 Genomic library 、 Biology 、 High fidelity
摘要: Multiplexed assays allow functional testing of large synthetic libraries genetic elements, but are limited by the designability, length, fidelity and scale input DNA. Here, we improve DropSynth, a low-cost, multiplexed method that builds gene compartmentalizing assembling microarray-derived oligonucleotides in vortexed emulsions. By optimizing enzyme choice, adding enzymatic error correction increasing scale, show DropSynth can build thousands gene-length fragments at >20% fidelity.
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sci-hub.st 参考文章(29)
Nikolai Eroshenko, Sriram Kosuri, Adam H. Marblestone, Nicholas Conway, George M. Church, Gene Assembly from Chip-Synthesized Oligonucleotides. Current protocols in chemical biology. ,vol. 4, pp. 1- 17 ,(2012) , 10.1002/9780470559277.CH110190
Dmitry A Shagin, Konstantin A Lukyanov, Laura L Vagner, Mikhail V Matz, Regulation of average length of complex PCR product. Nucleic Acids Research. ,vol. 27, ,(1999) , 10.1093/NAR/27.18.E23
Jerrod J Schwartz, Choli Lee, Jay Shendure, Accurate gene synthesis with tag-directed retrieval of sequence-verified DNA molecules Nature Methods. ,vol. 9, pp. 913- 915 ,(2012) , 10.1038/NMETH.2137
J. Smith, P. Modrich, Removal of polymerase-produced mutant sequences from PCR products Proceedings of the National Academy of Sciences of the United States of America. ,vol. 94, pp. 6847- 6850 ,(1997) , 10.1073/PNAS.94.13.6847
Alex Y. Borovkov, Andrey V. Loskutov, Mark D. Robida, Kristen M. Day, Jose A. Cano, Tien Le Olson, Hetal Patel, Kevin Brown, Preston D. Hunter, Kathryn F. Sykes, High-quality gene assembly directly from unpurified mixtures of microarray-synthesized oligonucleotides Nucleic Acids Research. ,vol. 38, ,(2010) , 10.1093/NAR/GKQ677
R. T. Hietpas, J. D. Jensen, D. N. A. Bolon, Experimental illumination of a fitness landscape. Proceedings of the National Academy of Sciences of the United States of America. ,vol. 108, pp. 7896- 7901 ,(2011) , 10.1073/PNAS.1016024108
Hwangbeom Kim, Hyojun Han, Jinwoo Ahn, Joongoo Lee, Namjin Cho, Hoon Jang, Hyoki Kim, Sunghoon Kwon, Duhee Bang, ‘Shotgun DNA synthesis’ for the high-throughput construction of large DNA molecules Nucleic Acids Research. ,vol. 40, ,(2012) , 10.1093/NAR/GKS546
Timothy H.-C. Hsiau, David Sukovich, Phillip Elms, Robin N. Prince, Tobias Stritmatter, Paul Ruan, Bo Curry, Paige Anderson, Jeff Sampson, J. Christopher Anderson, A method for multiplex gene synthesis employing error correction based on expression. PLOS ONE. ,vol. 10, ,(2015) , 10.1371/JOURNAL.PONE.0119927
Tilo Buschmann, Leonid V Bystrykh, Levenshtein error-correcting barcodes for multiplexed DNA sequencing. BMC Bioinformatics. ,vol. 14, pp. 272- 272 ,(2013) , 10.1186/1471-2105-14-272
Cosmas D Arnold, Daniel Gerlach, Christoph Stelzer, Łukasz M Boryń, Martina Rath, Alexander Stark, None, Genome-wide quantitative enhancer activity maps identified by STARR-seq. Science. ,vol. 339, pp. 1074- 1077 ,(2013) , 10.1126/SCIENCE.1232542