The DOI matches its Crossref record; 26 of the 27 references it cites were found in a registry or an index.
Microdroplet-based PCR enrichment for large-scale targeted sequencing
Open accessAbstract
Targeted enrichment of specific loci of the human genome is a promising approach to enable sequencing-based studies of genetic variation in large populations. Here we describe an enrichment approach based on microdroplet PCR, which enables 1.5 million amplifications in parallel. We sequenced six samples enriched by microdroplet or traditional singleplex PCR using primers targeting 435 exons of 47 genes. Both methods generated similarly high-quality data: 84% of the uniquely mapping reads fell within the targeted sequences; coverage was uniform across approximately 90% of targeted bases; sequence variants were called with >99% accuracy; and reproducibility between samples was high (r(2) = 0.9). We scaled the microdroplet PCR to 3,976 amplicons totaling 1.49 Mb of sequence, sequenced the resulting sample with both Illumina GAII and Roche 454, and obtained data with equally high specificity and sensitivity. Our results demonstrate that microdroplet technology is well suited for processing DNA for massively parallel enrichment of specific subsets of the human genome for targeted sequencing.
Overview
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Integrity signals
Registry
Cites
Statistics
Verification
Claims the paper makes
The claims and statistics of this paper haven’t been checked yet.
Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.
Claims other papers make about this one
Each is checked against this paper’s own text: does it say what the citing paper says it does?
The source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
In the citing paper
“In genome research, an average depth of sequence coverage of 30-fold has been accepted as sufficient for single-nucleotide polymorphism (SNP) and nucleotide insertion or deletion (indel) detection (15–22).”
From Carrier testing for severe childhood recessive diseases by next-generation sequencing (2011) · cites this paper as [16]
Experimental — accuracy not yet measuredThe source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
In the citing paper
“Target capture and next-generation sequencing (NGS) have shown efficacy and, recently, scalability for resequencing human genomes and exomes, providing an alternative potential paradigm for comprehensive carrier testing (15–22).”
From Carrier testing for severe childhood recessive diseases by next-generation sequencing (2011) · cites this paper as [16]
Experimental — accuracy not yet measuredThe source backs part of the claim, not all of it — or backs it for a different population, condition or comparison.
In the citing paper
“Alternatively, microdroplet PCR should theoretically achieve a sensitivity of ~99%, albeit at higher cost (16, 47).”
From Carrier testing for severe childhood recessive diseases by next-generation sequencing (2011) · cites this paper as [16]
Experimental — accuracy not yet measuredThe cited source says what the claim attributes to it.
In the citing paper
“Preliminary experiments suggested that existing protocols for Agilent SureSelect hybrid capture (15) and RainDance microdroplet polymerase chain reaction (PCR) (16) but not Febit HybSelect microarray-based biochip capture (30) or Olink padlock probe ligation and PCR (31) yielded consistent target enrichment.”
From Carrier testing for severe childhood recessive diseases by next-generation sequencing (2011) · cites this paper as [16]
Self-check
Percentages, sums and reported differences re-computed from the paper’s own numbers.
The self-check needs the paper’s full text.
Citations
Cites
Comprehensive genomic characterization defines human glioblastoma genes and core pathways
The Cancer Genome Atlas Research Network (2008) · Nature · doi:10.1038/nature07385 · Cited by 7,735 · cited 1×
“(7) Primer library droplets are dispensed to the microfluidic chip.”
A haplotype map of the human genome
The International HapMap Consortium (2005) · Nature · doi:10.1038/nature04226 · Cited by 5,858
The ENCODE Project Consortium (2007) · Nature · doi:10.1038/nature05874 · Cited by 5,223
Accurate whole human genome sequencing using reversible terminator chemistry
Bentley, D. R., Balasubramanian, S., Swerdlow, H. P., Smith, G. P., Milton, J., Brown, C. G., Hall, K. P., Evers, D. J., Barnes, C. L., Bignell, H. R., et al. (2008) · Nature · doi:10.1038/nature07517 · Cited by 3,684 · cited 1×
“(4) Primer library: primer pair droplets of library elements are mixed together so that each library element has an equal representation.”
The Consensus Coding Sequences of Human Breast and Colorectal Cancers
Sjöblom, T., Jones, S., Wood, L. D., Parsons, D. W., Lin, J., Barber, T. D., Mandelker, D. L., Leary, R. J., Ptak, J., Silliman, N., et al. (2006) · Science · doi:10.1126/science.1133427 · Cited by 3,161
A map of human genome sequence variation containing 1.42 million single nucleotide polymorphisms
The International SNP Map Working Group, Laboratories:, C. S. H., Sachidanandam, R., Weissman, D., Schmidt, S. C., Kakol, J. M., Stein, L. D., National Center for Biotechnology Information:, Marth, G., Sherry, S., et al. (2001) · Nature · doi:10.1038/35057149 · Cited by 2,977 · cited 1×
“intervals that were sequenced in five of the HapMap samples in our study as part of the HapMap Consortiu”
MethodSomatic mutations affect key pathways in lung adenocarcinoma
Ding, L., Getz, G., Wheeler, D. A., Mardis, E. R., McLellan, M. D., Cibulskis, K., Sougnez, C., Greulich, H., Muzny, D. M., Morgan, M. B., et al. (2008) · Nature · doi:10.1038/nature07423 · Cited by 2,681 · cited 1×
“(6) Purified genomic DNA is mixed together with all of the components of the PCR reaction (DNA polymerase, dNTPs and buffer) except for the PCR primers.”
Mapping short DNA sequencing reads and calling variants using mapping quality scores
Li, H., Ruan, J., & Durbin, R. (2008) · Genome Research · doi:10.1101/gr.078212.108 · Cited by 2,638
Formation of dispersions using “flow focusing” in microchannels
Anna, S. L., Bontoux, N., & Stone, H. A. (2003) · Applied Physics Letters · doi:10.1063/1.1537519 · Cited by 2,387
Targeted capture and massively parallel sequencing of 12 human exomes
Ng, S. B., Turner, E. H., Robertson, P. D., Flygare, S. D., Bigham, A. W., Lee, C., Shaffer, T., Wong, M., Bhattacharjee, A., Eichler, E. E., et al. (2009) · Nature · doi:10.1038/nature08250 · Cited by 2,030
Flagged first, then by citations
1 of the 27 references couldn’t be matched to a known paper.
Cited by
Sorted by citations
Sequencing technologies – the next generation
Metzker, M. L. (2010) · Nature Reviews Genetics · doi:10.1038/nrg2626 · Cited by 7,066
Single-cell analysis and sorting using droplet-based microfluidics
Mažutis, L., Gilbert, J., Ung, W. L., Weitz, D. A., Griffiths, A. D., & Heyman, J. A. (2013) · Nature Protocols · doi:10.1038/nprot.2013.046 · Cited by 1,387
Single-cell sequencing-based technologies will revolutionize whole-organism science
Shapiro, E., Biezuner, T., & Linnarsson, S. (2013) · Nature Reviews Genetics · doi:10.1038/nrg3542 · Cited by 1,177
Target-enrichment strategies for next-generation sequencing
Mamanova, L., Coffey, A. J., Scott, C. E., Kozarewa, I., Turner, E. H., Kumar, A., Howard, E., Shendure, J., & Turner, D. (2010) · Nature Methods · doi:10.1038/nmeth.1419 · Cited by 1,258
“The proof of concept for this approach has been published recently 22 .”
Microdroplets in Microfluidics: An Evolving Platform for Discoveries in Chemistry and Biology
Theberge, A. B., Courtois, F., Schaerli, Y., Fischlechner, M., Abell, C., Hollfelder, F., & Huck, W. T. S. (2010) · Angewandte Chemie International Edition · doi:10.1002/anie.200906653 · Cited by 1,059
Anchored multiplex PCR for targeted next-generation sequencing
Zheng, Z., Liebers, M. R., Zhelyazkova, B. H., Cao, Y., Panditi, D., Lynch, K. D., Chen, J., Robinson, H. E., Shim, H. S., Chmielecki, J., et al. (2014) · Nature Medicine · doi:10.1038/nm.3729 · Cited by 913
Variable Clonal Repopulation Dynamics Influence Chemotherapy Response in Colorectal Cancer
Kreso, A., O'Brien, C. A., van Galen, P., Gan, O. I., Notta, F., Brown, A. M. K., Ng, K., Ma, J., Wienholds, E., Dunant, C. F., et al. (2012) · Science · doi:10.1126/science.1227670 · Cited by 755
Carrier testing for severe childhood recessive diseases by next-generation sequencing
Bell, C. J., Dinwiddie, D. L., Miller, N. A., Hateley, S. L., Ganusova, E. E., Mudge, J., Langley, R. J., Zhang, L., Lee, C. C., Schilkey, F. D., et al. (2011) · Science Translational Medicine · doi:10.1126/scitranslmed.3001756 · Cited by 706
“Alternatively, microdroplet PCR should theoretically achieve a sensitivity of ~99%, albeit at higher cost (16, 47).”
BackgroundMethodLibrary construction for next-generation sequencing: Overviews and challenges
Head, S. R., Komori, H. K., LaMere, S. A., Whisenant, T. C., Nieuwerburgh, F. V., Salomon, D. R., & Ordoukhanian, P. T. (2014) · BioTechniques · doi:10.2144/000114133 · Cited by 716
Applications of next-generation sequencing to phylogeography and phylogenetics
McCormack, J. E., Hird, S. M., Zellmer, A. J., Carstens, B. C., & Brumfield, R. T. (2011) · Molecular Phylogenetics and Evolution · doi:10.1016/j.ympev.2011.12.007 · Cited by 666
Multiplexed DNA Sequence Capture of Mitochondrial Genomes Using PCR Products
Maričić, T., Whitten, M., & Pääbo, S. (2010) · PLoS ONE · doi:10.1371/journal.pone.0014004 · Cited by 581
“Other methods rely on micro-droplet-based selection [7].”
Droplet Microfluidics—A Tool for Single‐Cell Analysis
Joensson, H. N., & Svahn, H. A. (2012) · Angewandte Chemie International Edition · doi:10.1002/anie.201200460 · Cited by 563
Quantitative and sensitive detection of rare mutations using droplet-based microfluidics
Pekin, D., Skhiri, Y., Baret, J., Corre, D. L., Mažutis, L., Salem, C. B., Millot, F., Harrak, A. E., Hutchison, J. B., Larson, J. W., et al. (2011) · Lab on a Chip · doi:10.1039/c1lc20128j · Cited by 536
Self-citationGenetic heterogeneity of diffuse large B-cell lymphoma
Zhang, J., Grubor, V. D., Love, C., Banerjee, A., Richards, K. L., Mieczkowski, P. A., Dunphy, C. H., Choi, W., Au, W. Y., Srivastava, G., et al. (2013) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1205299110 · Cited by 543
Studying clonal dynamics in response to cancer therapy using high-complexity barcoding
Bhang, H.-E. C., Ruddy, D. A., Krishnamurthy Radhakrishna, V., Caushi, J. X., Zhao, R., Hims, M. M., Singh, A. P., Kao, I., Rakiec, D., Shaw, P., et al. (2015) · Nature Medicine · doi:10.1038/nm.3841 · Cited by 507
High-throughput injection with microfluidics using picoinjectors
Abate, A., Hung, T., Mary, P., Agresti, J., & Weitz, D. (2010) · Proceedings of the National Academy of Sciences of the United States of America · doi:10.1073/pnas.1006888107 · Cited by 477
Next-generation sequencing for cancer diagnostics: a practical perspective.
Meldrum, C. J., Doyle, M., & Tothill, R. W. (2011) · Europe PMC (PubMed Central) · Cited by 356
Microfluidics for medical diagnostics and biosensors
Rivet, C. A., Lee, H., Hirsch, A., Hamilton, S. K., & Lu, H. (2010) · Chemical Engineering Science · doi:10.1016/j.ces.2010.08.015 · Cited by 372
Droplet microfluidics: recent developments and future applications
Solvas, X. C. I., & deMello, A. J. (2010) · Chemical Communications · doi:10.1039/c0cc02474k · Cited by 361
Targeted capture in evolutionary and ecological genomics
Jones, M. R., & Good, J. M. (2015) · Molecular Ecology · doi:10.1111/mec.13304 · Cited by 370
Next-generation sequencing in the clinic: Promises and challenges
Xuan, J., Yu, Y., Qing, T., Guo, L., & Shi, L. M. (2012) · Cancer Letters · doi:10.1016/j.canlet.2012.11.025 · Cited by 355
“Despite these drawbacks, PCR-based strategies may find wide applications in clinical practice, considering their high specificity that can ensure clinical accuracy.”
MethodInfluentialHiatt, J. B., Pritchard, C. C., Salipante, S. J., O’Roak, B. J., & Shendure, J. (2013) · Genome Research · doi:10.1101/gr.147686.112 · Cited by 335
“These include multiplex hybrid capture (Lipson et al. 2012; Wagle et al. 2012) and highly multiplexed PCR (Tewhey et al. 2009; Harismendy et al. 2011; Forshew et al. 2012).”
InfluentialGenotyping-by-Sequencing in Plants
Deschamps, S., Llaca, V., & May, G. D. (2012) · Biology · doi:10.3390/biology1030460 · Cited by 348
“strategies for genome reduction such as multiplexed amplification of target sequences [105], molecular inversion probes (MIPs) [106] or the use of probes to capture DNA fragments by direct hybridization prior to sequencing [107,108] are available but can be labor intensive and rely heavily on existing sequence informa…”
BackgroundScaling up: A guide to high‐throughput genomic approaches for biodiversity analysis
Porter, T. M., & Hajibabaei, M. (2018) · Molecular Ecology · doi:10.1111/mec.14478 · Cited by 332
“It has been shown that a large portion of sequences, however, map to the universal sequence and targetspecific sequences (Tewhey et al., 2009).”
InfluentialExome sequencing: the sweet spot before whole genomes
Teer, J. K., & Mullikin, J. C. (2010) · Human Molecular Genetics · doi:10.1093/hmg/ddq333 · Cited by 303
“Instead of using plates with hundreds of wells, aqueous microdroplets can segregate thousands of individual reactions in the same tube, allowing for a much more highly parallel use of PCR (22) (commercially available from Raindance).”
BackgroundInfluentialDigital Assays Part I: Partitioning Statistics and Digital PCR
Basu, A. S. (2017) · SLAS TECHNOLOGY · doi:10.1177/2472630317705680 · Cited by 307
“Using a variant of the Raindance system, Tewhey et al.107 amplified nearly 4000 targets simultaneously using a highly multiplexed primer library.”
MethodThe future is now: single-cell genomics of bacteria and archaea
Blainey, P. C. (2013) · FEMS Microbiology Reviews · doi:10.1111/1574-6976.12015 · Cited by 296
“The microdroplets can then be processed en masse for genomic analysis (Tewhey et al., 2009; Zeng et al., 2010).”
Advances in microfluidic PCR for point-of-care infectious disease diagnostics
Park, S., Zhang, Y., Lin, S., Wang, T., & Yang, S. S. (2011) · Biotechnology Advances · doi:10.1016/j.biotechadv.2011.06.017 · Cited by 300
“…amplification and quantification in millions of droplets have been demonstrated for large scale genomic sequencing (Kiss et al. , 2008, Mazutis et al. , 2009a, Tewhey et al. , 2009) and digital detection of pathogenic E. coli O157 cells in a high background of normal K12 cells (Zeng et al. , 2010).”
Shen, F., Du, W., Kreutz, J. E., Fok, A., & Ismagilov, R. F. (2010) · Lab on a Chip · doi:10.1039/c004521g · Cited by 295
Guidelines for DNA taxonomy, with a focus on the meiofauna
Fontaneto, D., Flot, J., & Tang, C. Q. (2015) · Marine Biodiversity · doi:10.1007/s12526-015-0319-7 · Cited by 288
“It is also possible to perform hundreds of PCRs in parallel in a single test tube using microdroplets, and then simultaneously sequence all of the resulting amplicons (Tewhey et al. 2009).”
Development and future of droplet microfluidics
Nan, L., Zhang, H., Weitz, D. A., & Shum, H. C. (2024) · Lab on a Chip · doi:10.1039/d3lc00729d · Cited by 274
Targeted enrichment of genomic DNA regions for next-generation sequencing
Mertes, F., ElSharawy, A., Sauer, S., van Helvoort, J. M. L. M., van der Zaag, P. J., Franke, A., Nilsson, M. F., Lehrach, H., & Brookes, A. J. (2011) · Briefings in Functional Genomics · doi:10.1093/bfgp/elr033 · Cited by 269
Leung, K., Zahn, H., Leaver, T., Konwar, K. M., Hanson, N. W., Pagé, A. P., Lo, C., Chain, P., Hallam, S., & Hansen, C. L. (2012) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1106752109 · Cited by 244
“Two-phase flow systems that manipulate picoliter (pL) volume droplets in closed channels have been shown to be ideally suited to high-speed serial analysis for use in high-throughput screening applications (11) and sample preparation for genomics (12), while the programmable manipulation of nanoliter (nL) volume dropl…”
BackgroundShen, F., Davydova, E. K., Du, W., Kreutz, J. E., Piepenburg, O., & Ismagilov, R. F. (2011) · Analytical Chemistry · doi:10.1021/ac200247e · Cited by 240
Key Principles and Clinical Applications of “Next-Generation” DNA Sequencing
Rizzo, J. M., & Buck, M. J. (2012) · Cancer Prevention Research · doi:10.1158/1940-6207.capr-11-0432 · Cited by 251
Targeted enrichment strategies for next‐generation plant biology
Cronn, R., Knaus, B. J., Liston, A., Maughan, P. J., Parks, M., Syring, J. V., & Udall, J. A. (2012) · American Journal of Botany · doi:10.3732/ajb.1100356 · Cited by 218
Dynamic self-assembly and control of microfluidic particle crystals
Lee, W., Amini, H., Stone, H. A., & Carlo, D. D. (2010) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1010297107 · Cited by 209
“P and programmable control of microscale droplets and particles is poised to facilitate new capabilities in biomedicine (1), materials synthesis (2), and computation (3–5), similar to how robust control of electrons enabled a myriad of applications in computation and communication.”
BackgroundSystematic comparison of three genomic enrichment methods for massively parallel DNA sequencingPreprint
Teer, J. K., Bonnycastle, L. L., Chines, P. S., Hansen, N. F., Aoyama, N., Swift, A. J., Abaan, H. Ö., Albert, T. J., Program, N. C. S., Margulies, E. H., et al. (2010) · Genome Research · doi:10.1101/gr.106716.110 · Cited by 206
Activation of the NOTCH Pathway in Head and Neck Cancer
Sun, W., Gaykalova, D. A., Ochs, M. F., Mambo, E., Arnaoutakis, D., Liu, Y., Loyo, M., Agrawal, N., Howard, J. D., Li, R., et al. (2013) · Cancer Research · doi:10.1158/0008-5472.can-13-1259 · Cited by 206
“The exome sequencing of NOTCH1 was performed by Asuragen (Austin, Texas) (29-31).”
Chip in a lab: microfluidics for next generation life science research
Streets, A. M., & Huang, Y. (2013) · Biomicrofluidics · doi:10.1063/1.4789751 · Cited by 180
Allele-specific DNA methylation: beyond imprinting
Tycko, B. (2010) · Human Molecular Genetics · doi:10.1093/hmg/ddq376 · Cited by 179
Wei, X., Ju, X.-C., Yi, X., Zhu, Q., Qu, N., Liu, T., Chen, Y., Jiang, H., Yang, G., Ruan, Z., et al. (2011) · PLoS ONE · doi:10.1371/journal.pone.0029500 · Cited by 169
“Several sophisticated target-capture methods have been developed to either capture the entire human exome or provide customized capture of regions of interest [7–10], including the Agilent SureSelect target-enrichment system and the Nimblegen solid-phase hybrid array.”
MethodMicrofluidic droplets: new integrated workflows for biological experiments
Kintses, B., van Vliet, L., Devenish, S. R. A., & Hollfelder, F. (2010) · Current Opinion in Chemical Biology · doi:10.1016/j.cbpa.2010.08.013 · Cited by 168
Miniaturization and Parallelization of Biological and Chemical Assays in Microfluidic Devices
Vyawahare, S., Griffiths, A. D., & Merten, C. A. (2010) · Chemistry & Biology · doi:10.1016/j.chembiol.2010.09.007 · Cited by 174
A glimpse into past, present, and future DNA sequencing
Morey, M., Marmiesse, A. F., Castiñeiras, D. E., Fraga, J. M., Couce, M. L., & Cocho, J. A. (2013) · Molecular Genetics and Metabolism · doi:10.1016/j.ymgme.2013.04.024 · Cited by 177
“After thermal cycling, the emulsion is broken down and the products are subjected to library construction [61].”
BackgroundDiagnostics based on nucleic acid sequence variant profiling: PCR, hybridization, and NGS approaches
Khodakov, D. A., Wang, C., & Zhang, D. Y. (2016) · Advanced Drug Delivery Reviews · doi:10.1016/j.addr.2016.04.005 · Cited by 159
The next-generation sequencing technology and application
Zhou, X., Ren, L., Meng, Q., Li, Y., Yu, Y., & Yu, J. M. (2010) · Protein & Cell · doi:10.1007/s13238-010-0065-3 · Cited by 161
“Two methods of target-region resequencing are widely used: PCR-based candidate gene (Dracatos et al., 2009; Goossens et al., 2009; Harismendy and Frazer, 2009; Tewhey et al., 2009) and whole exome approaches (Hodges et al.”
MethodUtilization of next-generation sequencing platforms in plant genomics and genetic variant discovery
Deschamps, S., & Campbell, M. A. (2009) · Molecular Breeding · doi:10.1007/s11032-009-9357-9 · Cited by 153
Self-priming compartmentalization digital LAMP for point-of-care
Zhu, Q., Gao, Y., Yu, B., Ren, H., Qiu, L., Han, S., Jin, W., Jin, Q., & Mu, Y. (2012) · Lab on a Chip · doi:10.1039/c2lc40774d · Cited by 147
Ballew, B. J., Vijai, J., De, S., Sarek, G., Vannier, J., Stracker, T. H., Schrader, K. A., Small, T. N., O'Reilly, R., Manschreck, C., et al. (2013) · PLoS Genetics · doi:10.1371/journal.pgen.1003695 · Cited by 150
Digital PCR on an integrated self-priming compartmentalization chip
Zhu, Q., Qiu, L., Yu, B., Xu, Y., Gao, Y., Pan, T., Tian, Q., Song, Q., Jin, W., Jin, Q., et al. (2014) · Lab on a Chip · doi:10.1039/c3lc51327k · Cited by 144
Ocwieja, K. E., Sherrill-Mix, S. A., Mukherjee, R., Custers-Allen, R., David, P. H., Brown, M., Wang, S., Link, D. R., Olson, J., Travers, K. J., et al. (2012) · Nucleic Acids Research · doi:10.1093/nar/gks753 · Cited by 156
“We also used end point PCR and analysis on ethidium-bromide-stained gels to confirm that the relative ratios of transcripts containing junctions to A3, A4a, A4b and A4c were different in HOS and T cells (Figure 4B).”
InfluentialSelf-citationExisting and Emerging Technologies for Tumor Genomic Profiling
MacConaill, L. E. (2013) · Journal of Clinical Oncology · doi:10.1200/jco.2012.46.5948 · Cited by 144
Multiphase flow in microfluidics: From droplets and bubbles to the encapsulated structures
Sattari, A., Hanafizadeh, P., & Hoorfar, M. (2020) · Advances in Colloid and Interface Science · doi:10.1016/j.cis.2020.102208 · Cited by 141
Miniaturization of biological assays — Overview on microwell devices for single-cell analyses
Lindström, S., & Svahn, H. A. (2010) · Biochimica et Biophysica Acta (BBA) - General Subjects · doi:10.1016/j.bbagen.2010.04.009 · Cited by 141
The PhyloPythiaS Web Server for Taxonomic Assignment of Metagenome Sequences
Patil, K. R., Roune, L., & McHardy, A. C. (2012) · PLoS ONE · doi:10.1371/journal.pone.0038581 · Cited by 133
“In the current study, we analyzed two datasets, one sequenced with Sanger and another with Illumina sequencing technology [21].”
UroMark—a urinary biomarker assay for the detection of bladder cancer
Feber, A., Dhami, P., Dong, L., de Winter, P., Tan, W. S., Martínez‐Fernández, M., Paul, D. S., Hynes-Allen, A., Rezaee, S., Gurung, P. M. S., et al. (2017) · Clinical Epigenetics · doi:10.1186/s13148-016-0303-5 · Cited by 131
“For example, the microdroplet-based PCR amplification of bisulfite-converted DNA followed by NGS of the amplified target loci (termed RainDrop BS-Seq) enables the sensitive, specific and simultaneous amplification of up to 4000 bisulfite-converted target loci [26].”
Voelkerding, K. V., Dames, S. A., & Durtschi, J. (2010) · Journal of Molecular Diagnostics · doi:10.2353/jmoldx.2010.100043 · Cited by 137
A ferrobotic system for automated microfluidic logistics
Yu, W., Lin, H., Wang, Y., He, X., Chen, N., Sun, K., Lo, D., Cheng, B., Yeung, C. J., Tan, J., et al. (2020) · Science Robotics · doi:10.1126/scirobotics.aba4411 · Cited by 129
Sequence-based marker development in wheat: Advances and applications to breeding
Paux, E., Sourdille, P., Mackay, I., & Feuillet, C. (2011) · Biotechnology Advances · doi:10.1016/j.biotechadv.2011.09.015 · Cited by 122
Liao, G. J. W., Lun, F. M. F., Zheng, Y. W. L., Chan, K. C. A., Leung, T. Y., Lau, T. K., Chiu, R. W. K., & Lo, Y. D. (2010) · Clinical Chemistry · doi:10.1373/clinchem.2010.154336 · Cited by 120
Nanoliter Multiplex PCR Arrays on a SlipChip
Shen, F., Du, W., Davydova, E. K., Karymov, M. A., Pandey, J., & Ismagilov, R. F. (2010) · Analytical Chemistry · doi:10.1021/ac1007249 · Cited by 115
The physics of 2D microfluidic droplet ensembles
Beatus, T., Bar‐Ziv, R., & Tlusty, T. (2012) · Physics Reports · doi:10.1016/j.physrep.2012.02.003 · Cited by 111
“…embryos (Funfak et al., 2007), PCR reactions of a single template molecule (Beer et al., 2007, 2008; Kiss et al., 2008; Schaerli et al., 2008; Tewhey et al., 2009), scanning of protein crystallization conditions (Lau et al., 2007; Leng and Salmon, 2009; Shim et al., 2007; Zheng et al., 2005,…”
BackgroundDetecting biomarkers with microdroplet technology
Taly, V., Pekin, D., Abed, A. I. E., & Laurent‐Puig, P. (2012) · Trends in Molecular Medicine · doi:10.1016/j.molmed.2012.05.001 · Cited by 117
Gasc, C., Peyretaillade, É., & Peyret, P. (2016) · Nucleic Acids Research · doi:10.1093/nar/gkw309 · Cited by 119
Moon, B., Jones, S. G., Hwang, D. K., & Tsai, S. S. H. (2015) · Lab on a Chip · doi:10.1039/c5lc00217f · Cited by 113
Perkins, G., Lu, H., Garlan, F., & Taly, V. (2016) · Advances in clinical chemistry · doi:10.1016/bs.acc.2016.10.001 · Cited by 114
Microfluidic platform for combinatorial synthesis in picolitre droplets
Theberge, A. B., Mayot, E., Harrak, A. E., Kleinschmidt, F., Huck, W. T. S., & Griffiths, A. D. (2012) · Lab on a Chip · doi:10.1039/c2lc21019c · Cited by 111
Lin, X., Tang, W., Ahmad, S., Lü, J., Colby, C. C., Zhu, J., & Yu, Q. (2012) · Hearing Research · doi:10.1016/j.heares.2012.01.004 · Cited by 115
Clinical application of amplicon-based next-generation sequencing in cancer
Chang, F., & Li, M. M. (2013) · Cancer Genetics · doi:10.1016/j.cancergen.2013.10.003 · Cited by 109
“The droplet PCR reactions are then destabilized and the PCR products are released and purified for sequencing (21).”
BackgroundOverview of Target Enrichment Strategies
Kozarewa, I., Armisen, J., Gardner, A. F., Slatko, B. E., & Hendrickson, C. L. (2015) · Current Protocols in Molecular Biology · doi:10.1002/0471142727.mb0721s112 · Cited by 105
Medlin, L. K., & Orozco, J. (2017) · Sensors · doi:10.3390/s17051184 · Cited by 104
“Field-induced coalescence of these droplet pairs results in the two droplets merging into a single PCR droplet, which is collected and processed as an emulsion PCR reaction [99].”
MethodAdvances in microfluidics in combating infectious diseases
Tay, A. K. P., Pavesi, A., Yazdi, S. R., Lim, C. T., & Warkiani, M. E. (2016) · Biotechnology Advances · doi:10.1016/j.biotechadv.2016.02.002 · Cited by 103
“Currently, droplet microfluidic PCR is also being tested (Tewhey et al., 2009).”
Droplet-Based Microfluidics: Enabling Impact on Drug Discovery
Dressler, O. J., Maceiczyk, R. M., Chang, S., & deMello, A. J. (2013) · SLAS DISCOVERY · doi:10.1177/1087057113510401 · Cited by 98
Detection of low prevalence somatic mutations in solid tumors with ultra-deep targeted sequencing
Harismendy, O., Schwab, R. B., Bao, L., Olson, J., Rozenzhak, S., Kotsopoulos, S. K., Pond, S. J. K., Crain, B., Chee, M. S., Messer, K. S., et al. (2011) · Genome biology · doi:10.1186/gb-2011-12-12-r124 · Cited by 96
“The RDT1000 generates each PCR droplet by merging a single DNA template droplet with a single primer droplet [13].”
MethodResultInfluentialSelf-citationA novel three-round multiplex PCR for SNP genotyping with next generation sequencing
Chen, K., Zhou, Y., Li, K., Qi, L.-X., Zhang, Q., Wang, M., & Xiao, J. (2016) · Analytical and Bioanalytical Chemistry · doi:10.1007/s00216-016-9536-6 · Cited by 97
Under-three minute PCR: Probing the limits of fast amplification
Wheeler, E. K., Hara, C. A., Frank, J., DeOtte, J. R., Hall, S., Benett, W. J., Spadaccini, C. M., & Beer, N. R. (2011) · The Analyst · doi:10.1039/c1an15365j · Cited by 97
Target Enrichment Approaches for Next-Generation Sequencing Applications in Oncology
Singh, R. R. (2022) · Diagnostics · doi:10.3390/diagnostics12071539 · Cited by 100
“They involve varied approaches, such as droplet microfluidics, where the PCR amplification, barcoding, and library preparation occur in microscopic droplets, which act as microreactors [18,43], and multilayered soft lithographics microfluidics systems, which use microfluidics in a cartridge to miniaturize and conduct…”
Zargar, S. M., Raatz, B., Sonah, H., MuslimaNazir, Bhat, J. A., Dar, Z. A., Agrawal, G. K., & Rakwal, R. (2015) · Journal of Crop Science and Biotechnology · doi:10.1007/s12892-015-0037-5 · Cited by 101
“Tewhey and co-workers in 2009 reported sequencing PCR amplicons from primers targeting 435 exons of 47 genes in six samples simultaneously using a microdroplet PCR platform from RainDance Technologies (Lexington, USA) (Tewhey et al. 2009).”
MethodComparison of Three Targeted Enrichment Strategies on the SOLiD Sequencing Platform
Hedges, D. J., Guettouche, T., Yang, S., Bademci, G., Diaz, A., Andersen, A., Hulme, W. F., Linker, S. B., Mehta, A., Edwards, Y. J. K., et al. (2011) · PLoS ONE · doi:10.1371/journal.pone.0018595 · Cited by 94
“Genomic enrichment via massively parallel PCR was conducted at Raindance Technologies, as previously described [6].”
MethodReview of massively parallel DNA sequencing technologies
Moorthie, S. A., Mattocks, C., & Wright, C. F. (2011) · The HUGO Journal · doi:10.1007/s11568-011-9156-3 · Cited by 101
“…from RainDance Technologies which uses an emPCR approach to simultaneously amplify up to 4,000 short DNA sequences in separate microdroplets (Tewhey et al. 2009), and the Access Array from Fluidigm which uses proprietary microfluidics to setup an array of 2,304 PCRs (48 samples 9 48 assays).”
Droplet-based microfluidics at the femtolitre scale
Leman, M., Abouakil, F., Griffiths, A. D., & Tabeling, P. (2014) · Lab on a Chip · doi:10.1039/c4lc01122h · Cited by 99
Trans genomic capture and sequencing of primate exomes reveals new targets of positive selectionPreprint
George, R. D., McVicker, G., Diederich, R., Ng, S. B. H., MacKenzie, A. P., Swanson, W. J., Shendure, J., & Thomas, J. H. (2011) · Genome Research · doi:10.1101/gr.121327.111 · Cited by 89
“New methods for targeted enrichment of the human exome allow high-coverage sequence to be generated for the coding fraction of the genome (Albert et al. 2007; Gnirke et al. 2009; Tewhey et al. 2009; Turner et al. 2009).”
Jones, M. A., Bhide, S., Chin, E., Ng, B. G., Rhodenizer, D., Zhang, V. W., Sun, J., Tanner, A. K., Freeze, H. H., & Hegde, M. (2011) · Genetics in Medicine · doi:10.1097/gim.0b013e318226fbf2 · Cited by 91
Cryptic species in plant-parasitic nematodes
Palomares‐Rius, J. E., Cantalapiedra‐Navarrete, C., & Castillo, P. (2014) · Nematology · doi:10.1163/15685411-00002831 · Cited by 90
Analyzing the cancer methylome through targeted bisulfite sequencing
Lee, E.-J., Luo, J., Wilson, J. M., & Shi, H. (2012) · Cancer Letters · doi:10.1016/j.canlet.2012.10.040 · Cited by 89
“PCR amplification bias is significantly decreased due to the nature of microdroplet emulsion PCR reactions [59].”
Shen, X., Liang, D., Feng, Y., Chen, M. Y., & Zhang, P. (2013) · Molecular Biology and Evolution · doi:10.1093/molbev/mst122 · Cited by 88
Höijer, I., Tsai, Y., Clark, T. A., Kotturi, P., Dahl, N., Stattin, E., Bondeson, M.-L., Feuk, L., Gyllensten, U. B., & Ameur, A. (2018) · Human Mutation · doi:10.1002/humu.23580 · Cited by 91
Novel Insight into Mutational Landscape of Head and Neck Squamous Cell Carcinoma
Gaykalova, D. A., Mambo, E., Choudhary, A. K., Houghton, J., Buddavarapu, K., Sanford, T., Darden, W., Adai, A. T., Hadd, A. G., Latham, G. J., et al. (2014) · PLoS ONE · doi:10.1371/journal.pone.0093102 · Cited by 92
“Somatic mutation calling The sequence read data generated from the GAIIx were demultiplexed, adapter and primer sequences were removed and trimmed to retain high quality data (q20 or higher) [37,39,40].”
MethodRapid and continuous magnetic separation in droplet microfluidic devices
Brouzes, E. R., Kruse, T., Kimmerling, R. J., & Strey, H. H. (2014) · Lab on a Chip · doi:10.1039/c4lc01327a · Cited by 87
From tubes to drops: droplet-based microfluidics for ultrahigh-throughput biology
Tran, T. M., Lan, F., Thompson, C. S., & Abate, A. R. (2013) · Journal of Physics D Applied Physics · doi:10.1088/0022-3727/46/11/114004 · Cited by 86
Advances in clinical next-generation sequencing: target enrichment and sequencing technologies
Ballester, L. Y., Luthra, R., Kanagal‐Shamanna, R., & Singh, R. R. (2015) · Expert Review of Molecular Diagnostics · doi:10.1586/14737159.2016.1133298 · Cited by 86
Bernasconi-Elias, P., Hu, T., Jenkins, D., Firestone, B., Gans, S., Kurth, E., Capodieci, P., Deplazes-Lauber, J., Petropoulos, K., Thiel, P., et al. (2016) · Oncogene · doi:10.1038/onc.2016.133 · Cited by 87
“Therefore, Sanger Sequencing (Genewiz), RainDance analysis (28) and RNAseq (29) were performed to determine whether any of these cell lines had NOTCH3 mutations.”
Hopp, K., Heyer, C. M., Sieben, C. J., Henke, S. A., Sundsbak, J. L., Patel, S., Patel, P., Consugar, M. B., Czarnecki, P. G., Gliem, T. J., et al. (2011) · Human Molecular Genetics · doi:10.1093/hmg/ddr151 · Cited by 85
Zellmer, A. J., Hanes, M. M., Hird, S. M., & Carstens, B. C. (2012) · Systematic Biology · doi:10.1093/sysbio/sys048 · Cited by 81
“…for systems where high levels of paralogy are expected, other methods of library construction may be preferable, such as PCR amplicon sequencing (e.g., Binladen et al. 2007; Meyer et al. 2008; Tewhey et al. 2009) or target enrichment (e.g., Albert et al. 2007; Okou et al. 2007; Gnirke et al. 2009).”
Smith, C. A., Li, X., Mize, T. H., Sharpe, T., Graziani, E. I., Abell, C., & Huck, W. T. S. (2013) · Analytical Chemistry · doi:10.1021/ac400453t · Cited by 79
Fluorogenic DNA sequencing in PDMS microreactors
Sims, P. A., Greenleaf, W. J., Duan, H., & Xie, X. S. (2011) · Nature Methods · doi:10.1038/nmeth.1629 · Cited by 79
Targeted resequencing of candidate genes using selector probes
Johansson, H., Isaksson, M., Falk-Sörqvist, E., Roos, F., Stenberg, J., Sjöblom, T., Botling, J., Micke, P., Edlund, K., Fredriksson, S., et al. (2010) · Nucleic Acids Research · doi:10.1093/nar/gkq1005 · Cited by 76
A scalable self-priming fractal branching microchannel net chip for digital PCR
Zhu, Q., Xu, Y., Qiu, L., Ma, C., Yu, B., Song, Q., Jin, W., Jin, Q., Liu, J., & Mu, Y. (2017) · Lab on a Chip · doi:10.1039/c7lc00267j · Cited by 75
Scheible, M. K. R., Loreille, O. M., Just, R. S., & Irwin, J. A. (2014) · Forensic Science International Genetics · doi:10.1016/j.fsigen.2014.04.010 · Cited by 75
Metadata
- Authors
- Tewhey, R., Warner, J. B., Nakano, M., Libby, B., Medkova, M., David, P. H., Kotsopoulos, S. K., Samuels, M. L., Hutchison, J. B., Larson, J. W., Topol, E. J., Weiner, M. P., Harismendy, O., Olson, J., Link, D. R., & Frazer, K. A.
- Affiliations
- Scripps Research Institute (US)
- Massachusetts Institute of Technology (US)
- University of California San Diego (US)
- Corresponding author
- Link, Darren R
- Frazer, Kelly A
- ORCID
- Venue
- Nature Biotechnology
- Published
- 2009-11
- Volume
- 27
- Issue
- 11
- Pages
- 1025-1031
- ISSN
- 0733-222X (ISSN-L)
- 1087-0156
- 1546-1696
- PMC
- PMC2779736
- OpenAlex
- W2164112721
- Cited by
- 454
- References
- 27
- Citations by year
- 2026: 3 · 2025: 9 · 2024: 14 · 2023: 18 · 2022: 13 · 2021: 13 · 2020: 11 · 2019: 13 · 2018: 20 · 2017: 34 · 2016: 25 · 2015: 31 · 2014: 42 · 2013: 49 · 2012: 58
- FWCI
- 31.84
- Type
- journal article
- Language
- en
- Update notices
- erratum (2010-02-01)
- Open access
- green
- www.ncbi.nlm.nih.gov
- Locations
- Nature Biotechnology (publishedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- eScholarship (California Digital Library) (submittedVersion)
- Funders
- National Institutes of Health (1r21ca125693-01)
- National Heart, Lung, and Blood Institute (P50HL077107)
- National Cancer Institute (1R21CA125693-01, R21 CA125693)
- National Center for Research Resources (1U54RR025204-01, UL1RR025774)
- Topics
- Physical Sciences › Chemical Engineering › Fluid Flow and Transfer Processes › Innovative Microfluidic and Catalytic Techniques Innovation
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Phylogenetic Studies
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Genomics and Rare Diseases
- Keywords
- target enrichment, microfluidics, targeted sequencing, exome sequencing, variant calling, human genome
- MeSH terms
- Base Sequence, Humans, Mutation, Reproducibility of Results, Polymerase Chain Reaction, Sequence Analysis, DNA, Microfluidics
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
Are you an author, or do you see a problem with this page? Request a review
