Droplet-based microfluidic systems for high-throughput single DNA molecule isothermal amplification and analysis.
We have developed a method for high-throughput isothermal amplification of single DNA molecules in a droplet-based microfluidic system. DNA amplification in droplets was analyzed using an intercalating fluorochrome, allowing fast and accurate "digital" quantification of the template DNA based on the Poisson distribution of DNA molecules in droplets. The clonal amplified DNA in each 2 pL droplet was further analyzed by measuring the enzymatic activity of the encoded proteins after fusion with a 15 pL droplet containing an in vitro translation system.
Mazutis
Linas
L
Institut de Science et d'Ingenierie Supramoleculaire, Universite de Strasbourg, CNRS UMR 7006, 8 allee Gaspard Monge, 67083 Strasbourg Cedex, France.
Araghi
Ali Fallah
AF
Miller
Oliver J
OJ
Baret
Jean-Christophe
JC
Frenz
Lucas
L
Janoshazi
Agnes
A
Taly
Valérie
V
Miller
Benjamin J
BJ
Hutchison
J Brian
JB
Link
Darren
D
Griffiths
Andrew D
AD
Ryckelynck
Michael
M
D004247:DNA / Q000032:analysis*; D007364:Intercalating Agents / Q000737:chemistry; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D013696:Temperature
D007364:Intercalating Agents; D004247:DNA