PCR in a Rayleigh-Bénard Convection Cell

S. Chandrasekhar, Hydrodynamic and Hydromagnetic Stability (Clarendon, Oxford, 1961).

Müller G., Neumann G., Weber W., J. Cryst. Growth 70, 78 (1984).

A 295-bp segment of the human β-Actin gene was amplified. Forward and reverse primer sequences were 5′-TCACCCACAATGTGCCCATCTACGA-3′ and 5′-CAGCGGAACCGCTCATTGCCAATGG-3′. Reactions contained 10 mM tris-HCl (pH 8.3); 50 mM KCl; 4 mM MgCl₂; 0.2 mM each dATP, dCTP, dGTP, and dUTP; 9 ng/μl human DNA; and 0.1 U/μl of AmpliTaq Polymerase (PE Applied Biosystems). Reactions were run for about 1.5 hours, aspirated from the reaction chambers, stained with SYBR-Green I (final concentration, 200×), and run on a 1% Agarose gel at 110 V for 1 hour.

Lindahl T., Nyberg B., Biochemistry 11, 3610 (1972).

Motion of a dilute aqueous suspension of fluorescent latex microspheres (6-μm diameter; Polysciences) was observed through a fluorescence stereoscope, imaged using an intensified charge-coupled device camera, and recorded to videotape. Averaging the digitized video stream over a time interval of 1 s produced particle paths representative of microsphere trajectories.

This is likely to be an effect of the high temperature at the bottom of the cell (97°C) causing single-stranded scission of the >50 kb template DNA fragments (4). A faint band is also observed at the same migration distance in positive reactions and template-containing negative controls generated in the thermocycler but does not appear in the photographs.

M.K. and V.M.U. contributed equally to the work and are listed alphabetically in the author list. M.K. and V.M.U. co-conceived the idea and executed the flow studies together. V.M.U. carried out image processing and analysis, and M.K. performed the PCR experiments. We acknowledge funding from the NIH (grant P01 HG01984-01).