Nucleic Acids Research, Vol 27, Issue 24 e34-e34, Copyright © 1999 by Oxford University Press
RD Mitra and GM Church
We describe a method to clone and amplify DNA by performing the polymerase
chain reaction (PCR) in a thin polyacrylamide film poured on a glass
microscope slide. The polyacrylamide matrix retards thediffusion of the
linear DNA molecules so that the amplification products remain localized
near their respective templates. At the end of the reaction, a number of
PCR colonies, or `polonies', have formed, each one grown from a single
template molecule. As many as 5 million clones can be amplified in parallel
on a single slide. If an Acrydite modification is included at the 5[prime]
end of one of the primers, the amplified DNA will be covalently attached to
the polyacrylamide matrix, allowing further enzymatic manipulations to be
performed on all clones simul-taneously. We describe techniques to make
replicas of these polony slides, and high throughput sequencing protocols
for this technology. Other applications are also discussed.
ARTICLES
In situ localized amplification and contact replication of many individual DNA molecules
Massachusetts Institute of Technology, Department of Electrical Engineering, Boston, MA 02139, USA; Harvard Medical School, Department of Genetics, Boston, MA 02115, USA
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