High-Speed Plasmid Isolation Using 96-Well, Size-Exclusion Filter Plates
Author(s) -
David Harris,
Marcy Engelstein,
R.M. Parry,
James H. Smith,
Masaharu Mabuchi,
J.C. Leonard
Publication year - 2002
Publication title -
biotechniques
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.617
H-Index - 131
eISSN - 1940-9818
pISSN - 0736-6205
DOI - 10.2144/02323pf02
Subject(s) - chaotropic agent , plasmid , lysis , plasmid preparation , chromatography , alkaline lysis , dna , dna extraction , isolation (microbiology) , biology , cloning (programming) , filtration (mathematics) , microbiology and biotechnology , chemistry , biochemistry , polymerase chain reaction , gene , computer science , dna vaccination , mathematics , pbr322 , programming language , statistics
The accelerating pace of genomics analysis has necessitated the abbreviation of DNA sample preparation protocols. We have developed a size-exclusion-based system for the rapid isolation ofplasmid DNA in a 96-well microplate format. This high-speed protocol employs a modified alkaline lysis methodfor the preparation of the bacterial lysate, followed by three short vacuum filtration steps. Unlike traditional bind/wash/elute methods, there is no need to use chaotropic salts or ethanol. The samples are recovered from the top side of the MultiScreen96 PLASMID plates. Starting with bacterial cell pellets, the entire prycedure for purifying the plasmid DNA can be performed in 30 min with a multichannel pipettor. The high yields, reproducibility, and quality of the plasmids make this system a good choice for any cloning or DNA sequencing operation.
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