A filtration method for plasmid isolation using microtiter filter plates.
A method for the simultaneous isolation of plasmid DNA from as many as 96 Escherichia coli clones in less than 2 h is described. It is based on a modified version of the alkaline lysis procedure originally described by Birnboim and Doly (Nucleic Acids Res. 7, 1513-1523, 1979). The handling of DNA samples is facilitated by the use of microtiter plates with membrane filter bottoms. All centrifugation steps are replaced by filtrations ("the filtration method"). The yield of plasmid DNA from 0.35 ml of an overnight culture is sufficient for restriction analysis of the plasmid clones. Up to 400 nucleotides' readable sequences could be obtained in cycle sequencing reactions with an unmodified sequencing protocol on an automated ABI sequencer.
Ruppert
A
A
Faculty of Chemistry, Department of Biochemistry and Molecular Biology, University of Hamburg, Germany.
Szalay
B
B
van den Boom
D
D
Horst
G
G
Köster
H
H
D001431:Bacteriological Techniques; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D005374:Filtration / Q000295:instrumentation / Q000379:methods*; D008567:Membranes, Artificial; D008832:Microchemistry; D008969:Molecular Sequence Data; D010957:Plasmids / Q000302:isolation & purification*; D015183:Restriction Mapping; D013462:Sulfur Radioisotopes; D017186:Titrimetry
D008567:Membranes, Artificial; D013462:Sulfur Radioisotopes