An efficient PCR mutagenesis strategy without gel purificiation step that is amenable to automation
B Séraphin, S Kandels-Lewis - Nucleic acids research, 1996 - academic.oup.com
B Séraphin, S Kandels-Lewis
Nucleic acids research, 1996•academic.oup.comWe describe here an improved megaprimer PCR mutagenesis strategy. The cumbersome
gel purification step that is usually used can be omitted by appropriately cleaving the first
and second DNA templates with restriction enzymes and enzymatically removing remaining
primers from the first PCR reaction. We show that this improved procedure is reproducible
and highly efficient. Furthermore this method is suitable for automation because all the steps
are now carried out in reaction tubes.
gel purification step that is usually used can be omitted by appropriately cleaving the first
and second DNA templates with restriction enzymes and enzymatically removing remaining
primers from the first PCR reaction. We show that this improved procedure is reproducible
and highly efficient. Furthermore this method is suitable for automation because all the steps
are now carried out in reaction tubes.
Abstract
We describe here an improved megaprimer PCR mutagenesis strategy. The cumbersome gel purification step that is usually used can be omitted by appropriately cleaving the first and second DNA templates with restriction enzymes and enzymatically removing remaining primers from the first PCR reaction. We show that this improved procedure is reproducible and highly efficient. Furthermore this method is suitable for automation because all the steps are now carried out in reaction tubes.
Oxford University Press