Methods for isolating nucleic acids using alkaline protease

Details Methods for isolating nucleic acids using alkaline protease Methods for isolating nucleic acids using alkaline protease

1996-07-29

1999-11-09

Naff, David M.

Chemistry: molecular biology and microbiology

Micro-organism, tissue cell culture or enzyme using process...

Preparing compound containing saccharide radical

435184, 435219, C12P 1934, C12N 999, C12N 950

Patent

active

059812354

ABSTRACT:
Solutions containing nucleic acids are treated with an alkaline protease to digest proteins such as nucleases that degrade the nucleic acids. In the isolation of nucleic acids, a biological sample containing nucleic acids is suspended in a solution containing water, buffer and chelating agent, the pH of the solution is adjusted to at least about 10 by adding a solution of sodium hydroxide and anionic detergent, an alkaline protease is incubated in the solution until nucleases are degraded, the pH of the solution is lowered to reduce activity of the alkaline protease by adding a solution having a pH between 3.5 and 4.5 and the alkaline protease is heat inactivated. Lowering of the pH may produce a cloudy solution which is cleared by centrifuging. Nucleic acids are isolated from the cleared solution by alcohol precipitation, or by using paramagnetic particles or a resin matrix containing silica particles. A chaotropic salt can be used to reversibly bind DNA to the resin matrix.

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