[Q] ThermoSequenase aging

Alexander Kraev kraev at bc.biol.ethz.ch
Mon Jun 24 11:59:55 EST 1996


I have used both kits US78500 (radioactive) and RPN2438 (fluorescent),
as well as polymerase separately with homemade termination mixes and
did not notice that it is a particularly unstable enzyme. It is 
definitely more tolerant to clumsiness and general abuse than 
Sequenase V2.0. The problem you describe looks like that of a reduced
polymerase activity, but this reduction may come not only from "ageing",
but also from other, i.e. physical factors, such as an erratic cycler. 
We had two years back a problem with regular PCR, when everybody started 
getting smears instead of bands and it was traced to the cycler being 
offset by 3-5 degrees at denaturation temperature. Did you get the same
problem with the control DNA included?

Alexander Kraev, PhD                     Internet:    
kraev at bc.biol.ethz.ch
> Lab.of Biochemistry III                            Phone:   0041-1-632-31-47
> Swiss Federal Inst. Of Technology           FAX:      0041-1-632-12-13
> Universitaetsstr. 16                    Home Page: http://www.bc.biol.ethz.ch/BiochemistryIII/                                 
> CH-8092 Zurich                                     /Sasha/kraev.html





More information about the Methods mailing list