[Q] ThermoSequenase aging

Eric Kofoid kofoid at BIOLOGY.UTAH.EDU
Mon Jun 24 11:35:02 EST 1996

I began using Amersham's ThermoSequenase kit in February, and got
excellent results with double stranded PCR templates. After a couple of
months, I began to notice some banding in the sequence ladders, but the
reagents were nearly exhausted so I simply bought a new kit.

Results were good at first but massive banding began occuring after only a
couple of weeks. All reagents from both kits were stored at -20 C or kept
on ice when in use. I use end-labelled primers and have purchased new
label and polynucleotide kinase with no improvement. I have double checked
primer/template pairs which worked well the first time and find heavy
banding with the aged kits.

Before I start the lengthy task of checking each reagent, can anybody
offer any suggestions? Is this a particularly fragile enzyme? Do the
termination mixes age unusually fast? Has anybody else noticed a short
shelf life for this kit?



Eric Kofoid        Dept.Biology, U.Utah       kofoid at biology.utah.edu


Date:   23-Jun-1996
Posted-date: 23-Jun-1996

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