Hsp92II - ? ligase activity; NlaIII - ?different grades

Ian & Viive Howell ihowell at ozemail.com.au
Fri Sep 5 05:47:54 EST 1997


Does anyone have any experience using the restrction endonuclease
Hsp92II?

I  recently begun using this enzyme to detect a single base change
mutation in a PCR fragment (168bp).  (The mutation creates a
restriction site.)  On electrophoresis of the digested and undigested
fragments, an extra band ~ double the size of the original band always
appears in the digested samples only.  This occurs whenever the enzyme
is added - with or without BSA, with or without inactivation at 65C.
It also occurred with other fragments.  Promega gave me 2 different
batches of Hsp92II - with no change. I have now changed to using the
isoschizomer NlaIII and this phenomenom has disappeared.  Has anyone
else experienced this problem or know why it occurred?

When searching for a replacement enzyme I found 2 different price
ranges for NlaIII:
New England Biolabs (NlaIII) and Promega (Hsp92II) - AUD$82/500 U;
Sigma and Amersham (NlaIII)  ~ AUD $269/ 20 U.
The companies cound not explain the > 1000 fold price difference.  Are
there 2 different grades of this enzyme??

Viive Howell
Clinical Chemistry & Molecular Genetics
ICPMR
Westmead Hospital
NSW, Australia




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