Invitrogen platinum pfx enzyme not so fantastic - problems with pcr product gel migration (Aug/10/2004 )
I posted a problem earlier about my pcr product not migrating from the agarose gel well.
I was just informed by invitrogen customer service that:
The problem you have mentioned is one that we have observed before with
Platinum® Pfx DNA Polymerase. Please see the notes below.
Problem: Ethidium bromide-stained DNA in the wells of the agarose gel
amplification with Platinum® Pfx DNA Polymerase.
Solution: This is likely caused by a combination of the following:
* Platinum® Pfx DNA Polymerase has more protein per unit of enzyme than
* Platinum® Pfx DNA Polymerase may have different DNA-binding
enzymes. If the protein remains bound to the DNA, migration is
the gel. If this
reduces the yield of product migrating into the gel, add SDS to the
loading buffer to a final
concentration of 0.1%.
I thought that this might help someone else with this problem. I might suggest using another enzyme if blunt-ended product is needed!
So, it was the protein! I have been quite successful with Stratagene's Pfu. Works great all the time!