TA Clonning

Matthias Pfeiffer pfeiffer at thermohybaid.de
Mon Sep 10 06:38:10 EST 2001


Hi Peter,
Scott is exacly right: The insert used for cloning with the TOPO-TA should
not be 5´ phosphorylated, instead you need an OH-end. Thus, no primer
phosphorylation is required.
We used the system for routinely cloning our customised synthetic genes.
www.thermohybaid.com/cgi-bin/pg.exe/instrument.html?page_code=Syntgene
In our hands the kit worked out fine, but the amount of insert has to be
estimated with care. The ligation reaction was room temperature. Hope this
helps. Matthias


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