Researchers made avoidable mistakes when planning their cloning procedures, and records of cloning procedures were incomplete. The 1-base overhangs produced by AhdI may be hard to ligate.As the PI of an academic lab, Ben Glick was frustrated by the huge amount of effort being wasted during cloning. G A C N N N N N G T C C T G N N N N N C A G SmaI can be used at 37☌ for brief incubations.Īfter cleavage, BamHI-HF® (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility.īclI is typically used at 50-55☌, but is 50% active at 37☌.Įfficient cleavage requires at least two copies of the SgrAI recognition sequence. PaeR7I does not recognize the sequence CTCTCGAG.Įfficient cleavage requires at least two copies of the SacII recognition sequence.Ĭleavage may be enhanced when more than one copy of the XmaI recognition sequence is present. Prolonged incubation with NdeI may lead to removal of additional nucleotides. Sticky ends from different SfiI sites may not be compatible.
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