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KanR sequence


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#1 Rsm

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Posted 07 March 2012 - 05:18 AM

What is the minimal cassette for Kanamycin resistance in bacteria? I have use one from peCFP-N1 (see attached file) from "Bacterial promoter for expression of Kanr gene" until "thymidine kinase polyadenylation signal" (~1.5kb), but it does not seem to confer KanR when I clone it into an AmpR plasmid. Is there something missing, or is it too big?

Any help really appreciated!

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#2 phage434

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Posted 07 March 2012 - 06:23 AM

It's unusual to have a bacterial promoter as far away from the start codon as in this vector. Depending on where you decided to clone, you may have omitted a 5' UP element from the sequence, 5' of the -35 region. In general, I would not be using this vector as a source of the KanR cassette.

#3 Rsm

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Posted 07 March 2012 - 06:42 AM

Thanks phage434. I don't have other vectors for KanR... Well, I have one with AmpR, so I could clone the other way round (AmpR into KanR). But again, what is the minimal cassette for this? If I use the attached vector (pTAL-d2eGFP), then I clone the part from -35 until stop codon? Or do I need some pA sequence, more 5' information etc?

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