plasmid isolation for sequensing
Christos Nitsos
Wednesday, 07 October 2009 09:36 UTC
I have a gene inserted in a pET29 vector and I want to send it for sequensing. The problem is that the laboaratory where I ll send the sample requires isolation by spin column and I dont have a coulumn isolation kit in the lab. In our lab we use the typical alkaline lysis protocol for plasmid DNA isolation. However I do have a total DNA isolation kit from Quiagen (Quiamp DNA Mini kit using proteinase K for cell lysis). Does anyone know if i can combine the two protocols? i.e. perform an alkaline lysis and then purify the lysate through the quiagen DNA column, instead of the ethanol precipitation and washing steps? is the chemistry compatible? If anyone knows this it would be a great help
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Replies
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I would stick to a manual procedure for plasmid dna isolation, foreinstance one of those suggested maniatis and sambrook. prior to send the sample for sequencing, try some digestions to check the dna purity.
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