Qiagen midi-prep Kit protocol says :
4ml P1 buffer
4ml P2 buffer
Will it be okay if i added 8ml P1 and 4ml P2 in the experiment?
how is this going to affect the plamid concentration?
Thank you!
If you add too much of the first buffer, you should scale up P2 and N3 accordingly. Once the sample is bound to the columns you can go back to the normal volumes of the wash and elution buffers.
P2 is the lysis buffer, so at a lower concentration you may have gotten incomplete lysis which would hurt your yield.
Hi Supriya,
The P1 basically contains buffer ( Tris usually) for re-suspension of bacterial pallet, EDTA to prevent DNAase from degrading your DNA and RNAase to degrade bacterial RNA.
As such addition of more of this buffer will not have any adverse effect on your plasmid prep.
Hi there,
As pointed out by Chris, this is the concentration of the different reagents (SDS, NaOH in P2 and acetate in N3) after addition that is crucial so proportions between volumes have to be respected whatever the starting volume of P1. If you double P1then you double P2 and N3.
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