Protocol Online logo
Top : Forum Archives: : Molecular Biology

Colony lift, help!!! - (Jun/07/2005 )


I did a colony lift with nitrocellulosa membranes from my plates. But I need to rehybridize with the same probe and my question is: Do you know if I have to make an stripping before rehybridize? I'm not sure, because I think the nitrocellulosa membranes are very weak and maybe can't resist and stripping.

Any idea?? sad.gif

Thank you!!!



It would be easier to answer if you had specified if you are doing
South-Western or Southern probing. Try Nylon in the future if you are attaching nucleic acid probes, its more versatile for this.
Nitrocellulose can be stripped. Be careful not to tear it, it should withstand the heat of stripping just fine.. Some people strip with 0.5 M NaOH at room temp but some heat may be required. I strip with boiling SDS, but more gentle conditions may be necessary with nitrocellulose. If you have significant background from the first probe, yes, you need to strip.