15-mer binding to membrane. Is it possible?

Dr. Peter Gegenheimer PGegen at RNAworld.edu
Fri Feb 13 21:51:55 EST 1998


On Fri, 13 Feb 1998 14:47:12, osterop at tch.fgg.eur.nl ("Arthur 
Osterop Ph.D.") wrote:

> Hy netters,
> 
> I want to detect anti-sense oligonucleotides (15 bases long)using Southern 
> blot hybridisation with a labeled sense oligo as probe. Now I am running into 
> the fact that the binding capacity of membranes for small oligo's is very 
> low, and probably the pore width is too large to fetch the oligo's.
> Can anyone advice me on what brand of membrane is suited to use in this 
> application?
> 
> Thanks in advance,
> 
> Arthur Osterop, PhD
> Erasmus Univ. R'dam, NL
> osterop at tch.fgg.eur.nl

Your best bet is to use a positively-charged Nylon membrance, like 
GeneScreen Plus or equivalent, and do the transfer under mildly 
alkaline conditions (>= 50 mM NaOH). We haven't done transfers with 
oligos this small, but I don know that alkaline transfers to 
(+)-Nylon give tightest binding, and that we have electro-blotted 
RNA fragments as small as ~38 nt to (+)-charged Nylon.

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