[Zbrafish] about designing morpholino

Jon D.Moulton via zbrafish%40net.bio.net (by jmoulton from gene-tools.com)
Tue Jun 10 12:25:30 EST 2008


Prajakta Limaye wrote:
"I want to design morpholino against LIM Domain only 1
(lmo1). Please guide me how to design morpholino. "


Good day Prajakta,

Gene Tools is happy to assist with Morpholino design.  There is no 
charge for oligo design.  You may submit sequence on our website at the 
"Ordering" page, where there is a link titled "Free Oligo Design" or you 
can go directly to the oligo design sequence submission page by 
following this URL:
http://www.gene-tools.com/Oligo_Design
When submitting sequence, please put parentheses around the start codon 
for translation-blocking oligos and write exons in upper case and 
introns in lower case for splice-blocking oligos.

Here are some general guidelines we use for Morpholino oligo design.

Design guidelines
Ideally 25-mer length, can decrease a few bases as needed (shorter 
oligos are better for high CG);
40% to 60% GC
   (lower CG means affinity might be too low to block processes, higher 
favors non-specific binding of subsequences;
   shift toward lower CG for organisms living below 37C);
Up to 36% G (higher G causes loss of water solubility) but avoid the 
upper end of the acceptable range if possible;
Run of 3 G maximum (runs of 4+ G can associate through Hoogsteen binding 
to form tetramers (a.k.a. G quartets));
When possible, avoid a G or A on the 3' end of a modified oligo as these 
may decrease coupling efficiency of the modification;
---------------------------------------------------------------------
Stem loop:
  16 contiguous hydrogen bonds maximum
Self-complementarity:
  16 contiguous hydrogen bonds maximum
Example: AGCGCT has 16 H-bonds (2+3+3+3+3+2 = 16)
Check for non-WatsonCrick G-T pairing, which can participate in 
self-complementarities; we score these as 1 H-bond.
---------------------------------------------------------------------

If you have access to Current Protocols in Nucleic Acid Chemistry, you 
might find this chapter useful.
Moulton JD.  Using morpholinos to control gene expression.  Curr Protoc 
Nucleic Acid Chem. 2007 Jan;Chapter 4:Unit 4.30.

Let me know how I can help.

Regards,

  - Jon

     Jon D. Moulton, Ph.D.
     Diagnostics and Special Projects
     GENE TOOLS, LLC
     jmoulton from gene-tools.com
     www.gene-tools.com
     (541) 929-7840 x1201
     http://network.nature.com/group/morpholinos


prajakta dongre wrote:
> Dear All
>
> I want to design morpholino against LIM Domain only 1
> (lmo1). Please guide me how to design morpholino. 
>
>
> Thanking you
> Yours sincerely
> Prajakta Limaye
> c/o Dr. Upendra Nongthomba
> Molecular reproduction, Development and Genetics
> Indian Institute of Science
> Bangalore - 560012
>
>
>       
>
> _______________________________________________
> Zbrafish mailing list
> Zbrafish from net.bio.net
> http://www.bio.net/biomail/listinfo/zbrafish
>   



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