[Zbrafish] morpholino blues

Gilbert Weidinger via zbrafish%40net.bio.net (by gilbert.weidinger from biotec.tu-dresden.de)
Fri May 23 10:54:13 EST 2008


hi all,

I wonder whether it's conceivable that some genes cannot be targeted 
with morpholinos. In many years of using MOs I have come to accept that 
only about 50% of them work, but now we have a case in the lab where 
NONE out of 4 MOs targeting the same gene has any effect.

We have 2 splice blockers: they do not alter splicing as determined by 
RT-PCR.
We have 2 translation blockers: they do not inhibit translation of RNA 
reporter constructs (5'UTR fused to cherry).

We inject up to 10µg/µl and use both the RT-PCR assay and the reporter 
RNAs successfully for other MOs (against other genes). Genetools claims 
that 70% of all MOs knock down their target...

Of note is that all 4 MOs are tagged with fluorescein. We don't have a 
lot of experience with these. Is it possible that they are less 
efficient than untagged MOs?

I'd appreciate getting some feedback on whether there are other stories 
like this out there or whether we are simply exceptionally unlucky.

thanks
gilbert

-----------------------------------------------------------
Gilbert Weidinger, PhD

Group Leader SFB 655
Biotechnology Center & Center for Regenerative Therapies
Technical University of Dresden

Mail address:

Biotechnology Center
Tatzberg 47-51
01307 Dresden
Germany

Tel. office:    0049 351 463 40120
Tel. lab:       0049 351 463 40108
Tel. secretary: 0049 351 463 40345
Fax:            0049 351 463 40348
email:          gilbert.weidinger from biotec.tu-dresden.de
web:		http://www.biotec.tu-dresden.de/weidinger



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