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Using dose synergy to demonstrate specificity

Using dose synergy to demonstrate specificity - RNA Techniques Forum

Using dose synergy to demonstrate specificity - Post and discuss RNA methods and RNA science related topics. RNA extraction, cDNA synthesis, RNA EMSA, and other RNA protocols.


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Old 05-05-2009, 04:27 PM
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Default Using dose synergy to demonstrate specificity



Brent Bill has a new paper in the journal Zebrafish which tells us how to use dose synergy to demonstrate specificity of a gene knockdown. In the usual two-nonoverlapping-Morpholino experiment, two oligos targeting the same RNA are used in separate experiments. If both of the oligos elicit the same phenotypic change, that is good support for the hypothesis that the change is due to knockdown of the targeted oligo (and not an interaction with an off-target RNA). The new paper proposes using dose synergy as a test for specificity. Again two non-overlapping oligos are used, but for this experiment they are co-injected so that each oligo is at a concentration considerably less than half of that normally needed to elicit a phenotype if the oligo is injected individually. If coinjection alters the phenotype though dose synergy, that supports the hypothesis that the knockdown is specific for the target.

For each new oligo sequence, the dose needed to elicit a phenotype must be determined experimentally. Often different oligos will require different doses to cause a knockdown, even if the oligos are targeting the same RNA. Since the oligo sequences are different, their RNA affinities are probably different. Secondary structure in the RNA might make invasion of duplexed regions slower for one oligo than for the other. However, copying the phenotype and showing dose synergy are together good methods for demonstrating specificity of oligos even if they are used at different doses.


Here is Brent Bill's paper:

Bill BR, Petzold AM, Clark KJ, Schimmenti LA, Ekker SC. A primer for morpholino use in zebrafish. Zebrafish. 2009 Mar;6(1):69-77.

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demonstrate , dose , morpholino , specificity , synergy , two-noniverlapping-oligo


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