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Tm of a Morpholino - Effective? Specific?

A Morpholino user asked me about his oligo design, which I had marked with a Tm of 82.5°C. He wondered about efficacy and specificity of such an oligo. Here's my response.

I think you can get pretty good activity from an oligo with a Tm of 80°C. I mark oligos with Tm of 80°C to 85°C as "fairly low" or "moderately low" RNA affinity so that when deciding between two good oligo designs, you might take the one without a fairly/moderately low Tm note. If I didn't mark it, there would be no way to tell between a very good oligo and a pretty good oligo. I don't start worrying until the Tm is down in the 70s, and those get a "very low" or "quite low" Tm note (at that point I am trying to warn people away). I won't send out oligo designs in the 60s or less. Once an oligo Tm is much over 100°C, I'll include a note saying that this might be too high an RNA affinity for good target specificity.

Since your oligo has a Tm at the lower end of the good range, that is good for the oligo specificity. More bases will need to pair to a target RNA before the oligo has any activity.

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