From: ****
Sent: Wednesday, January 18, 2023 10:17 AM
To: Jon Moulton
Subject: Translation-blocking Vivo-MO reduces gene expression on the level of mRNA?
Hello Jon,
I find that the translation-blocking Vivo-MO is able to significantly reduce the copies on the level of mRNA. I use qPCR and RNAseq to confirm that the mRNA of my target gene is down-regulated by more than 4-fold at 24 hours after injection, and by more than 10-fold at 7 days after injection. Such knockdown effect is impressive, but theoretically, translation blocking MO only blocks translation, which should reduces the level of protein, rather than the level of mRNA. I am just curious about why translation-blocking MO also have effect on the mRNA level.
I also try splice-modifying Vivo-MO, which also knockdown gene expression significantly. But in my case, translation-blocking MO always induces stronger phenotype than the splice-modifying one. I hope this information will be useful to other researchers.
Have a nice day.
Best,
****
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Hi ****,
Binding a Morpholino to an RNA can change the stability of the RNA -- we hypothesize that this is due to change of the secondary structure making the RNA more susceptible to nucleolytic attack. However, we cannot predict what kind of change will occur: when Morpholinos bind RNA there have been reports of degradation, stabilization and lack of effect on RNA lifetime. Because of this we suggest that all translation blocking Morpholinos should be assessed at the level of protein using an immunochemical assay; this gives a more reliable signal.
Thanks for reporting your observation of a decrease in the RNA on binding a Morpholino translation-blocking oligo. May I have your permission to post these two messages on my blog? I would be happy to remove identifying information from your email if you like (let me know your preference). I think this would be useful exchange for others to read.
Thanks!
- Jon
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