Open preprint reviews by Mikhail Koksharov

Comparative assessment of fluorescent proteins for in vivo imaging in an animal model system

Jennifer K Heppert, Daniel J Dickinson, Ariel M Pani, Christopher D Higgins, Annette Steward, Julie Ahringer, Jeffrey R Kuhn, Bob Goldstein

It's really useful data for the scientific community!

"To ask whether differences in fluorescent

protein abundance could account for the differences in fluorescence intensity we
observed, we analyzed protein levels in each of our single-copy transgenic
strains by western blot (Supplemental Figure S1). We observed approximately 2-
fold higher levels of mex-5 driven GFP::PH protein compared with mNG::PH
protein (Supplemental Figure S1, C, paired t-test, p=0.0408), which may be due
to partial transgene silencing or post-transcriptional regulation of these

1) Have you checked the mRNA levels and half-lives? As I've observed myself in mammalian cells, mRNA half-lives associated with a CDS can be very different if (functionally the same) reporter protein is derived from different species or subjected to codon optimizations.
This in turn can affect protein levels.

2) Can the differences be due to different protein half-lives of these FPs?

3) Regarding the Fig S1-C:
Is it possible to compare protein levels with some technique that would give better CIs?
Also, there may be some problem with the confidence procedure since some CIs cross zero.

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