I am basically interested in understanding the graph obtained after ribosome profiling and how can you know from the graph in which translational state are the mRNAs?
The mRNAs present in the polysome fraction are recruited more often by the translational apperature of the cell. A shift of a given mRNA into the polysomal fraction is often connected with higher translational efficiency. In the polysomal profiles you often see the shift, of a mRNA, after a treatment before it is detected on the proteome level.
If you isolate RNA from the individual fractions, you can easily amplify 18S and 28S RNA and follow the distribution of 40S and 60S SUs als well as monosomes and polysomes in an agarose gel , which will match your gradient profile. The same way you can detect shifts of specific mRNAs. An example can be found in Naarmann-de Vries, I.S. et al., J. of Cell Science (2016).
I would rather like to extend a question in this discussion. I want to check translational block of a gene using polysome analysis but I m confused how to differentiate between actively translating genes from translationally blocked genes??
because both kind of genes could have multiple ribosome units on them. Kindly shed some light.
Many thanks..
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