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A retrovirus with RNA genome infected a host cell. You would like to isolate the host cell’s RNA (rRNA, tRNA, and mRNA) from the virus RNA. What properties can you rely on to determine the three types of host cell RNA- for example, in terms of their size, complexity, GC content, or abundance in the cell? The undetermined RNA can be identified as the virus RNA.

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    $\begingroup$ This is a homework question, right? $\endgroup$ – Chris Apr 12 '15 at 16:03
  • $\begingroup$ @Chris I am a Bioengineering student preparing a presentation for the next lecture, so it kinda is a homework. $\endgroup$ – Sam Apr 12 '15 at 16:08
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    $\begingroup$ So what's your idea of solcing this problem? $\endgroup$ – Chris Apr 12 '15 at 16:11
  • $\begingroup$ Here in Germany we are at the beginning of the 2. semester and up until now what i can think of is using radioactive Uracil, but that would only 'seperate' the DNA from all RNA. Another idea would be using Polymerase-Chain-Reaction. $\endgroup$ – Sam Apr 12 '15 at 16:21
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    $\begingroup$ @Sam Well if you know the virus and know its gene sequence, you might be able to use RT-PCR to get DNA copies of its RNA. I can't think of a way to take total RNA from a cell and physically separate host RNA from viral RNA. $\endgroup$ – user137 Apr 13 '15 at 14:42
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Thermo Fisher has a protocol for the separation of host mammalian RNA from prokaryotic RNA, optimized for E. coli vs human, mouse or rat sequences. Capture oligonucleotides bind portions of the mammalian RNA and hybridize. These hybridized oligo/RNA are then removed from solution via "oligonucleotide-derivatized magnetic beads," after which the remaining RNA assumed to be from whatever prokaryote can be precipitated out w/ ethanol. If you know enough about your virus and your host, I don't see why a similar assay can't be created. Otherwise, Thermo Fisher has additional links to RNA isolation protocol.. most of the viral assays, however, utilize serum, plasma, etc. because host DNA/RNA will elute along with the viral RNA if they're present, as in samples containing cells.

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  • $\begingroup$ Retrovirus is not prokaryotic; so that kit may not work. But the the principle is, as you mentioned, the same: adsorb oligos that will bind specifically to viral RNA on beads and remove the viral RNA. Modified oligos such as LNA can be used for a stronger binding and longer life. $\endgroup$ – WYSIWYG Apr 14 '15 at 4:41
  • $\begingroup$ Right, i was thinking more of a modified assay based on the same principles. $\endgroup$ – CKM Apr 14 '15 at 22:33

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