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How close can one pack protein and tRNA's genes

| posted 18 Feb, 2016 16:58
OK, general question. We have a new phage with lots of tRNA genes packed interdigitated with glimmer/Gene protein predictions. There are long spaces between the tRNA genes so there is certainly room for a protein gene but some of the glimmer and genemark predictions come pretty close to the aragorn hits. How much space should there be between these features? Do we need to leave room for a promoter anytime we switch from a tRNA gene to a protein gene or vice versa?

Should I tell the students to leave at least 25-50 bp needed for the promoter? I will have students look at the protein vs. tRNA distribution in the host bacteria but wanted to check if there is some kind of general rule to maintain consistency across all SEA phages annotations.
Edited 18 Feb, 2016 17:34
| posted 24 Feb, 2016 15:03
Nope, the tRNA cassettes are weird, and no you don't have to have a promoter for each of them or for each protein encoding gene interspersed within the cassette. We tend to steer clear of a tRNA and a protein occupying the same space, but there are definitely genomes where they get pretty close (take a look at the Ms). The only promoter rule is still about switching directions– presumably, switching transcriptional directions would still require two promoters even for tRNAs.
| posted 24 Feb, 2016 17:46
OK thanks.
Looking more closely we have now found at least several tRNA's where there are NO bases between the end of the tRNA and the first base of the auto-annotation. Splicing of the RNA to excise the tRNA would thus produce a leaderless message. I have been talking to our local faculty who works almost exclusively with Streptomyces (the phage host in this case) and apparently in some of these strains leaderless messages are not that uncommon, so I think we will go ahead and annotate both no matter how close as along as they don't overlap. I tend to agree with the general annotation policy that it is better to annotate a false positive than a false negative; so when things are truly ambiguous over-predict don't under-predict.

He also raised the possibility that this is an intentional organization that could be used as some sort of regulation of expression, by splicing out an overlapping tRNA you effectively kill the protein encoded gene, resulting in less protein production than if the tRNA wasn't there. I don't know if this has ever been described before but would be very cool if true.

| posted 02 Feb, 2017 19:41
To follow up with this discussion, we have a tRNA gene identified by Aragorn on the +3 frame and a clear ORF on the -2 frame. In other words, they are in opposite orientation on the genome.

However, the both look very "REAL". The Frames output is below.

https://drive.google.com/open?id=0B_7KYneUM8kdVWRKWjJrQlFBajQ

We are planning to call both unless we hear otherwise here.

Let me know your thoughts. Thank you. GF
Edited 16 Feb, 2017 15:20
| posted 02 Feb, 2017 19:42
{Deleting Duplicate Message}
Edited 02 Feb, 2017 20:09
| posted 09 Feb, 2017 16:08
Has tRNA-scan-SE been changed? It now directs to:

https://galaxy.pasteur.fr/forms::trnascan and none of the advanced functions detailed in the Annotation Guide apply (pp. 97-100)
| posted 10 Feb, 2017 15:59
Yes the old tRNA-scan-SE site was shut down as of last November. You will need to follow the updated protocol which is on the seaphages faculty info page, Bioinformatics section:

Click here for Revised tRNA evaluation protocol (Section 9.5).
| posted 10 Feb, 2017 17:57
Perfect. Thanks!
| posted 14 Feb, 2017 16:01
tRNAscan-SE 2.0 has moved again: http://lowelab.ucsc.edu/tRNAscan-SE/
| posted 16 Feb, 2017 15:17
GregFrederick@letu.edu
To follow up with this discussion, we have a tRNA gene identified by Aragorn on the +3 frame and a clear ORF on the -2 frame. In other words, they are in opposite orientation on the genome.

However, the both look very "REAL". The Frames output is below.

https://drive.google.com/open?id=0B_7KYneUM8kdVWRKWjJrQlFBajQ

We are planning to call both unless we hear otherwise here.

Let me know your thoughts. Thank you. GF

To clarify: The ORF and the tRNA gene in question are overlapping, and on the opposite strands. See the picture of the region in the image at the link.

I did not see a reply to this question. We are about finished with the annotation portion of the course. However, we are uncertain how to deal with this region.

Thoughts?
Edited 16 Feb, 2017 15:22
 
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