Link to this post | posted 27 Jan, 2018 01:31
ptsourkas	Is there a cutoff value of the SD score for start codons, i.e. a value below which you would eliminate a start codon from consideration? I use Kibler6 and Karlin medium per the DNA Master guide. Thanks, Philippos

Link to this post | posted 27 Jan, 2018 18:12

cdshaffer

According to the "Guiding Principles" I would say there is no lower cutoff. Context matters for interpretation of SD scores, for example, "If the genes are part of an operon, a 4bp overlap (ATGA), where a start codon overlaps the stop codon of the upstream gene, is preferred by the ribosome. Therefore RBS scores may have little bearing in this type of gene arrangement." Also, this: "The preferred start site usually has a favorable RBS score within all the potential start codons, but not necessarily the best. A notable exception is the integrase in many genomes, which has a very low RBS score. Our experimental data suggests that some genes do not have an SD sequence." {Bold added by me}

Link to this post | posted 24 Feb, 2020 14:03

Pollenz

We are annotating a Gordonia DJ phage Secretariat. It appears there is an region across most of these DJ genomes that begins about 32,000bp and contains ~15 small genes that are divergent across the different phages and all the genes are separated by 20-100bp gaps between them (no 1-4bp overlaps or small <10bp gaps). The issue is that the called start for many of these in NOT the LO that in many cases will significantly reduce the gaps between these genes. It appears that the RBS data is very poor (example, Z value 0.446, spacer 8, final score -8, compared to the "called" start of 1.77/11/-5.3) for many of these. All of the genes are hypothetical, so no evidence of how these genes are related (an opernon??). The Coding Potential may drop off at the LO on some of these, but its not that different from many genes that have been called. We know that there are many aspects that impact translation initiation beyond the RBS/SD such as RNA structure, operons, translation of previous genes, etc. What are your thoughts on selecting the LO and reducing gaps when the RBS data is so poor? RS Pollenz

Link to this post | posted 24 Feb, 2020 14:50

lhughes

We see a similar region in the Cluster BI1 phages. When we look more closely at the Phamerator map, we also see the criss-crossing lines of short regions of homology that I see in the map of the Cluster DJ phages. There appear to be some short areas of conserved sequences between those genes (that is what we find in the BI1 phages). I would consider that these conserved sequences probably serve some purpose and are likely intergenic, so extending to LO just to do so might not be the best approach without further analysis of what is in those gaps. I have a graduate student who is doing that for BI1 phages right now to try to understand what is going on there better. Lee

Link to this post | posted 24 Feb, 2020 15:22
Pollenz	Yes, very interesting. There is one sequence of ~44bp that is replicated PRIOR to 5 consecutive genes….. RS Pollenz

Link to this post | posted 25 Feb, 2020 12:58
Pollenz	The short duplicated areas in the DJ phages appear to have sequence similarity to the sequences in the Cluster BI1 phages even though they come from different species. The DJ phages have eight of these directly upstream of start codons: CGCAtatATTACAtGCCtTATAATGAGACCCCTAcGAAAGG(xxxxxxx 6-9bp)ATG RS Pollenz