Hello,

Making the assumption that your data is DNA (and not RNA), the tools under "NGS: SAM Tools" and "Operate on Genomic Intervals" can generate coordinates of the mapped reads which then can be correlated with known genes/ORFs from your bacterial genome (or related genomes, if you can obtain those those mapped to the same reference genome to use as predictions).


General analysis path:

Starting with your SAM file, use these tools first to obtain an interval file representing your read coverage:
1 - SAM-to-BAM
2 - Generate pileup
3 - Pileup-to-Interval

Next, import a reference known gene set. Sources may include UCSC, NCBI, or other. Download from that source (if not directly available via a "Get Data" source) and upload the file into Galaxy using FTP ("Get Data -> Upload"). If this data is in GTF/T format, you can convert it to Interval using "Convert Formats -> GFF-to-BED" (BED is a stricter form of Interval, use the pencil icon in the datasets box to Edit Attributes to change data type to Interval).

Then if using output from "Pileup-to-Interval" and a reference known gene/transcript dataset mapped to the same reference genome, use the tools in "Operate on Genomic Intervals" to perform comparisons based on genomic coordinates. Each tool has a description on the main tool form, but there are also screencasts explaining the functions here under "3. Interval Operation Tutorial"
http://wiki.g2.bx.psu.edu/Learn/Screencasts
Also see: "Regional Variation -> Feature coverage" for localized comparisons

Once an intersection of coordinates is complete, you may need to use the tools in "Text Manipulation", "Filter and Sort", or "Join, Subtract and Group" to merge in gene identifiers. Exactly what order to use these tools greatly depends on the input reference gene/transcript dataset formats.

If you are doing transcript predictions, the tool "EMBOSS -> getorf" Finds and extracts open reading frames (ORFs)" may be helpful. This tool requires sequence as input. Once predicted transcript coordinates are obtained, extract sequence from the reference genome using "Fetch Sequences -> Extract Genomic DNA" to use as input.

Hopefully this helps to get you started. Please let us know if we can help again,

Best,

Jen
Galaxy team

On 7/12/11 7:02 AM, Joanne Rampersad wrote:
Hi
I am sequencing a bacterial genome and have assembled my Illumina
reads (40 bp single) using bowtie with a reference genome. This
generated a sam file.
I would like to obtain a listing of the open reading frames from the
bacterial genome and the corresponding genes that they are most
similar to.
Can you please give the tools/steps  necessary to do this?

many thanks

Jo
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