We have sequenced Synechococcus sp. PCC 7002 genome. Already the same strain has been sequenced and annotated by many before. Whole-genome map is also available online. The gene of interest is not present in NCBI gene bank assembly of the same strain. What we want to know is that wether our strain has been mutated and whether it has this particular gene or not? I have assembled and annotated the sequenced files using velvet and Prokka but do not know the further exact procedure to get efficient results.
Get the nif genes of closest to Synechococcus sp from NCBI,
Blast nif genes gene against your assembled genome.
If no hits in blast, then you may go for targeted re-sequencing
target gene sequencing.
Use Long-read technologies
find a sequence of this gene in NCBI and use BLAST to look for it in the genome you just assembled.
Which kind of cluster are you looking for? They are tools which screen for clusters depending on the function. If its an individual gene with unknown sequence function I would go for protein domain search. You need to run blast of the whole proteome and screen for suspected proteins. Let me know your objective and I may come with more help.
Best
Ireneusz Stolarek Thanks...that looks quite simple. I will try that. Thanks
Loubna Youssar Nif genes (encodes for enzymes involved in atmospheric nitrogen), mostly located on the chromosome, but in some organisms even found in plasmids.
The easiest way is to blast nif genes already known against your genome. Use Galaxy europe server for that.
Get the nif genes of closest to Synechococcus sp from NCBI,
Blast nif genes gene against your assembled genome.
If no hits in blast, then you may go for targeted re-sequencing
target gene sequencing.
Use Long-read technologies
Use BLAST of the nif genes from the closest species from NCBI. You can use tblastn agianst the whole genome and then using the cordinate of the BLAST hits predict the gene using any prokaryotic gene prediction tool such as GeneMarkS, Glimmer3 Or Prodigal and annotate it and if it hits to nif genes then select it. Now you can use muscle program followed by codeml to check if the mutation undergone positive or constraint selection.
Thanks everyone for the valuable info....i did use blast... and i found a good tutorial by Kat Holt and David Edwards for begginers in comparative genomics. Here is the link
https://holtlab.net/2017/07/01/update-to-comparative-bacterial-genomics-tutorial/
Follow This Link Please:
https://www.yourgenome.org/facts/how-do-you-identify-the-genes-in-a-genome
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