The sequence of the best hit ( Ruminococcus faecis) is located at the right upper row of your sequence).Īlignment your sequence against the hit sequence ( Ruminococcus faecis) by clicking (c).Īfter the automated pairwise alignment, you will see the erroneous region that does not match to other reference sequences (box (a) of the above). Check your sequence (from ab1 file) is at the bottom (b). Move the cursor (a) to the bottom of the screen. Your sequence is located at the bottom of the alignment. Start EzEditor2 software and open the file that you have just downloaded. This file contains your sequences and reference sequences that show the highest sequence similarities. Click (c) to view the detailed result of “Identify”.Ĭlick (a) to download a data file for EzEditor2 tool. We will start the trimming process using a sequence alignment tool called EzEditor2. As you will see the chromatogram shown above, your sequence contains errors at the front end (as well as backend). This is cutoff is applicable for high-quality sequence only. The generally accepted 16S similarity cutoff is 98.7%. In this case, sequence similarity is 95.35% and you may be excited to discover a potentially new species. The hit species (a) is displayed with pairwise sequence similarity (b). Go to and paste your sequence as new query. Then, search through the EzBioCloud’s Identify. This file contains the result of the fairly good sequencing reaction. This contains a 16S sequence of a human fecal bacterium. If you used the sequence data without trimming the ends, your sequence will have a substantial amount of errors. Because of the sequencing chemistry, both ends of sequence contain substantial errors.
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Quality and manual inspection of the raw data is possible with the chromatogram.
An ab1 file contains both the chromatogram and sequence data (after base calling).The following example ab1 files are required.Ĭharacteristics of Sanger sequencing data You need a sequence alignment tool for 16S gene sequences.If your sequencing service provider did not give you ab1 files, ask for ab1 files. An ab1 file contains the chromatogram information from ABI sequencers ( Learn more).
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This document explains how to use EzBioCloud’s “Identity” service using single Sanger sequencing data.
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