Verify clones with Sanger reads

Upload your sequencing traces, align them to the expected plasmid, and read the differences.

Check one trace

1. Run Sanger Verification & Alignment from the Tools panel or Ctrl/Cmd+K, or press Ctrl/Cmd+Shift+L. It opens as the Alignment & Verification tab.
2. Drop your files on Drop AB1 or FASTQ files here, or use the upload button.
3. Pick a trace under Recent Traces.
4. Click Align to Reference, search for the plasmid, then click Align.
5. Step through differences with Previous mismatch and Next mismatch.
6. To trim low-quality ends before aligning, use the trim control in the toolbar.

To compare two pasted sequences instead, choose Pairwise Alignment on the start screen.

Screen a batch of clones

1. On the start screen, choose Multi-Trace Alignment.
2. Under Save to, pick a project.
3. Pick one reference plasmid and tick the traces.
4. Set Trim stringency: Low keeps most data, Medium is balanced, High trims hard.
5. Optionally fill in Batch name and pick an entry to link.
6. Click Run Alignment. You can leave the page; open the finished batch from Jobs.

Several plasmids in one batch

1. Tick Multi-reference mode.
2. Each trace is matched to a plasmid by file name. Check the matches and fix any that look wrong.
3. To set matches from a file, click Import sample sheet and choose a CSV or TSV.

How calls are made

Assemble reads into a contig

Use Assemble Sequencing Reads in the Tools panel to join overlapping reads into one consensus.