Align and Verify
Course Content
Align to a Reference DNA Sequence
Simulate Agarose Gel Electrophoresis
Common Questions About Using SnapGene
What sequencing file formats can be aligned to a reference sequence in SnapGene?
SnapGene accepts Sanger trace files in .ab1, .scf, and .ztr formats, as well as .fastq and .fasta consensus sequences from whole plasmid sequencing (Oxford Nanopore). Plain sequence files can also be aligned. The workflow differs depending on the input type: Sanger traces use the Align Sanger Reads tool; whole-plasmid reads use a dedicated Align a Whole Plasmid Sequence workflow that requires a circular reference sequence.
How does SnapGene display discrepancies between sequencing reads and the reference?
Discrepancies are highlighted in the alignment view and can be navigated sequentially using jump arrows. For each mismatch, you can expand the trace view to inspect peak quality, check read orientation, and see how the difference would affect any translated features. Low-quality ends of traces are automatically hidden during import based on configurable stringency settings.
Is the Align to Reference tool the same as a map-to-reference assembly?
No. SnapGene's documentation explicitly distinguishes these. The Align to Reference tool aligns each read independently to the reference sequence and is intended for construct validation. It does not assemble reads into a consensus. For de novo assembly of overlapping Sanger reads, SnapGene provides a separate Assemble Contigs tool based on the CAP3 algorithm.
Can I mark a construct as experimentally confirmed after alignment?
Yes. Once all aligned reads are in agreement with the reference sequence, you can open the Info panel and check the "Confirmed experimentally" option. This records the verification status in the sequence file.
What can SnapGene's agarose gel simulation predict?
The simulation predicts band patterns for restriction digests (up to four enzymes per lane), PCR amplification products, and uncut circular DNA, including supercoiled migration. You can configure the number of lanes (up to 26), the percentage of agarose, the electrophoresis run time, buffer type, and select from a list of commercially available MW markers. Hovering over any band shows fragment size, cut sites, and coordinates.
What is the difference between this video series and the Working with Primers series?
This series covers construct verification after sequencing: aligning Sanger or whole plasmid reads to a predicted reference and simulating gel electrophoresis to confirm digest or PCR results. The Working with Primers series covers the earlier design stage: creating and importing primers, designing primers with 5' extensions, and simulating directed mutagenesis.
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