Define Regions

A region is a contiguous range of positions within one reference sequence. Most analysis steps can restrict their work to one or more such regions rather than the full reference length.

Define a region by coordinates

Specify the first (5’) and last (3’) position using 1-based numbering (the first base of the reference is position 1). Both endpoints are included.

Use --region-coords (-c) followed by the reference name and the two coordinates:

seismic filter -c ref-1 34 71 out/

defines a region spanning positions 34–71 of ref-1. Repeat -c to define multiple regions.

Define a region by primer sequences

For amplicon samples prepared with RT-PCR, it is usually more convenient to define a region by primer sequences than by coordinates. SEISMIC-RNA locates the exact primer-binding sites in the reference (no mismatches are permitted) and creates a region spanning from one base downstream of the forward primer’s 3’ end to one base upstream of the reverse primer’s 5’ end.

Use --region-primers (-P) followed by the reference name and the two primer sequences (give the reverse primer as written on the oligonucleotide itself, not as its reverse complement):

seismic filter -P ref-1 TCGAAT GTTACG out/

To exclude a few extra bases near each primer end (to reduce edge artifacts), add --primer-gap N (default 0).

Define regions in a file

To define many regions reproducibly or share definitions across multiple runs, list them in a CSV file and pass it with --regions-file (-i):

seismic filter -i regions.csv out/

See Metadata for Regions for the CSV format.

Multiple regions per reference

Repeat -c, -P, or rows in a --regions-file to define as many regions as you need for each reference. Regions may overlap. Regions for one reference have no effect on any other reference.

Region names

  • Regions from -c or -P are named {first}-{last} (e.g. 34-71).

  • Regions from --regions-file use the name in the Region column.

  • If no region is defined for a reference, one region named full spanning the entire reference is created automatically.

See also