Table 2 Amplicon Filter Module parameter descriptions and considerations
Parameter | Description | Notes |
|---|---|---|
Strict mode | VaST ignores missing or ambiguous data in input matrix | Speeds up preprocessing but some sites are lost |
Window size | Maximum distance between adjacent sites that can be combined into a single amplicon | The desired amplicon length should be considered when setting the window size. A larger window may increase the number of variant sites that are included in the amplicons making them more efficient |
Primer zone size | Size of the region upstream and downstream of the target to evaluate in the amplicon filter | The primer zones begin immediately before the first and immediately after the last target site in the window, so the maximum amplicon size is 2 × primer zone size + window size. A smaller primer zone may limit the number of primer options. |
Strain Cutoff | The number of strains at a primer zone site that can have a non-conserved call before the site is flagged. | A strain cutoff greater than one will not guarantee that the primer zone sequences are conserved across all of the strains but it may be appropriate in cases where one or a few strains have low sequence coverage |
Primer zone filter percent | The percent of primer zone positions that must be present in un-flagged segments of the primer zone that are longer than the primer zone filter length. | A higher primer zone filter percent will increase the total number of primer options in amplicons that pass the filter |
Primer zone filter length | The length of un-flagged primer zone segments that count toward the primer zone filter percent | The primer zone filter length should be at least as long as the minimum acceptable primer length to ensure that conserved primers can be found within the primer zone |