Ion AmpliSeq™ HD Custom panels: DNA hotspot design input specifications
For Ion AmpliSeq™ HD Custom DNA hotspot designs, use a text file with genomic coordinates to upload several hotspot targets at once. The file can be in CSV or BED format, with the following restrictions.
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The genomic coordinates must include at least 1 base and at most 125 bases.
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The start coordinate should always be less than the end coordinate.
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Genomic regions are represented as number pairs (start base, end base). The system of genomic coordinates used in all Ion AmpliSeq™ inputs follows the convention of "0‑based start, 1 based end" also known as "Zero-based, half-open". For example, in a zero based system the first 100 bases of a chromosome are defined as chromStart=0, chromEnd=100, and span the bases that are numbered 0-99. Similarly, the region comprising the second 100 bases would be represented as (100, 200). For more information on this system of coordinates, visit http://genomewiki.ucsc.edu/index.php/Coordinate_Transforms.
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Ion AmpliSeq™ Designer does not support designing primers for mitochondria targets. Designs for any coordinates on “chrM” will be rejected. If the mitochondrial DNA sequence is submitted for a custom reference design, the design will proceed. However, we recommend an accompanying list of polymorphisms to avoid at primer binding sites. We strongly recommend not mixing mitochondrial amplicons with those of nuclear DNA.
The following input files are accepted for Ion AmpliSeq™ HD Custom DNA hotspot designs. To view example files, in the target addition pane, click Input Specifications, then click Download this example for each file type of interest. You can copy/edit the example files to create panel‑specific input files.
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File type |
Description |
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SNP List |
An SNP list is a TXT file of list that contains SNP IDs. It is similar to a CSV file, but contains only one column. COSMIC and dbSNP are available for hg19 designs. |
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CSV File |
A comma separated values (CSV) file that contains multiple columns. Each row can either be a region or a SNP. The required fields for a region row are in the following order: Type (“REGION”), a name for the region (any name of your choice), chromosome number (any one of chr1–chr22, chrX, chrY), a starting genomic coordinate, and an ending genomic coordinate. The fields for a SNP row are: Type (“SNP”) and SNP ID, or SNP ID only. The file is not required to have a header but it can include one as shown the following example. Example CSV File contents, where each column is separated by a comma: |
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BED File |
A BED file is a tab-delimited format file, with one line per hotspot. The three required fields are listed in order: Chromosome (any one of chr1–chr22, chrX, chrY), a starting genomic coordinate, and an ending genomic coordinate. We recommend an additional column for a name of your choice, but it is not required. A header in the file does not prevent the file from being uploaded but generates a warning error. For more information on BED files, visit http://genome.ucsc.edu/FAQ/FAQformat.html#format1. |