Reformat and Upload a Coordinates File
| 1. | Open the original TruSeq Custom Amplicon design coordinate input file (*.csv) in a spreadsheet editor, such as Excel. |
| 2. | Delete the following columns: |
| • | TargetType |
| • | PaddingPerExon |
| • | Include3UTR |
| • | Include5UTR |
| • | Labels |
| 3. | Rename the StartCoordinate column into Start. |
| 4. | Rename the StopCoordinate column into End. |
| 5. | Insert a column to the left of the Name column and enter Type in the header. |
| 6. | Enter REGION for all entries in the Type column. |
| 7. | Make sure that all entries in the Chromosome column include prefixes that match the chromosome format of the reference genome. See Expected Chromosome Format. |
| 8. | Make sure that all entries in the Name column are 20 alphanumeric characters or less. Hyphens, periods, underscores, and whitespaces are allowed. |
The name field can be empty.
| 9. | Save and close the coordinate input file. |
| 10. | Create a new AmpliSeq for Illumina Gene design to target exons and large (> 125 bp) regions or an AmpliSeq for Illumina Hotspot design to target SNVs and small (< 125 bp) regions. See Create an AmpliSeq for Illumina Gene Design or Create an AmpliSeq for Illumina Hotspot Design. |
Make sure to select the species that matches the original TruSeq Custom Amplicon design. For human TruSeq Custom Amplicon designs, select Human (hg19).
| 11. | Configure the new AmpliSeq for Illumina design by selecting the Sample Type and Max Amplicon Length that most closely match those of the original TruSeq Custom Amplicon design. |
Use the following table for selecting equivalent amplicon sizes between the two assay types:
|
TruSeq Custom Amplicon design Amplicon Size |
AmpliSeq for Illumina custom design Max Amplicon Size |
|---|---|
|
150 |
140 |
|
175 |
175 |
|
250 |
275 |
|
425 |
375 |
| 12. | Use the default setting for Stringency. |
| 13. | Select Next. |
| 14. | Upload the reformatted coordinate input file into the AmpliSeq for Illumina design. See Add Targets by Uploading Files. |