To set ROI regions from a BED or GBK file
If you select Mitochondrial amplicon analysis, then in addition to loading the GenBank Mitochondrial reference file, you must load a BED file that includes the amplicon regions. You can also load an amplicon BED file for targeted sequencing analysis to display bases that align outside of the target regions or to primer regions as soft-clipped. If you choose to load an amplicon BED file to carry out targeted sequencing analysis, then following alignment, reads are matched to an amplicon as defined in the BED file. Any bases that fall outside the amplicon region, for example, primer sequences, are soft-clipped. If a read cannot be properly matched to an amplicon, then the entire read is soft-clipped.
 | The Amplicon BED file should be used for only for amplicon-based targeted sequencing methods, not for capture-based methods and the BED file should define specific amplicon regions, not including primers. |
The ROI Viewer track also displays a solid pink link indicating the amplicon regions. (See Track panes.) In this case, ROIs can be defined in either a BED file or GenBank file.
| | Setting regions from GBK files is applicable only if you load a GBK reference file. You can download GenBank format references from the NCBI website (http://www.ncbi.nlm.nih.gov/). If the GBK file does not have the necessary information about the ROIs, then you can manually add the information to the file. See Advanced GBK Editor tool. |
1. Load the reference file.
| | For the next step, Set Amplicon BED file should be used for only for amplicon-based targeted sequencing methods, not for capture-based methods. |
2. Select one of the following as appropriate – Set Amplicon BED file or Set ROIs from GBK files.
3. Do one of the following:
• If you selected Set Amplicon BED file, click Set to open a dialog box, and then browse to and select the appropriate BED file.
• If you selected Set ROIs from GBK files, no further action is required.