Example 1: Finishing a PacBio HGAP assembly

Example workflow for finishing PacBio HGAP Assembly

1. Load a multiFASTA file of the assembly by selecting “Load Assembly” from the file menu

2. View the assembly by selecting “View Assembly” from the “View” menu, ensure “All” is selected in the dropdown box next to “Contigs to view:”

3. To identify spurious contigs perform a self-comparison between contigs using the “Self-comparison tool” from the “View” menu. Phenomena such as prophage tail fibre allele switching can cause contig breaks and spurious contig to form. Remove small contigs that align internally to larger contigs from the assembly.

4. To identify plasmids and reconstruct the chromosome perform another self-comparison. This time ensure the “show intra contig hits” and “only show edge hits” checkboxes are ticked. Show intra-contig hits will allow us to identify circularising contigs and “only show edge hits” will treat the resulting comparison as a graph so when scaffolds are made it will remove overlapping edges.

5. Identify circularising contigs and write to individual FASTA files remove contig from canvas once written.

6. Order and reorientate the remaining contigs, choose “Select All” from the tools menu.

7. Check order and orientation of contigs is correct in the Contig list and then select “Write to FASTA” from the tools menu.

8. Map reads back to output contigs using your favourite read mapper to confirm new assembly.