MiCall performs post-assembly stitching after de novo assembly. This document explains why there are two stitchers and where each detailed design lives:
The shared problem
De novo assemblers such as IVA or Haploflow do not always turn input reads into a single contiguous sequence. They may return multiple contigs: fragmented sequences that can represent adjacent or overlapping portions of the same biological sequence, sometimes encoding the same region more than once.
A post-assembly stitching/refinement step can therefore improve the assembly by systematically arranging those contigs and resolving discrepancies in overlapping regions.
MiCall has two different ways to perform that refinement.
Referencefull stitcher
The referencefull stitcher
(micall/utils/referencefull_contig_stitcher.py,
micall contig_stitcher with-references)
is allowed to use a reference sequence and positions relative to that
reference.
That gives it valuable information unavailable from the contigs alone. In particular, it can reason about ordering and adjacency using the reference coordinate system and can often produce substantially more complete assemblies. Its additional inputs are the reference sequence and, optionally, remap read counts — not the raw-read junction validation used on the referenceless path.
This is useful and intentional.
However, its output is therefore reference-guided. If the biological sequence genuinely differs structurally from the reference — for example through a large deletion, inversion, rearrangement, duplication, or other scrambled structure — reference-derived ordering can potentially obscure or normalize that structure.
Referenceless stitcher
The referenceless stitcher
(micall/utils/referenceless_contig_stitcher.py,
micall contig_stitcher without-references)
exists for a different purpose.
Its goal is:
Improve the initial de novo assembly without using a reference sequence or reference-derived structural assumptions.
It relies only on evidence intrinsic to the sample, such as:
- the contig sequences themselves;
- sequence overlap between contigs;
- short-read evidence;
- other sample-derived linkage evidence actually available to the implementation.
Reference independence is a deliberate property of this output, not an implementation deficiency.
The important use case is that a user may want an improved de novo assembly while still preserving unusual biological structure exactly as supported by the sample. Note that referenceless refinement is a step applied after de novo assembly; “unstitched” output is not automatically reference-free in the same sense.
Why both exist
Neither stitcher is simply “better” than the other. They answer different questions:
referencefull:
What assembly can we obtain when reference-derived structure
is allowed as evidence?
referenceless:
What improvement over the de novo assembly can be justified
from sample-intrinsic evidence alone?
The referencefull result can be more complete. The referenceless result has stronger reference-independence semantics. These are complementary algorithms — different assembly interpretations of the same contigs.
Conceptually — not the literal pipeline dataflow — MiCall has two refinement semantics:
de novo contigs (e.g. IVA, Haploflow)
|
+-- referencefull refinement -------> more complete,
| (reference + contigs reference-guided assembly
| [+ optional remap counts])
|
+-- referenceless refinement --------> improved assembly with
(contigs + reads, reference-independent
no reference structure) structure preserved
In the current denovo pipeline (micall/drivers/sample.py), the
referencefull stitcher in fact runs on the referenceless stitcher’s
output, so the two branches above describe the refinement semantics
rather than two independent pipeline paths.
See the detailed designs for the algorithm behind each interpretation: