Generation Information and Q&A
What are the common alignment tools used?
Please see this page for more details.
Can I align optical mapping assemblies to assemblies, instead of optical maps to assemblies?
Yes. You can take any of the optical maps, optical mapping assemblies, or in-silico digested sequences as alignment queries or alignment reference freely.
Are the optical maps well-aligned?
There are several indicators for the quality of an alignment. These include but not limited to: number of signals aligned, ratio of aligned signals to extra or missing signals, and scaling error. Usually, alignment software already combines these factors to calculate a score or determine the statistical significance that reflects the alignment quality. However, in complex genomic regions such as those with exceptionally high or low signal density, repetitive elements and segment duplication, or that structural variations exist between the reference and sample genomes, the alignment quality parameters may not be accurate and you may need to customize your own quality assessment scheme.
How should I set the alignment parameters?
Parameters are very different across different software, and you should read the respective software manual for details. Here we give you two general tips about alignment parameters.
First, alignment parameters are different for queries and references with different lengths. For example, aligning the optical mapping assemblies (long query) to sequence scaffolds (long reference) is different from aligning optical maps (short query) to optical mapping assemblies (long reference). You should adjust your parameters according to the software manual. Note that some alignment software only targets certain query-reference combination.
Second, adjustment of alignment quality filtering parameters is usually a trade-off between precision and recall. Depending on the tasks you want to accomplish, choose a more stringent parameter if you want to the results more precise, or a more lenient parameter if you want to capture more alignments.
Detailed commands
Alignment (Molecules to References / Assemblies)
Alignment of short molecules to a reference.
java -jar OMTools.jar OMBlastMapper --refmapin Reference.ref --optmapin Molecules.data --optresout AlignmentResult.oma --alignmentjoinmode 1 --filtermode 1 --thread 40
This command aligns molecules onto the reference with filter (–filtermode) and split-mapping (indel) (–alignmentjoinmode) enabled using 40 threads (–thread).
Alignment (References / Assemblies to References / Assemblies)
Since both references and assemblies are longer and contain more signals and fewer errors, users could set a higher score cut-off for alignment. Also, translocation detected should be more confident. The potential overlapping part could also be a problem.
java -jar OMTools.jar OMBlastMapper --refmapin Reference.ref --optmapin Assembly.data --optresout AlignmentResult.oma --alignmentjoinmode 3 --filtermode 1 --minscore 50 --thread 10 --writeunmap false
This command aligns assemblies onto the reference with filter (–filtermode) and split-mapping (translocation) (–alignmentjoinmode) enabled using 10 threads (–thread). The minimum score needs be increased to at least 50 (–minscore) when split-mapping (translocation) is enabled. The unaligned contigs is not written (–writeunmap).
Re-alignment
After initial alignment, you want to refine the alignment at the target region.
java -jar OMTools.jar OMBlastMapper --refmapin Reference.ref --optmapin Assembly.data --optresin InitialAlignmentResult.oma --optresout RealignmentResult.oma --alignmentjoinmode 1 --filtermode 1 --minscore 15 --minmatch 3 --thread 10 --writeunmap false
This command re-aligns assemblies onto the reference based on the initial alignment result. The alignment is limited to align on the initial aligned region extended by query size.
Alignment filtering
Filtering removes alignment results that do not fulfill the criteria.
java -jar OMTools.jar ResultTools --refmapin Reference.cmap --optresin AlignmentResult.oma --optresout FilteredAlignmentResult.oma --filtermode 1 --minscore 50
This command filters AlignmentResult.oma by using a minimum score of 50.
Alignment format conversion
Format conversion allows users to convert alignment formats to the desired one. Note that some information may be lost throughout the conversion process.
java -jar OMTools.jar ResultTools --optresin AlignmentResult.oma --optresout AlignmentResult.xmap
This command converts alignment result in oma format to xmap format.
Renaming
Rename the query or reference inside the alignment results.
java -jar OMTools.jar ResultTools --optresin AlignmentResult.oma --optresout RenamedAlignmentResult.oma -qprefix Query1_ -rprefix Ref1_
This command adds the prefix “Ref1_” to the references and “Query1_” to the queries in the alignment file.
Alignment selection
Alignment selection allows quick extraction of alignments according to the query ID or the aligned region.
java -jar OMTools.jar ResultTools --optresin AlignmentResult.oma --optresout RenamedAlignmentResult.oma --dataid Query1 Query2 Query3
This command extracts alignments with query name “Query1”, “Query2”, “Query3” to the output.
java -jar OMTools.jar ResultTools --optresin AlignmentResult.oma --optresout RenamedAlignmentResult.oma --region chr1:1-1000000
This command extracts alignments at chr1:1-1000000.