Intro to Ultima Bioinformatics
Get Started Analyzing your Ultima Genomics Data
Analysis Best Practices for multiple sequencing applications including: ppmSeq, germline WGS, somatic WGS, single-cell, spatial, methylation, proteomics and RNA-Seq
Generalized Ultima Analysis Pipeline
General Outline for Steps for Secondary Analysis Steps of Ultima Data
Trimming
The Ultima Trimmer software tool divides each read into different segments and processes each segment, including pass-through, trimming, as well as barcode and adapter matching. Information about each segment can be written to configurable SAM tags.
Alignment and Tagging
Reads are tagged (barcodes, UMI, insert, etc.) and aligned (if the specified application requires alignment to a reference). For more information on Realignment: Ultima Aligner (UA) Github Page:
Demultiplexing
Data output is sorted, per sample, into folders based on the sample barcode used for demultiplexing.
UMI-Aware Sorting and De-duplication
Unique molecular identifiers can be used to identify clonal copies of molecules and remove them.
Sample QC
Application-specific QC files are generated. Examples: ppmSeq, Methylation.
File Output
Read data is output in a variant calling-ready CRAM file or an application-specific FASTQ file ready for third-party analysis tools.
For more information on Output File Structure per application see: Solaris Informatics Run Output Reference Guide
For more information on implementing applications on the UG 100 sequencing platform see: UG Supported Applications Guide
Application Specific Links
ppmSeq
ppmSeq (paired-plus-minus sequencing) is Ultima's ground-breaking WGS method to achieve accuracy of one part-per-million or better for calling SNVs via sequencing of both strands of a template molecule. ppmSeq identifies and removes disagreement between strands of a captured DNA molecule caused by sample errors. ppmSeq on the UG 100 sequencing platform will output a variant calling-ready CRAM file as well as a ppmSeq-specific QC file.
Whole Genome Sequencing - Germline Variant Calling
Ultima utilizes Efficient DV, an analysis pipeline designed to call variants from aligned CRAM files, using a version of DeepVariant (DV) adapted for Ultima Genomics data. Germline variant calling also includes copy number variation (CNV) and structural variant (SV) calling pipelines.
WGS Somatic
Ultima utilizes Efficient DV, an analysis pipeline designed to call variants from aligned CRAM files, using a version of DeepVariant adapted for Ultima Genomics somatic data. Whole Genome somatic variant calling also includes structural variant (SV) calling pipelines.
Single-Cell/Spatial
Single cell and spatial analysis on the UG 100 sequencing platform is limited to secondary analysis pipelines that generate output files tailored for direct input into downstream analysis tools specific for supported assay preparation kits.
Methylation
Whole genome methylation sequencing on the UG 100 sequencing platform includes read trimming, alignment (using a methylation-aware reference), and sorting. This is followed by automated on-instrument secondary methylation analysis using Methyl Dackel which provides output of methylation statistics and QC information.
Proteomics
Ultima Genomics’ UG 100 sequencing platform provides on-instrument analysis support for Olink proteomics libraries including sequence matching of Olink barcodes to reference whitelists. The identified barcodes are tabulated to generate a counts file (hist.csv) that is designed for input directly into Olink’s NPX analysis software.
RNA-seq
RNA-Seq on the UG 100 sequencing platform outputs an unaligned CRAM file that is ready for input into downstream alignment tools such as Spliced Transcripts Alignment to a Reference (STAR).