CLC LightSpeed Module 25.0

Released on December 3, 2024

Improvements to tools

LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal

Updates to these tools are likely to result in differences in the identified variants compared to results from earlier versions.

• LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal
  • Annotate variants with average quality.
  • Produce an improved mapping through testing of more seeds for reads with low alignment scores.
  • Support analysis of UMI data generated using the QIAseq Multimodal DNA/RNA Library Kit.
• LightSpeed Fastq to Germline Variants
  • Supports analysis of data from non-diploid organisms.
  • Provides the following new options for variant detection:
    • Sensitivity and precision
    • Minimum average quality
    • Minimum QUAL
    • Minimum allele count
  • Has updated default settings:
    • The maximum repeat element length that variants must have to be annotated as Homopolymer/STR Yes, has been lowered from 3 to 2.
    • Variants annotated as Homopolymer/STR Yes, are not included in the variant track if the frequency is below 25%.
• LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal
  • Provide the following new options for variant detection:
    • Minimum frequency (%)
    • Minimum average quality
  • Have updated default settings:
    • The Quality trim limit has been changed from 0.01 to 0.05, leading to less stringent trimming. The default quality trim limit is now the same for germline and somatic analyses.
    • SNV and Indel minimum allele count has been lowered from 3 to 2, this can lead to an increase in the number of detected variants.
    • SNV and Indel minimum per-strand allele count has been lowered from 1 to 0, this can lead to an increase in the number of detected variants.
    • The maximum repeat element length that variants must have to be annotated as Homopolymer/STR Yes, has been lowered from 3 to 2.
    • Variants annotated as Homopolymer/STR Yes, are not included in the variant track if the frequency is below 5%.
  • The maximum value of the SNV and Indel significance thresholds using global error rate has been increased from 0.1 to 1. Increasing the threshold allows detection of more variants.
• Various minor improvements

Copy Number Variant Detection (WGS)

Updates to this tool are likely to result in differences in the identified CNVs compared to results from earlier versions.

• Copy Number Variant Detection (WGS) is no longer in beta.
• Purity and ploidy can be estimated from the data.
• Regions with low mappability can be handled through providing a Umap mappability track:
  • Windows where the Umap score is low are not included when calculating the average coverage of a sample.
  • CNVs where the Umap score is low are filtered out.
  • Umap mappability tracks for hg19 and hg38 are available under the QIAGEN Sets in the Reference Data Manager.
• Estimation of sex chromosome status (XX and XY) has been improved.
• When normalizing the coverage on sex chromosomes, the estimated ploidy is taken into account. Windows on X and Y that are estimated to be copy number neutral, will therefore have an average normalized read count that approximates 1, irrespective of the the number of X and Y chromosomes in the sample.
• Neighbouring CNVs with the same copy number can be merged.
• The output CNV track includes new annotations:
  • Distance to the nearest centromere, only provided when a centromere track is provided
  • Average mappability, only provided when a Umap mappability track is provided.
• The output “Masked regions” track lists regions that were masked:
  • Regions where the ambiguous nucleotide content in the reference sequence exceeded 25%.
  • Regions that had low mappability, only included when a Umap mappability track is provided.
• The report includes estimated purity, ploidy and sex chromosome status (XX or XY).
• The plots in the report have improved colouring and axis labels.
• Various minor improvements

Changes and improvements to template workflows

Changes

• The template workflow Fastq to Annotated Germline Variants has been replaced with Fastq to Germline Variants (WGS).
• The template workflow Fastq to Annotated Somatic Variants has been replaced with Fastq to Somatic Variants (WGS).
• The template workflow Fastq to Annotated Somatic Variants (Tumor Normal) has been replaced with Fastq to Somatic Variants (Tumor Normal) (WGS).
• Each WGS template workflow produces a read mapping and includes QC for Read Mapping.
• The template workflow Fastq to Annotated Germline Variants with Coverage Analysis has been renamed to Fastq to Germline Variants (WES).
• The template workflow Fastq to Annotated Somatic Variants with Coverage Analysis has been renamed to Fastq to Somatic Variants (WES).
• The template workflow Fastq to Annotated Somatic Variants (Tumor Normal) with Coverage Analysis has been renamed to Fastq to Somatic Variants (Tumor Normal) (WES).

Improvements

• The following workflows have been updated to use MANE genes, mRNA, and CDS Reference Data Elements:
  • Fastq to Germline Variants (WGS)
  • Fastq to Germline Variants (WES)
  • Fastq to Somatic Variants (WGS)
  • Fastq to Somatic Variants (WES)
  • Fastq to Somatic Variants (Tumor Normal) (WGS)
  • Fastq to Somatic Variants (Tumor Normal) (WES)
  • QIAseq Pro Fastq to Annotated Germline Variants
  • QIAseq Pro Fastq to Annotated Somatic Variants
• The Indel strand balance threshold has been increased to 1.0 in the template workflows QIAseq Fastq to Somatic Variants and QIAseq Pro Fastq to Somatic Variants. This allows detection of more indels.
• Using {input:1} or {2:1}, output names from template workflows including LightSpeed Fastq to Germline Variants or LightSpeed Fastq to Somatic Variants include the name of the first input fastq file.
• Using {input:1} or {2:1}, output names from template workflows including LightSpeed Fastq to Somatic Variants Tumor Normal include the name of the first tumor input fastq file.
• All template workflows use {2:1} as output naming.
• Various minor improvements

Bug fixes

• Fixed an issue that could cause LightSpeed Fastq to Germline Variants to call deletions that did not have read support for the precise start and end position.
• Fixed an issue that could cause the histogram showing read lengths before and after adapter trimming in the report from the LightSpeed Fastq to Variants tools to show wrong values.
• Fixed an issue that could cause Copy Number Variant Detection (WGS) to call overlapping CNVs.
• Various minor bug fixes

CLC LightSpeed Module 24.1

Released on June 25, 2024

New features and improvements

New tools

• Copy Number Variant Detection (WGS) can be used to identify copy number variants from whole genome sequencing (WGS) data.
• Calculate TMB Score (LightSpeed) can calculate Tumor Mutational Burden (TMB) scores from variant tracks produced by LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal.

UMI improvements available in LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal

• Duplex UMI protocols are now supported.
• Presets for UMI length and common sequence trimming are available for a number of third party duplex UMI protocols.
• Variants are annotated with information about the UMI reads supporting them, for example the proportion of singleton UMI reads and average read pairs per UMI.
• Fastq files where the UMI sequences are present in the fastq file headers are supported.
• Broken reads are grouped to UMI reads. Previously, they were kept as singletons.
• UMI grouping is faster on samples with high coverage.
• The maximum number of read pairs that can be grouped into one UMI read pair has been increased from 20,000 to 100,000.

Reporting

• Reports from LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal have been updated:
  • Mapping information in the summary section and in the detailed sections provide numbers for proper, broken, specific proper and non-specific proper read pairs instead of the previous “Final read pairs”.
  • Information about insert size, quality metrics on input reads and the length of realigned/reassembled regions has been added.
  • When used as input to Create Sample Report, a set of summary items that can be used for quality control are now available.
• All LightSpeed template workflows include Create Sample Report and output a sample report with quality control traffic lights.

Performance related improvements

• The following improvements have been made to LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal:
  • The read mapping has been improved through updates to seeding of reads as well as to realignment and reassembly. The improved read mapping can lead to better variant calls, in particular, fewer false positive variants may be detected.
  • Improvements have been made that make it possible to call more tandem duplications, such as those affecting FLT3.
• The number of regions that are ignored during variant calling by LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal has been greatly reduced.
• LightSpeed Fastq to Germline Variants has a new option that can be used to set minimum allele frequency. The option is enabled by default and the threshold is set to 0.15.

Other improvements

• New options allowing filtering of variants based on homopolymer and short tandem repeat (STR) information are available in LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal.
• Variants called by LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal are annotated with homopolymer and short tandem repeat (STR) information.
• Unmapped reads can be output from LightSpeed Fastq to Germline Variants and LightSpeed Fastq to Somatic Variants.
• In LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal a new option that allows quality trimming from the 3′ end only is available. The option is recommended when using deduplication and is enabled by default.
• The history on output elements from LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal contains numbers that align with the values given in the updated reports.
• Various minor improvements

Bug fixes

Issues affecting LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal

• Fixed an issue that could cause primer trimming to truncate deletions partially overlapping a primer.
• Fixed an issue in deduplication that caused consensus read pairs calculated from non-specific reads to become specific.
• Fixed an issue that could cause reference alleles to be duplicated.
• Fixed an issue that in rare cases caused significantly extended runtime of realignment. This could happen when read mappings contained highly complex regions where it was difficult to determine the best alignment.

Issues affecting LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal

• Fixed an issue that could cause too short versions of longer true positive deletions to be called instead of the full length deletion.
• Fixed an issue that caused the ignored regions track to contain regions outside target regions.

Issues affecting LightSpeed Fastq to Germline Variants

• Fixed an issue that could cause the count and coverage on insertions and deletions to be slightly inflated.
• Fixed an issue where adjacent germline variants with a count of one would not be linked together to form one longer variant.

Other bug fixes

• Fixed as issue that caused the PHS-3200Z QIAseq Targeted DNA Pro Human Comprehensive Hereditary Cancer Research Panel to be missing in the QIAseq Panel Analysis Assistant.
• Various minor bug fixes

CLC LightSpeed Module 24.0

Released on January 9, 2024

Improvements

• QIAseq template workflows are available in QIAseq Panel Analysis Assistant.
• QIAseq Pro template workflows use the GenomeReferenceConsortium_masking_hg38_no_alt_analysis_set masking track by default.
• Output names and output folder structures have been aligned between the different template workflows.
• When reads longer than 800 base pairs are provided as input, LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal will cancel the analysis.

Bug fixes

• Fixed an issue affecting LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal, that, very rarely, could cause variants in regions with many overlapping variants to not be reported.
• Fixed an issue affecting LightSpeed Fastq to Germline Variants, LightSpeed Fastq to Somatic Variants and LightSpeed Fastq to Somatic Variants Tumor Normal, that could cause the individual reads in broken pairs to be mapped to positions that did not give the best alignment score.

CLC LightSpeed Module 23.1

Released on December 07, 2023

New tools

• LigthSpeed Fastq to Somatic Variants takes fastq files as input, maps and realigns the reads and calls somatic variants. Each step has been optimized for rapid processing.
• LigthSpeed Fastq to Somatic Variants Tumor Normal takes tumor and normal fastq files as input, maps and realigns the reads, calls somatic variants and subtracts the variants that are also present in the normal reads. Each step has been optimized for rapid processing.

New workflows

• Workflows are available for somatic variant calling from WES and WGS data as well as for generating control coverage tables for CNV detection:
  • Fastq to Annotated Somatic Variants
  • Fastq to Annotated Somatic Variants with Coverage Analysis
  • Fastq to Annotated Somatic Variants (Tumor Normal)
  • Fastq to Annotated Somatic Variants (Tumor Normal) with Coverage Analysis 
  • Fastq to Somatic CNV Control
• Workflows are available for analysis of QIAseq Targeted DNA and QIAseq Targeted DNA Pro panel data, as well as for generating control coverage tables for CNV detection from QIAseq data:
  • QIAseq Fastq to Annotated Germline Variants 
  • QIAseq Fastq to Annotated Somatic Variants
  • QIAseq Fastq to Germline CNV Control
  • QIAseq Fastq to Somatic CNV Control
  • QIAseq Pro Fastq to Annotated Germline Variants 
  • QIAseq Pro Fastq to Annotated Somatic Variants
  • QIAseq Pro Fastq to Germline CNV Control
  • QIAseq Pro Fastq to Somatic CNV Control

Changes

• The template workflow Fastq to CNV Control has been renamed to Fastq to Germline CNV Control.

Improvements to LightSpeed Fastq to Germline Variants

• The following improvements have been made to LightSpeed Fastq to Germline Variants:
  • Analysis of QIAseq Targeted DNA and QIAseq Targeted DNA Pro panel data is supported. Updates include new functionality for UMI grouping, common sequence trimming, and primer soft-clipping.
  • Tandem duplications and inversions can be inferred from unaligned ends.
  • In variant tracks, fragment count and coverage replace read count and coverage. This means that one paired read is now only counted once, before, read 1 and read 2 were counted separately.
  • Variants are annotated with forward and reverse count, forward and reverse coverage and forward/reverse balance.
  • Realignment has been improved to better balance insertion of SNVs and indels in the read mapping. Previously realignment could favour inclusion of indels over SNVs.
  • Deduplication can now collapse read pairs where the 3' end of read 1 and/or read 2 is not in the same position. This improves deduplication of reads that have been quality trimmed.
  • Creating a read mapping is significantly faster.
  • The report can now be processed by Create Sample Report and Combine Reports.
• The template workflow Fastq to Annotated Germline Variants with Coverage Analysis includes the element Create Sample Report and outputs a sample report.

Bug fixes

• Fixed an issue where insertions and deletions called by LightSpeed Fastq to Germline Variants could have too high count. This could happen when there were similar overlapping insertions or deletions represented in the reads.
• Fixed an issue that caused LightSpeed Fastq to Germline Variants to fail if fastq files did not end with a new line.
• Fixed an issue that caused LightSpeed Fastq to Germline Variants to hang indefinitely if R1/R2 pairs of fastq files contained a different number of reads.

Compatibility

At time of release, CLC LightSpeed Module 23.1 is available for CLC Genomics Workbench and Server 23.0.5 or higher. It will be compatible with later releases in this major release line.

CLC LightSpeed Module 23.0.2

Released on May 25, 2023

Bug fixes

• Fixed an issue that caused LightSpeed Fastq to Germline Variants to assume that fastq files for paired reads were provided in the order R1, R2, and that read pairs were provided consecutively.
  If R1 and R2 were swapped for a given read pair, the same variants would be called, but the direction of the paired reads in the mapping was reverse of what it should have been.
  If R1 and R2 were swapped between different read pairs leading to a different number of reads in R1 and R2, the tool would hang indefinitely producing no results.
  Windows users are likely not affected, because the system alphabetically re-orders fastq files. Mac users can be affected because the order that files are selected is retained when starting an analysis.
• Various minor bug fixes

CLC LightSpeed Module 23.0.1

Released on February 23, 2023

Improvements

• Speed improvement to outputting read tracks from LightSpeed Fastq to Germline Variants.

Bug fixes

• Fixed issues affecting LightSpeed Fastq to Germline Variants that could cause variant count, coverage and frequency to be incorrect:
  • When the option “Ignore non-specific matches” was used, non-specific matches were still included when calculating count, coverage and frequency. Variants in regions with ambiguously mapped reads were affected.
  • Broken read pairs were included when calculating count, coverage and frequency. Variants in regions with broken reads were affected.
  • Variant calls and zygosity were not affected by the issues, see Important QIAGEN CLC software notifications for further details.
• Various minor bug fixes

CLC LightSpeed Module 23.0

Released on January 17, 2023

This is the first release of this product.