Why does FASTQC show unexpectedly high sequence duplication levels (PCR-duplicates)?

FASTQC is primarily designed to QC whole-genome shotgun sequencing data. Importantly, it is significantly limited in its analyses because it only works on single reads instead of read-pairs. As a consequence FASTQC tends to generate unnerving warnings for multiple Illumina

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When should I trim my Illumina reads and how should I do it?

Should I trim adapters from my Illumina reads? This depends on the objective of your experiments. In case you are sequencing for counting applications like differential gene expression (DGE) RNA-seq analysis, ChIP-seq, ATAC-seq, read trimming is generally not required anymore

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Where can I find the UMIs in the Tag-Seq data? When and how should I trim my Tag-Seq data? What is the low complexity stretch in the Tag-Seq data?

By default, we will generate Tag-Seq and Batch-Tag-Seq gene expression profiling data that incorporate Unique Molecular Identifiers (UMIs) in the sequence reads. (This FAQ provides information on the usage of UMIs:  https://dnatech.genomecenter.ucdavis.edu/faqs/should-i-remove-pcr-duplicates-from-my-rna-seq-data/ ). Please note that the UMIs provide optional

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Should I remove PCR duplicates from my RNA-seq data?

Should I remove PCR duplicates from my RNA-seq data? The short and generalized answer to the question “Should I remove PCR duplicates from my RNA-seq data?” is in most cases NO.  For some scenarios, de-duplification can be helpful, but only

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Which data will I receive from the PacBio Sequel II sequencer? Will they have quality scores?

We will deliver the complete data set generated by the PacBio Sequel to you securely via Bioshare. For push-button type secondary analyses (combining data for up to 2 SMRT-cells e.g. for demultiplexing, CCS, long amplicon, or IsoSeq analysis) we can run these

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What data will I receive for Illumina sequencing? Demultiplexing, Trimming, Filtering

By default you will receive gzip compressed FASTQ data, as individual files  for each sample (demultiplexed).  The demultiplexing is included in the service if you provide us the barcodes sequences on the submission form. The files will be available for download

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How do I download my sequencing data?

We deliver sequencing data via two portals: SLIMS for Illumina data, and BioShare for PacBio and Nanopore data. Both portals offer secure access to the data and support several download protocols. The emails that will notify you about new sequencing

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Which strand is sequenced for my strand-specific RNA-seq data?

Strand-Specific RNA-Seq Libraries RNA-Seq (conventional) after Poly-A enrichment or ribodepletion: By default we generate strand-specific RNA-seq libraries. Strand-specific (also known as stranded or directional) RNA-seq libraries substantially enhance the value of an RNA-seq experiment. They add information on the originating

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My FASTQ file contains some “N”s. Is there a problem with my data?

Please note that when opening an Illumina sequence fastq file it is expected that the first few thousand reads are of comparatively low quality and frequently contain “N”s.  An “N” means that the Illumina software was not able to make

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How should the miRNA/small-RNA data be trimmed?

We are using the PerkinElmer NEXTflex™ Small RNA-Seq kit for the generation of micro RNA and small RNA-seq libraries because it significantly reduces sequence-specific biases in the library preparation.  For this purpose the adapters oligonucleotides contain 4 randomized bases at the ligation

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