Sequence of Indices (Barcodes) Used for Multiplexing
QuantSeq libraries are intended for a high degree of multiplexing. With QuantSeq for Illumina up to 9,216 samples can be uniquely barcoded in one lane by using the up to 96 external i7 indices (7001-7096) included in the kit together with the 96 external i5 indices (5001-5096), which are part of the Lexogen i5 6 nt Dual Indexing Add-on Kit (Cat. No. 047).
Automated Data Analysis Pipeline
The QuantSeq data analysis pipeline has been implemented on the Bluebee® Genomics Platform and in the Partek Flow software, offering to every user, even without bioinformatics experience, the opportunity to analyze QuantSeq samples in a convenient and fast way.
The pipeline on the Bluebee® Genomics Platform is accessible through a complimentary code provided with each QuantSeq kit. Learn more and get started at https://www.bluebee.com/lexogen/.
Activation codes for additional runs can be purchased from Lexogen via the webshop.
The QuantSeq pipeline in the Partek Flow software is available for any Partek Flow user at http://www.partek.com/lexogen-quantseq-pipeline.
QuantSeq – FWD Data Analysis
The following analysis pipeline demonstrates a principal workflow.
The used tools are freely available and to a great extend part of the Galaxy platform1. All proprietary Software, for instance CLC workbench, should perform similarly.
The script examples uses mainly the bash internal for-loop; in Galaxy for instance, the single files have to be processed manually one by one.
Following software packages (and their dependencies) should be installed when following the given protocol:
- STAR aligner
Create a working directory and the subdirs fastq, qualitycheck and download the de-multiplexed fastq files into the fastq dir.
Let the fastq files be:
Let your STAR index files be in /data/star/human, the file truseq_rna.fa.gz is provided by bbmap and is here symbolically linked to /data/resources/truseq_rna.fa.gz.
The file /data/resources/polyA.fa.gz is a simple GNU-zipped fasta file containing a single entry with 18 ‘A’s.
1Goecks, J, Nekrutenko, A, Taylor, J and The Galaxy Team. Galaxy: a comprehensive approach for supporting accessible, reproducible, and transparent computational research in the life sciences. Genome Biol. 2010 Aug 25;11(8):R86.