QIAseq FastSelect –rRNA HMR is a new formulation that works to remove mitochondrial and cytoplasmic rRNA from human, mouse, rat and other mammalian samples. QIAseq FastSelect HMR shows greater potency than QIAGEN’s species-specific formulations currently on the market.
A QIAseq FastSelect –rRNA HMR Kit works to remove unwanted RNAs from your RNA-seq library using 1 step. This is significantly easier than hybridization or RNase H methods, which require 29 to 37 steps (e.g., Supplier I, Supplier N and Supplier T).
B QIAseq FastSelect –rRNA HMR Kit works in only 14 minutes, which saves hours versus other methods, and is 30% faster than existing QIAseq FastSelect RNA Removal Kits.
Simply add QIAseq FastSelect reagent (rRNA Removal and/or Globin Removal) to your RNA sample, perform fragmentation (if required), then initiate a stepwise cool-down from 75°C to 25°C over 14 minutes. Researchers can then complete the remaining library preparation steps without any additional changes to their workflow. QIAseq FastSelect works with or without RNA fragmentation, providing the flexibility to use FFPE or degraded RNA samples, or high-quality RNA as part of a standard RNA-seq library construction workflow.
QIAseq FastSelect –rRNA HMR Kits, QIAseq FastSelect –Globin Kits and QIAseq FastSelect –rRNA/Globin Kit have been tested with the QIAseq Stranded Total RNA Lib Kit (QIAGEN), TruSeq Stranded (Illumina), NEBNext Ultra II Directional (New England Biolabs) and KAPA RNA HyperPrep (KAPA Biosystems) library preparation kits.
QIAseq FastSelect –rRNA HMR Kit removes more than 95% of the rRNA from human, mouse, rat, horse, cow, hamster and sheep samples and >80% removal for dog, monkey, chicken, rabbit and pig samples.
QIAseq FastSelect –rRNA HMR was designed to remove both cytoplasmic (5S, 5.8S, 18S, 28S) and mitochondrial (12S, 16S) rRNA for human, mouse and rat samples.
QIAseq FastSelect –Globin was designed to remove adult, fetal and embryonic globin mRNA from human, mouse and rat samples.
QIAseq FastSelect –rRNA HMR is a new formulation of QIAseq FastSelect which provides a single-tube solution for human, mouse and rat samples. The new formulation shows an increased potency in removing rRNA with highly reproducible results. The improved QIAseq FastSelect –rRNA HMR Kit exhibits greater rRNA removal than kit previous versions.
QIAseq FastSelect –rRNA HMR Kit is a single-tube solution for human, mouse, rat and other mammalian samples. Gene expression data from human, mouse and rat universal RNA samples was compared using QIAseq FastSelect –rRNA HMR Kit and the previous versions of the kit. This data shows that the single-tube QIAseq FastSelect –rRNA HMR Kit can replace previous versions of QIAseq FastSelect RNA Removal Kits, which used species-specific human, mouse and rat solutions.
To test the reproducibility of the QIAseq FastSelect –rRNA HMR Kit and QIAseq Stranded Total RNA Lib Kit,100 ng of human universal RNA (HURR) was used in duplicate. Following RNA-seq library preparation and sequencing, the data was aligned using CLC Genomics Workbench. Mapped read percentages were highly reproducible, with 49% mapping to exons, 45% mapping to introns and 5% mapping to intergenic regions. Only 1.2% of the reads mapped to rRNA following treatment – a reduction of >95%.
To test the reproducibility of the QIAseq FastSelect –rRNA HMR Kit and QIAseq Stranded Total RNA Lib Kit, 100 ng of human universal RNA (HURR) was used in duplicate. Following RNA-seq library preparation and sequencing, the data was aligned using CLC Genomics Workbench. An average of 14,929 genes were detected with an RPKM >0.3.
To test the reproducibility of the QIAseq FastSelect –rRNA HMR Kit and QIAseq Stranded Total RNA Lib Kit, 100 ng of human universal RNA (HURR) was used in duplicate. Following RNA-seq library preparation and sequencing, the data was aligned using CLC Genomics Workbench. Expressed genes with an RPKM >0.3 were compared between the 2 replicates resulting in a high degree of concordance.
Stranded transcriptome libraries were prepared from 100 ng and 1 µg aliquots of human blood total RNA using the QIAseq FastSelect –rRNA/Globin Kit and QIAseq Stranded Total RNA Lib Kit.
To remove rRNA and globin mRNA during the QIAseq library prep, QIAseq FastSelect –rRNA HMR + –Globin was combined with each RNA sample and 5x RT Buffer. The samples were then heated (fragmented) at 95°C for 15 min and stepwise cooled to 25°C over 14 minutes. The remaining library prep steps were then performed according to the QIAseq Stranded Total RNA Lib Kit instructions. Sequencing was performed on a NextSeq 550 and data analysis was performed using CLC Biomedical Workbench. The percent or reads mapping to rRNA and globin is shown.
Stranded transcriptome libraries were prepared from 100 ng and 1 µg aliquots of human blood total RNA using the QIAseq FastSelect –rRNA/Globin Kit and QIAseq Stranded Total RNA Lib Kit.
To remove rRNA and globin mRNA during the QIAseq library prep, QIAseq FastSelect –rRNA HMR + –Globin was combined with each RNA sample and 5x RT Buffer. The samples were then heated (fragmented) at 95°C for 15 min and stepwise cooled to 25°C over14 minutes. The remaining library prep steps were then performed according to the QIAseq Stranded Total RNA Lib Kit instructions. Sequencing was performed on a NextSeq 550 and data analysis was performed using CLC Biomedical Workbench. Mapped read percentages were highly reproducible with 35% mapping to exon, 58% mapping to introns and 5% mapping to intergenic regions. Only 1–2% of the reads mapped to rRNA and less than 1% mapped to globin transcripts.
Stranded transcriptome libraries were prepared from 100 ng and 1 µg aliquots of human blood total RNA using the QIAseq FastSelect –rRNA/Globin Kit and QIAseq Stranded Total RNA Lib Kit.
To remove rRNA and globin mRNA during the QIAseq library prep, QIAseq FastSelect –rRNA HMR + –Globin was combined with each RNA sample and 5x RT Buffer. The samples were then heated (fragmented) at 95°C for 15 min and stepwise cooled to 25°C over 14 minutes. The remaining library prep steps were then performed according to the QIAseq Stranded Total RNA Lib Kit instructions. Sequencing was performed on a NextSeq 550 and data analysis was performed using CLC Biomedical Workbench. Gene expression results for 100 ng and 1 µg of RNA, for genes with an RPKM >0.3, show a high degree of correlation.
FFPE RNA reference standards A FFPE Horizon 5-Fusion Multiplex Positive and B FFPE Horizon 5-Fusion Multiplex Negative Control were used to prepare duplicate RNA-seq libraries using QIAseq FastSelect –rRNA HMR and QIAseq Stranded Total RNA Lib Kits. QIAseq FastSelect consistently removed >95% of the rRNA from the sample even though the input RNA was highly fragmented.
FFPE RNA reference standards A FFPE Horizon 5-Fusion Multiplex Positive and B FFPE Horizon 5-Fusion Multiplex Negative Control were used to prepare duplicate RNA-seq libraries using QIAseq FastSelect –rRNA HMR and QIAseq Stranded Total RNA Lib Kits. Removal of rRNA leads to an increase in exon mapping. FFPE Horizon 5-Fusion Multiplex Positive and Negative Control samples show an exon mapping rate of approximately 40% versus 10% in the untreated samples.
When rRNA depletion is performed using QIAseq FastSelect (Log2 normalized gene expression, targets with RPKM >0.3), gene expression results are highly reproducible.
QIAseq FastSelect –rRNA HRM removes rRNA from FFPE RNA samples.
A QIAseq FastSelect results in highly efficient removal of rRNA from fragmented samples. The percentage of reads mapped to rRNA is shown. B Exon/intron/intergenic read mapping is shown. QIAseq FastSelect results in a substantial increase in the percentage of reads mapped to exonic regions.
QIAseq FastSelect –rRNA HRM removes rRNA from FFPE RNA samples.
When rRNA depletion is performed using QIAseq FastSelect (Log2 normalized gene expression, targets with RPKM >0.3), QIAseq FastSelect treatment vs. no treatment results suggest that QIAseq FastSelect does not perturb the natural expression profile of samples.
