KAPA RNA HyperPrep Kits

Efficient rRNA depletion using KAPA RNA HyperPrep with RiboErase (HMR) leads to fewer non-informative reads and the detection of more unique transcripts compared to other suppliers.

Libraries were generated in quadruplicate using variable inputs of Universal Human Reference (UHR) RNA (Agilent Technologies) with rRNA de[pletion prior to library construction. Where present, error bars represent the standard deviation. For 25 ng samples, paired end (2 x 100 bp) sequencing was performed using an Illumina® HiSeq® 2500 instrument. Reads aligning to rRNA were removed, and reads were randomly subsampled to 14 M for comparative analyses. Transcripts were quantified using RNA-SeQC. The 10 ng input amount is lower than the validated minimum input for both KAPA RNA HyperPrep workflows; for these samples, paired-end (2 x 75 bp) sequencing was performed using an Illumina NextSeq 500 instrument. Reads were randomly subsampled to 14 M for comparative analysis prior to removing reads aligning to rRNA and subsequent marking of duplicates. Transcripts were quantified using Kallisto (data not plotted due to analysis and sequencing depth differences).

KAPA RNA HyperPrep Kit with RiboErase (HMR) Globin simultaneously removes rRNA and globin mRNA from blood-derived RNA. Together, effective RNAase H depletion and highly efficient library construction result in fewer non-informative reads 

(A) compared to the workflow from Supplier I, which employs bead-based depletion (orange). This translates to more complex libraries (B) and a larger number of unique transcripts detected (C).

Libraries were prepared from different inputs of RNA extracted from human blood, as indicated on the x-axis of each graph. The 25 ng input is lower than the recommended minimum input for the Supplier I workflow. Paired-end (2 x 125 bp) sequencing was performed on an Illumina® HiSeq® 2500 instrument. Data were sub-sampled to 17 M reads per sample for analysis. Each bar represents the average of three technical replicates. Transcript abundance was quantified using Kallisto. To assess off-target depletion, transcript abundances were aggregated at the gene level and TMM-normalized prior to differential expression analysis. The expression profiles of libraries generated with or without globin depletion were compared to assess off-target depletion for each workflow.

Efficient mRNA capture using KAPA mRNA HyperPrep leads to fewer non-informative reads and the detection of more unique transcripts compared to other suppliers. 

Libraries were generated in quadruplicate using variable inputs of Universal Human Reference (UHR) RNA (Agilent Technologies) with mRNA capture prior to library construction. For 50 ng samples, paired end (2 x 100 bp) sequencing was performed using an Illumina® HiSeq® 2500 instrument. Reads aligning to rRNA were removed, and reads were randomly subsampled to 14 M for comparative analyses. Transcripts were quantified using RNA-SeQC. The 10 ng input amount is lower than the validated minimum input for both KAPA RNA HyperPrep workflows; for these samples, paired-end (2 x 75 bp) sequencing was performed using an Illumina NextSeq 500 instrument. Reads were randomly subsampled to 14 M for comparative analysis prior to removing reads aligning to rRNA and subsequent marking of duplicates. Transcripts were quantified using Kallisto (data not plotted due to analysis and sequencing depth differences).

Single-cell full-length total RNA sequencing uncovers dynamics of recursive splicing and enhancer RNAs

Hayashi et al. describe the first method for full-length total RNA-sequencing for single cells, which they developed using KAPA HiFi DNA Polymerase, KAPA RNA HyperPrep Kit with RiboErase (HMR), and KAPA Library Quantification Kit. This method — random displacement amplification sequencing (RamDA-seq) — enables the investigation of gene expression dynamics, RNA-processing, and transcriptional regulation in single cells.

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Comparing library preparation methods for SARS-CoV-2 multiplex amplicon sequencing on the Illumina MiSeq platform

Batty, E.M. et al.

bioRxiv 20202020
In the article, Batty, E.M. et al. present their findings from a comparison of three different library preparation methods used in the ARTIC Network protocol for SARS CoV- 2 multiplex amplicon sequencing on the Illumina® MiSeq® platform. The authors report that the ligation-based KAPA HyperPrep method resulted in higher library yield, more total reads, and equal or better overall coverage than the tagmentation-based Nextera Flex or Nextera XT workflows.

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Blocking of the CD80/86 axis as a therapeutic approach to prevent progression to more severe forms of COVID-19

Julia et al.

aRxiV.org2020

Julia et al. used KAPA RNA HyperPrep with RiboErase (HMR) Globin to investigate the transcriptional responses of SARS-CoV-2-infected patient samples to the immunomodulating drug Abatacept, and to compare this response to that of cells and blood from Rheumatoid Arthritis patients. The authors used this data to determine whether blocking the CD80/86 axis is a viable therapeutic target to prevent severe immune responses of COVID19 patients.

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Hydroxychloroquine inhibits trained immunity – implications for COVID-19

Rother et al.

medRxiV.org2020

Rother, N. et al. constructed total RNA-seq libraries using KAPA RNA HyperPrep Kit with RiboErase (HMR) to perform transcriptome analysis of isolated monocytes of COVID-19 patients. These cells demonstrated increased expression of interferon-stimulated genes, which are likely markers of disease severity as they are associated with a poor clinical outcome. Using the Kapa HyperPrep Kit, the authors also constructed ChIP-seq libraries to investigate the effect of hydroxychloroquine on trained immunity at the functional and epigenetic level.

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COVID-19 Related Genes in Sputum Cells in Asthma: Relationship to Demographic Features and Corticosteroids.

Peters M. C. et al.

PubMed2020

Peters et al. used the KAPA mRNA Hyper Prep Kit in Whole Transcriptome RNA-seq to investigate host expression levels of angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2), two proteins believed to mediate SARS –CoV2 viral infection of host cells.

SARS-CoV-22020RNA-Seq

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Clinical benefit of remdesivir in rhesus macaques infected with SARS-CoV-2

Williamson et al.

bioRxiV.org2020

Williamson et al. use the KAPA HyperPlus Library Preparation Kit and KAPA Unique Dual-Indexed Adapters to prepare sequencing libraries from double-stranded cDNAs converted from SARS-CoV-2 viral RNA. Williamson et al. also amplified the resulting libraries using KAPA HiFi HotStart ReadyMix and quantified them with the KAPA Library Quantification Kit prior to paired-end sequencing.

LQK2020SARS-CoV-2DNA-Seq

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A Territory-wide study of COVID-19 cases and clusters with unknown source in Hong Kong community: A clinical, epidemiological and phylogenomic investigation.

Leung K. S. et al.

medRxiV.org2020

Leung et al. prepared sequencing-ready libraries with the KAPA HyperPrep DNA Library Prep Kit following generation of amplicons by multiplex PCR using ARCTIC network nCoV-2019 primers; through that workflow, Leung et al. investigated the epidemiological and phylogenomic characteristics of COVID-19 cases in Hong Kong.

SARS-CoV-22020DNA-Seq

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Patient-derived mutations impact pathogenicity of SARS-CoV-2

Yao H. et al.

medRxiV.org2020

Yao et al. performed whole viral genome sequencing using KAPA RNA-Seq library prep kit from COVID-19 patient isolates to understand whether acquired mutations of the SARS-CoV-2 genome have an impact on the pathogenicity of the virus.

SARS-CoV-22020RNA-Seq

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Transcriptional profiling of immune and inflammatory responses in the context of SARS-CoV-2 fungal superinfection in a human airway epithelial model

de Lamballerie, C.N. et al.

bioRxiV.org2020

Following construction of poly(A)-enriched RNA-seq libraries with the KAPA mRNA HyperPrep Kit, de Lamballerie et al. compared the host transcriptional response to SARS-CoV-2 infection alone to that of SARS-CoV-2 with Aspergillus superinfection to elucidate the impact of fungal superinfections on the course and severity of viral infections in respiratory epithelial cells.

2020RNA-SeqSARS-CoV-2KAPA mRNA HyperPrep Kits

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Expression of SARS-CoV-2 Entry Molecules ACE2 and TMPRSS2 in the Gut of Patients with IBD
Burgueno et al.

Inflammatory Bowel Disease2020

Burgueno et al. used the KAPA RNA HyperPrep Kit with RiboErase (HMR) and the LightCycler® 480 System to understand the expression of the viral entry molecules angiotensin I converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) in the intestine. Additionally, the authors investigated how inflammatory states in the gut of patients suffering from inflammatory bowel disease (IBD) and taking immune-modu

2020RNA-SeqSARS-CoV-2LightCycler 480KAPA RNA HyperPrep Kits with RiboErase

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The circulating SARS-CoV-2 spike variant N439K maintains fitness while evading antibody-mediated immunity

Thomson, E.C. et al.

bioRxiv 20202020

Immune evasion mutations that maintain virulence and fitness can emerge within the SARS-CoV-2 spike protein, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics. Thomson, E.C. et al. used the ARTIC network amplicon sequencing protocol and the https://doi.org/10.1101/2020.11.04.355842 to conduct epidemiological and genome surveillance of SARS-CoV-2 and

SARS-CoV-22020DNA Library Preparation

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