CORALL Total RNA-Seq Library Prep Kit
The CORALL Total RNA-Seq Library Prep Kit enables fast and cost-efficient generation of UMI labelled, stranded libraries for whole transcriptome analyses using Illumina® NGS platforms.
CORALL generates transcriptome-wide smooth, uniform read coverage, comparable to that of competitor kits (Fig. 1).
Figure 1 | Accumulated transcript body coverage (whole transcriptome). Coverage across all transcripts was generated using the geneBody_coverage.py tool provided by RSeQC (transcripts length normalized to 100 %).
CORALL delivers excellent gene discovery rates matching conventional competitor kits (Fig. 2A). 95% of genes are commonly detected between CORALL and each of the two competitors (at >10 counts per million (CPM), Fig. 2B). This high level of library complexity ensures faithful representation of the transcriptome, enabling sensitive expression profiling.
Figure 2 | Gene detection. A) Gene discovery rates. The number of detected genes is plotted against the total number of reads mapping uniquely to exons (calculated with featureCounts). B) Overlap of detected genes. The Venn diagram illustrates overlaps between CORALL and competitor kits for genes detected with normalized expression levels >10 CPM (for uniquely mapping reads).
Superior End-to-End Coverage
CORALL’s comprehensive coverage delivers improved transcript start and end site representation. Read coverage was analyzed using the ERCC spike-in controls, which feature precise, known transcription start and end sites (TSS and TES, respectively). CORALL reads map more accurately to the exact ERCC TSS (Fig. 3A) than competitor libraries, which fail to cover the true start sites. Additionally, CORALL provides elevated coverage at TES (Fig. 3B).
CORALL libraries can be prepared from poly(A)-enriched or rRNA-depleted RNA (100 – 200 ng of total RNA yields approximately 1 – 2 ng of rRNA-depleted RNA). We recommend using Lexogen’s RiboCop rRNA Depletion Kit V1.2 (Cat. No. 037) or Poly(A) RNA Selection Kit (Cat. No. 039). Total RNA without prior depletion or enrichment (100 pg to 100 ng) can also be used as input.
Unique Molecular Identifiers (UMIs) seamlessly included
UMIs are seamlessly introduced into CORALL libraries as an inherent part of the protocol. As a result Read 1 contains the UMI information making it directly accessible in the cost-efficient single-read mode.
CORALL libraries can be multiplexed using up to 96 i7 indices (included in the kit). Additional i5 indices can also be introduced using the Lexogen i5 6 nt Dual Indexing Add-on Kits (Cat. No. 047). Used together, Lexogen’s 96 i7 and 96 i5 indices enable up to 9,216 different index combinations for sequencing.
Fast and Easy All-in-One Protocol
The streamlined RNA fragmentation-free protocol generates ready-to-sequence libraries in 4.5 hours.
or rRNA depleted RNA as input.
of Displacement Stop Primers (DSP) with partial Illumina-compatible
P7 sequences, to the RNA template. No prior RNA fragmentation is
necessary, as the insert size is determined by the distance between
hybridized DSPs. Reverse transcription extends each DSP to the next,
where transcription is effectively stopped. This stop prevents spurious
second strand synthesis, maintaining excellent strand specificity.
cDNA fragments then introduces partial Illumina-compatible P5
sequences and Unique Molecular Identifiers (UMIs).
double-stranded cDNA is amplified. In doing so, i7 and (optional)
i5 indices as well as complete adapter sequences required for
cluster generation on Illumina instruments are added.
during sequencing and can therefore be assessed even in Single-Read
mode. All purification steps are based on magnetic beads, rendering
the protocol highly suitable for automation.
Frequently Asked Questions
Please find a list of the most frequently asked questions below. If you cannot find the answer to your question here or want to know more about our products, please contact firstname.lastname@example.org.
CORALL Total RNA-Seq Library Prep Kit for Illumina
User Guide – Release 28.02.2019
PCR Add-on Kit for Illumina Instruction Manual – update 03.07.2019
i5 Dual Indexing Add-on Kits (5001-5096) Instruction Manual – update 24.05.2018 (release of additional i5 Unique Dual Indexing Add-on Kit (5001-5096))
Data Analysis Pipeline Instruction Manual – Release 17.07.2019
Material Safety Datasheets
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