Our Comprehensive Data Analysis Package
Eclipsebio’s standard analysis uses human (hg38), mouse (mm10), rat (rn6), or c. elegans (ce11). Contact us for alternate species including viral genomes.
RAW SEQUENCING DATA | FASTQ
Reads directly off sequencer and demultiplexed per sample for data back-up and publication
GENOME ALIGNMENTS | BAM
Reads filtered of repetitive elements, aligned, and PCR deduplicated for additional downstream analysis
COVERAGE TRACKS | BIGWIG
Normalized read coverage on positive and negative strands for visualization in a genome viewer such as IGV
SCORED PEAKS | BED
Genomic regions scored after input normalization as putative binding sites (aka “peaks”) for target exploration. Includes log2 enrichment scores and p-values per peak.
BINDING SITE SUMMARY REPORT | HTML
Report for enriched peaks with interactive tables and plots for publication ready graphics. Includes enriched GO terms, KEGG pathways, motifs, and repetitive element analysis.
The RBP-eCLIP kit is a simplified and robust library preparation method to identify your RNA-binding proteins targets. Eliminate time-consuming prep work by ordering one kit with 30+ validated reagents. Start immediately and generate eCLIP libraries in 5 days.
An 8 sample kit for simplified and robust library preparation method to identify your RNA-binding proteins targets
Optional Data Analysis (Sequencing NOT included).
Recommended for Researchers with Experience in Library Prep, Immunoprecipitation, and Western Blotting
Each Kit Order Includes Up to 1 Hour of 1-1 Support (via Virtual Meeting/Email)
The RBP-eCLIP kit is a simplified and robust library preparation method to identify your RNA-binding proteins targets.
Eliminate time-consuming prep work by ordering one kit with 30+ validated reagents. Start immediately and generate eCLIP libraries in 5 days.
Published method and protocol improvements increase workflow efficiency and identify true binding sites
Kits includes optimized protocol and validated reagents to ensure reproducibility for better results.
Our eCLIP library preparation kit is an optimized 5-day protocol based on Van Nostrand, Yeo et al. Nature Methods 2016 paper.
Why choose the eCLIP kit?
- 1000-fold improved library prep efficiency vs standard iCLIP-seq
- Increased experimental success rates, decreased wasted sequencing
- Significantly improved signal/noise ratio identifies true in vivo targets
Each kit includes:
- Enough index primers to sequence 16 libraries at once
- Size selection beads
- Quality tested transfer membrane
- Index primers
What are the sequencing parameters?
Eclipsebio’s kit is based on the single-end eCLIP variant described in:
Van Nostrand EL, Nguyen TB, et al. Robust, Cost-Effective Profiling of RNA Binding Protein Targets with Single-end Enhanced Crosslinking and Immunoprecipitation (seCLIP). Methods Mol Biol. 2017;1648:177-200. PMID: 28766298.
Libraries generated using the eCLIP-seq method are typically sequenced using standard SE50 or SE75 conditions on the Illumina HiSeq, NovaSeq, or NextSeq platforms. eCLIP-seq libraries are compatible with paired-end sequencing if desired by the user, however due to the small size of typical eCLIP RNA fragments (~200bp), most fragments are fully sequenced in standard single-end formats.
What is the recommended sequencing depth per sample?
Eclipsebio’s target is 25 million reads per eCLIP-seq dataset. How deeply to sequence an eCLIP-seq dataset is a challenging balance between cost and sufficient read depth to detect true binding events. In an effort to experimentally address this question, an analysis of eCLIP-seq datasets for 150 RNA binding proteins suggested that for 90% of datasets, saturation of peak information occurred at or below 8.5 million reads (See Supplementary Fig. 11 of Van Nostrand EL, et al. A Large-Scale Binding and Functional Map of Human RNA Binding Proteins. Nature (Accepted, in press)
However, we have found that targeting 25 million reads provides better coverage for abundant, broadly binding RNA binding proteins (such as HNRNPs) while still allowing pooling of ~14 eCLIP libraries per standard Illumina HiSeq 4000 lane.
What genomes are available for analysis?
We have the following genomes available for analysis: human (hg38), mouse (mm10), rat (rn6), c. elegans (ce11). Custom species may incur an additional setup fee, including the addition of a viral genome to one of our available species. Contact us at firstname.lastname@example.org for more information.
What are the indices for sequencing Sample-Sheet?
|i7 index name||i7 bases on Sample Sheet||i5 index name||i5 bases bases on Sample Sheet|
Tools + Resources