Services + Kits

RBP-eCLIP

A robust and reproducible framework to map RBP binding sites on RNAs transcriptome-wide

The RBP-eCLIP Service

Turnaround Time
6-8 Weeks (Project Dependent)

*Requires Validated Antibody
See List of Pre-Validated Antibodies

Starting Material
Cells: 20 M; UV Crosslinked
Tissue: 80 mg; Non-Crosslinked; Flash Frozen

Start your RBP-eCLIP Service Project
Connect With An Eclipsebio RNA Expert

Bioinformatics

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.

EXAMPLE REPORT

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.

RBP-eCLIP Kit

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)

Full Description

Product Highlights
The RBP-eCLIP kit is a simplified and robust library preparation method to identify your RNA-binding proteins targets.

Convenient
Eliminate time-consuming prep work by ordering one kit with 30+ validated reagents. Start immediately and generate eCLIP libraries in 5 days.

High performance
Published method and protocol improvements increase workflow efficiency and identify true binding sites

Reproducible
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

Protocols

RBP-eCLIP Protocol v2.01R

RBP-eCLIP Protocol v1.01R (Discontinued Catalog #ECK001)

FAQs

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 techsupport@eclipsebio.com for more information.

What are the indices for sequencing Sample-Sheet?