RNA lifecycle control is key to therapeutics, with RPIs orchestrating gene expression. RIP-Seq combines antibody-based enrichment with NGS to map RPI landscapes, delivering high-resolution functional data. Creative Biolabs' RIP-Seq service bridges genomic data and protein regulation, offering unbiased, high-resolution insights to map interactomes and accelerate therapeutic discovery.
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The human genome encodes hundreds of RBPs, each possessing unique RNA-binding specificity. These RBPs are often dysregulated in disease, making them crucial diagnostic indicators and therapeutic targets. Standard RNA sequencing reveals which transcripts are present, but cannot indicate if a specific protein is actively controlling those transcripts. RIP-Seq overcomes this limitation by capturing the actual RNP complexes, allowing us to understand the functional regulatory network at the post-transcriptional level.
The core mechanism is rooted in immunology and sequencing:
We offer a comprehensive and modular workflow, designed for full transparency and optimized for high-quality, reproducible results. This detailed process ensures clear communication and reliable outcomes, making it ideal for visualization as a project flowchart.
RBP-specific antibody (validated by Western Blot), viable cell lines, fresh/frozen tissue samples, or detailed experimental condition information (e.g., cell stimulation, drug treatment protocol).
Define experimental groups, biological replicates, and sequencing depth for statistical power, ensuring final data addresses research questions.
Lyse cells with non-denaturing buffers to maintain native RPI structure.
Pull down target RBP using high-quality antibody-coupled magnetic beads.
Separate RNA from RBP complexes (via Proteinase K) and purify.
Convert purified RNA to cDNA and tag with Illumina-compatible sequencing adaptors. Expected Outcome: High-quality sequencing library ready for deep sequencing.
Conduct high-throughput sequencing of libraries. Perform quality control on raw reads, map to the reference genome, and use peak calling to identify significant RBP binding sites.
Raw Sequence Data (FASTQ files) and alignment reports; Mapped RNA Targets Report (gene lists, p-values, fold enrichment); Functional Annotation Analysis (pathway enrichment, GO analysis of targeted transcripts).
The typical timeframe for the RIP-Seq service ranges from 6 to 10 weeks, depending on the sample complexity (e.g., primary tissue vs. established cell line) and the required depth of the bioinformatic analysis.
Creative Biolabs doesn't just run assays; we deliver custom-designed solutions that integrate directly into your most complex therapeutic pipelines. Our dedication to scientific rigor and customized service ensures your RBP mapping project yields high-impact, definitive results.
Bespoke Assay Development
Full protocol customization based on your specific RBP's characteristics (e.g., native vs. cross-linked RIP, optimized lysis buffers, and co-factor analysis) to ensure optimal binding capture.
Validated Antibody Assurance
Access to Creative Biolabs' extensive, pre-screened library of RBP-specific antibodies, minimizing batch-to-batch variation and guaranteeing superior immunoprecipitation efficiency and specificity.
Advanced Bioinformatic Interpretation
Utilization of in-house, proprietary peak-calling and differential binding analysis algorithms to accurately delineate genuine RPIs, even in low-abundance transcripts.
Scalable Throughput Capacity
High-capacity NGS facilities enable the parallel processing of dozens of samples for large-scale comparative studies, such as analyzing RPI shifts across different disease states or time points.
End-to-End Project Management
A dedicated scientific expert guides your project from initial sample submission and quality control to the final delivery of publication-ready figures and comprehensive reports.
Rigorous Quality Verification
Implementation of Quality-by-Design (QbD) principles across all stages, including pre-RIP input verification and post-IP qPCR validation, ensuring the reliability of your final dataset.
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While both methods map RBP targets, RIP-Seq (non-crosslinked) is generally faster, easier, and better suited for identifying the full complement of RNAs associated with an RBP, providing a high-confidence list of targets. CLIP-Seq requires UV cross-linking to create a covalent bond, offering higher resolution down to the single-nucleotide binding site, but it is technically much more challenging and sensitive to artifacts. We typically recommend RIP-Seq for initial target discovery and broad mapping. If you require single-nucleotide resolution, we are happy to discuss our advanced CLIP-Seq services.
The single most critical factor is the quality and specificity of the antibody used for the immunoprecipitation. A poor antibody yields high background noise and inaccurate results. Creative Biolabs mitigates this risk by offering a pre-validated antibody selection service or rigorously testing client-provided antibodies prior to project launch, ensuring maximum confidence in the final data.
Yes, our protocol is optimized to handle a diverse range of materials, including challenging low-input or highly degraded samples like primary tissues. We provide strict guidelines and preservation techniques during the project design phase to ensure sample integrity is maintained, maximizing the likelihood of successful RNP complex isolation.
Not at all. Our deliverables include the raw data, but the core output is the Functional Annotation Analysis—a clearly summarized, publication-ready report. This report details the enriched gene lists, pathway diagrams, and biological interpretations, making the data immediately accessible and actionable for your team without needing specialized bioinformatics personnel.
We always recommend planning for appropriate negative controls (e.g., IgG controls) to accurately assess non-specific background binding. Furthermore, the expression level of your target RBP is important; highly expressed proteins yield cleaner results. We offer pre-project RBP expression validation to confirm feasibility before you invest fully in the sequencing run.
Creative Biolabs' RIP-Seq based RNA-Protein Interaction Analysis Service delivers the functional insights necessary to accelerate drug discovery in the age of RNA therapeutics. By leveraging proprietary protocols, a validated antibody library, and advanced bioinformatic interpretation, we transform complex regulatory data into clear therapeutic roadmaps.
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