Cross-linking and Analysis of cDNA by High Throughput Sequencing

Cross-linking and immunoprecipitation (CLIP) is a denatured method used extensively to study RNA-protein interactions in vivo. Cross-linking and analysis of cDNA (CRAC) is an advanced CLIP approach to identify binding sites of RNA-binding proteins by high throughput sequencing. Lifeasible is a biotechnology company with rich experience in RNA-protein interactions analysis. We will be your trust partner to perform the CRAC assay.

CRAC relays on the fact that the binding of the protein to the RNA causes transcription errors (deletions or substitutions), which makes it possible to map the protein/RNA binding site at single nucleotide resolution upon high throughput sequencing. Like other CLIP methods, CRAC also begins with UV cross-linking of the RNA-protein complex and is immunoprecipitated using a specific antibody to the protein-of-interest. Following with separation of RNA-protein complexes by SDS-PAGE and membrane transfer, cDNA is synthesized and is sequenced. Cross-linking induced mutation sites (CIMS) can be used to map the binding sites where proteins directly interact with RNA (Figure 1).

We are dedicated to offering professional and customized one-stop CRAC services (Figure 1), including:

• UV treatment
• Cell lysis
• Immunoprecipitation
• Nonspecific RNA binding removal by washing
• 3’-RNA adaptor ligation
• Proteinase K digestion
• 5’-RNA adaptor ligation
• Reverse transcription
• Sequencing of cDNA
• Formal report and data interpretation

We are determined to provide the optimal protocol for our customers to guarantee each project’s success. We have profound expertise in RNA-protein interaction research and are willing to share our expertise with our customers to facilitate their brilliant studies. Our customer-centered, highly efficient services will ensure the quality of your results and save you time and money. Welcome to contact us for more information. We will be more than happy to help you.