Ribosome Display

Ribosome display library construction plays a crucial role in identifying high-affinity peptides, cDNAs, and antibodies. At Rx Biosciences, we specialize in constructing and screening ribosome display libraries to support the efficient in vitro selection of binding molecules. Compared to phage and yeast display libraries, ribosome display libraries offer significantly higher complexity—greater than 10¹⁴–10¹⁵. Due to this high complexity, they yield rare, high-affinity clones with picomolar affinities in less time than conventional display methods.

We begin by generating a diverse library of peptides, cDNAs, and antibody variants. Each sequence contains essential elements such as a T7 RNA polymerase promoter, a ribosome binding site, actinomycin/puromycin binding sequences, and RNA stability motifs. These features ensure efficient transcription and promote stable complex formation.

Once the library is prepared, we proceed with in vitro transcription to synthesize RNA transcripts. Next, we perform in vitro translation using cell-free protein lysates (derived from either prokaryotic or eukaryotic sources depending on project requirements), tRNA, an appropriate buffer, and high concentrations of Mg²⁺ ions. The in vitro translation mixture, along with its ribosomes, mRNA, and translated proteins, is then hybridized with the target. As a result, quaternary complexes form—comprising the target, in vitro-translated protein (target binder), its corresponding mRNA, and the ribosomes.

To enhance quaternary complex formation, we maintain the process at 4°C. During the washing step, unbound tertiary complexes are removed. Subsequently, we elute RNA from the bound quaternary complexes and convert it into DNA for further screening.

Through three to four iterative rounds of screening, we enrich and isolate complexes with strong binding affinities. After each round, we extract RNA from the selected complexes. We then reverse transcribe the RNA into cDNA and clone it into appropriate vectors.

To identify the selected ligands, we sequence the cloned DNA and translate the sequences into their corresponding amino acid chains. This final step reveals the binding potential and structural characteristics of the selected molecules.

By following this systematic approach, Rx Biosciences enables researchers to discover novel ligands for therapeutic, diagnostic, and research purposes. Our expertise in ribosome display library construction ensures high-throughput screening with reliable and accurate results. acid sequences. This final step reveals the structure and binding potential of the molecules we’ve selected.

Thanks to this systematic approach, Rx Biosciences helps researchers efficiently discover novel ligands for therapeutic, diagnostic, and research applications. Our expertise in ribosome display library construction ensures accurate, high-throughput screening with reliable results.