Format

Send to

Choose Destination
Methods. 2013 Mar 15;60(1):55-69. doi: 10.1016/j.ymeth.2012.11.004. Epub 2012 Nov 28.

In vitro selection of proteins with desired characteristics using mRNA-display.

Author information

1
Division of Human Genetics, Cincinnati Children's Hospital Medical Center, Cincinnati, OH 45229, USA. alexander.valencia@cchmc.org

Abstract

mRNA-display is an amplification-based, iterative rounds of in vitro protein selection technique that circumvents a number of difficulties associated with yeast two-hybrid and phage display. Because of the covalent linkage between the genotype and the phenotype, mRNA-display provides a powerful means for reading and amplifying a peptide or protein sequence after it has been selected from a library with very high diversity. The purpose of this article is to provide a summary of the field and practical framework of mRNA-display-based selections. We summarize the advantages and limitations of selections using mRNA-display as well as the recent applications, namely, the identification of novel affinity reagents, target-binding partners, and enzyme substrates from synthetic peptide or natural proteome libraries. Practically, we provide a detailed procedure for performing mRNA-display-based selections with the aim of identifying protease substrates and binding partners of a target protein. Furthermore, we describe how to confirm the function of the selected protein sequences by biochemical assays and bioinformatic tools.

PMID:
23201412
PMCID:
PMC3877681
DOI:
10.1016/j.ymeth.2012.11.004
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Elsevier Science Icon for PubMed Central
Loading ...
Support Center