text: Although there are many other molecular display technologies, such as phage display, bacterial display, yeast display, and ribosome display, mRNA display technology has many advantages over the others. The first three biological display libraries listed have polypeptides or proteins expressed on the respective microorganism’s surface and the accompanying coding information for each polypeptide or protein is retrievable from the microorganism’s genome. However, the library size for these three in vivo display systems is limited by the transformation efficiency of each organism. For example, the library size for phage and bacterial display is limited to 1-10 × 10^9 different members. The library size for yeast display is even smaller. Moreover, these cell-based display system only allow the screening and enrichment of peptides/proteins containing natural amino acids. In contrast, mRNA display and ribosome display are in vitro selection methods. They allow a library size as large as 10^15 different members. The large library size increases the probability to select very rare sequences, and also improves the diversity of the selected sequences. In addition, in vitro selection methods remove unwanted selection pressure, such as poor protein expression, and rapid protein degradation, which may reduce the diversity of the selected sequences. Finally, in vitro selection methods allow the application of in vitro mutagenesis and recombination techniques throughout the selection process.
  Although both ribosome display and mRNA display are in vitro selection methods, mRNA display has some advantage over the ribosome display technology. mRNA display utilizes covalent mRNA-polypeptide complexes linked through puromycin; whereas, ribosome display utilizes stalled, noncovalent ribosome-mRNA-polypeptide complexes. For ribosome display, selection stringency is limited to keep ribosome-mRNA-polypeptide in a complex because of the noncovalent ribosome-mRNA-polypeptide complexes. This may cause difficulties in reducing background binding during the selection cycle. Also, the polypeptides under selection in a ribosome display system are attached to an enormous rRNA-protein complex, a ribosome, which has a molecular weight of more than 2,000,000 Da. There might be some unpredictable interaction between the selection target and the ribosome, and this may lead to a loss of potential binders during the selection cycle. In contrast, the puromycin DNA spacer linker used in mRNA display technology is much smaller comparing to a ribosome. This linker may have less chance to interact with an immobilized selection target. Thus, mRNA display technology is more likely to give less biased results.