United States Patent 6,620,587
Taussig , et al. September 16, 2003
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Ribosome complexes as selection particles for in vitro display and evolution of proteins
Abstract
The invention provides a method of displaying nascent proteins or peptides as complexes with eukaryotic ribosomes and the mRNA encoding the protein or peptide following transcription and translation in vitro, of further selecting complexes carrying a particular nascent protein or peptide by means of binding to a ligand, antigen or antibody, and of subsequently recovering the genetic information encoding the protein or peptide from the selected ribosome complex by reverse transcription and polymerase chain reaction (RT-PCR). The RT-PCR recovery step is carried out directly on the intact ribosome complex, without prior dissociation to release the mRNA, thus contributing to maximal efficiency and sensitivity. The steps of display, selection and recovery can be repeated in consecutive cycles. The method is exemplified using single-chain antibody constructs as antibody-ribosome-mRNA complexes (ARMs).
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Inventors: Taussig; Michael John (Cambridge, GB); He; Mingyue (Cambridge, GB)
Assignee: Discerna Limited (Cambridge, GB)
Appl. No.: 424712
Filed: November 29, 1999
PCT Filed: May 28, 1998
PCT NO: PCT/GB98/01564
PCT PUB.NO.: WO98/54312
PCT PUB. Date: December 3, 1998
Foreign Application Priority Data
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May 28, 1997[GB] 9710829
Nov 26, 1997[GB] 9724850
Feb 28, 1998[GB] 9804195
Current U.S. Class: 435/7.1; 435/4; 435/6; 435/69.1; 435/69.6; 435/DIG.3; 435/DIG.15; 435/DIG.17; 436/501; 436/547
Intern'l Class: G01N 033/53; G01N 033/566; C12Q 001/00
Field of Search: 435/6,7.1,4,69.1,69.6,DIG. 3,DIG. 15,DIG. 17 436/501,547 |
