cDNA - protein fusions: covalent protein - gene conjugates for the in vitro selection of peptides and proteins.
We report a method for the synthesis of covalent cDNA-protein fusions for protein display applications. A branched mRNA template was developed which carries a peptidyl acceptor and a reverse transcription primer at the 3'-end. Translation in vitro followed by reverse transcription produced a protein covalently bonded to its encoding cDNA. Both single- and double-stranded cDNA-protein fusions were prepared. cDNA-protein fusions are stable in alkali and resistant to ribonucleases. Their simple preparation and their resistance towards degradation should make cDNA-protein fusions a useful tool for the in vitro selection and evolution of high affinity ligands from large libraries of polypeptides.
Kurz
M
M
Phylos, Inc., 128 Spring Street, Lexington, MA 02421, USA.
Gu
K
K
Al-Gawari
A
A
Lohse
P A
PA
D019677:Artificial Gene Fusion / Q000379:methods*; D001483:Base Sequence; D002478:Cells, Cultured; D003470:Culture Media; D004247:DNA / Q000737:chemistry; D018076:DNA, Complementary / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry; D006801:Humans; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes / Q000737:chemistry; D010455:Peptides / Q000737:chemistry*; D011233:Precipitin Tests; D011506:Proteins / Q000737:chemistry*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013698:Templates, Genetic; D014158:Transcription, Genetic
D003470:Culture Media; D018076:DNA, Complementary; D004277:DNA, Single-Stranded; D015345:Oligonucleotide Probes; D010455:Peptides; D011506:Proteins; D012333:RNA, Messenger; D004247:DNA