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Improvement of a puromycin-linker to extend the selection target varieties in cDNA display method

Paper ID Volume ID Publish Year Pages File Format Full-Text
23627 43459 2012 4 PDF Available
Title
Improvement of a puromycin-linker to extend the selection target varieties in cDNA display method
Abstract

cDNA display using a puromycin-linker to covalently bridge a protein and its coding cDNA is a stable and efficient in vitro protein selection method. The optimal design of the often-used puromycin-linker is vital for effective selection. In this report, an improved puromycin-linker containing deoxyinosine bases as cleavage sites, which are recognized by endonuclease V, was introduced to extend the variety of the selection targets to molecules such as RNA. The introduction of this linker enables efficient in vitro protein selection without contamination from RNase T1, which is used for the conventional linker containing ribonucleotide G bases. In addition, mRNA–protein fusion efficiency was found to not depend on the length of the flexible poly (ethylene glycol) (PEG) region of the linker. These findings will allow practical and easy-to-use in vitro protein selection by cDNA display.

► A puromycin-linker containing deoxyinosine bases as cleavage sites was developed. ► The performance of the new linker is equal to the previous linker using RNase T1. ► The linker enables in vitro protein selection without contamination by RNase. ► The length of the polyethylene glycol region of the linker can be flexible. ► These findings will extend the variety of the selection targets in cDNA display method.

Keywords
In vitro selection; mRNA/cDNA display; Puromycin; Directed evolution; Protein engineering
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Improvement of a puromycin-linker to extend the selection target varieties in cDNA display method
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 162, Issues 2–3, 31 December 2012, Pages 299–302
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us