Leishmania cell-free protein expression system

Kovtun, Oleksiy, Mureev, Sergey, Jung, WoomRam, Kubala, Marta H., Johnston, Wayne and Alexandrov, Kirill (2011) Leishmania cell-free protein expression system. Methods, 55 1: 58-64. doi:10.1016/j.ymeth.2011.06.006


Author Kovtun, Oleksiy
Mureev, Sergey
Jung, WoomRam
Kubala, Marta H.
Johnston, Wayne
Alexandrov, Kirill
Title Leishmania cell-free protein expression system
Formatted title
Leishmania cell-free protein expression system
Journal name Methods   Check publisher's open access policy
ISSN 1046-2023
1095-9130
Publication date 2011-09-01
Sub-type Article (original research)
DOI 10.1016/j.ymeth.2011.06.006
Volume 55
Issue 1
Start page 58
End page 64
Total pages 7
Place of publication Maryland Heights, MO, United States
Publisher Academic Press
Collection year 2012
Language eng
Formatted abstract
Cell-free protein expression is an important tool for a rapid production, engineering and labeling of recombinant proteins. However the complex protocols for preparation of eukaryotic cell-free protein expression systems result in high manufacturing costs and limit their utility. Recently we reported a novel cell-free expression system based on the lysate of a fermentable protozoan Leishmania tarentolae. Herein we describe a protocol for high throughput protein expression using Leishmania cell-free lysate. The protocol combines PCR-based synthesis and engineering of translation templates with a combined transcription-translation system. The protocol is adapted to multiwell plate format and allows translation of large protein libraries. In the presented example we translate in vitro and isolate a nearly complete complement of mammalian Rab GTPases. Further applications and developments of the system are discussed.
Keyword Cell-free protein expression
Leishmania tarentolae
Overlap extension PCR
High throughput protein expression
Rab GTPase
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes Methods in Structural Proteomics

 
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Created: Thu, 13 Oct 2011, 09:51:39 EST by Professor Kirill Alexandrov on behalf of Institute for Molecular Bioscience