Abstract
Yeast (Saccharomyces cerevisiae) as a model organism has long been established to study various aspects of eukaryotic cellular and molecular biology. Cell-free lysates prepared from different yeast strains have been used as a powerful tool to study eukaryotic protein expression in vitro. Recently, we established a yeast cell-free lysate system for in vitro translation long and short L1 capsid gene mRNAs of human papillomavirus type 58. We were able to significantly improve the translation efficiencies of the viral mRNAs in the established system by optimizing the concentrations of potassium and magnesium and controlling the physiological status of the yeast cells used for lysate preparation. We proved that a single specific amino acid can be rate limiting for translation of a target mRNA. Here, we describe the materials, method, and technique used for the development of an efficient yeast cell-free translation system.
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Acknowledgments
This work was partially supported by a National Health and Medical Research Council of Australia Industry Research Fellowship (301256 to KNZ) and grants from Cancer Council of Queensland (401623 to KNZ).
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Wang, X., Zhao, L., Zhao, KN. (2014). An Optimized Yeast Cell-Free Lysate System for In Vitro Translation of Human Virus mRNA. In: Alexandrov, K., Johnston, W. (eds) Cell-Free Protein Synthesis. Methods in Molecular Biology, vol 1118. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-782-2_14
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DOI: https://doi.org/10.1007/978-1-62703-782-2_14
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Publisher Name: Humana Press, Totowa, NJ
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