Cell-Free Gene Expression: Methods and Protocols

Part I: Tools for Cell-Free Expression Systems

1. Best Practices for DNA Template Preparation towards Improved Reproducibility in Cell-Free Protein Production

Eugenia F. Romantseva, Drew S. Tack, Nina Alperovich, David Ross, and Elizabeth A. Strychalski

2. Simple Extract Preparation Methods for E. coli-Based Cell-Free Expression

Alissa C. Mullin, Taylor Slouka, and Javin P. Oza

3. Preparation and Screening of Cell-Free Extract from Non-Growing Escherichia coli A19 Cells

Florian Hiering, Jurek Failmezger, and Martin Siemann-Herzberg

4. Cell-Free Protein Synthesis Using Pichia pastoris

Alex J. Spice, Rochelle Aw, and Karen M. Polizzi

5. A Streptomyces-Based Cell-Free Protein Synthesis System for High-Level Protein Expression

Huiling Xu, Wan-Qiu Liu, and Jian Li

6. In Vitro Reconstitution Platforms of Mammalian Cell-Free Expressed Membrane Proteins

Hossein Moghimianavval, Yen-Yu Hsu, Alessandro Groaz, and Allen P. Liu

7. High-Throughput Experimentation Using Cell-Free Protein Synthesis Systems

Conary Meyer, Chuqing Zhou, Zecong Fang, Marjorie L. Longo, Tingrui Pan, and Cheemeng Tan

8. Cell-Free Gene Expression from DNA Brushes

Michael Levy, Ohad Vonshak, Yiftach Divon, Ferdinand Greiss, Noa Avidan, Shirley S. Daube, and Roy H. Bar-Ziv

9. Efficient and Precise Protein Synthesis in a Cell-Free System Using a Set of In Vitro Transcribed tRNAs with Nucleotide Modifications

Kazuaki Amikura, Keita Hibi, and Yoshihiro Shimizu

10. Measurement of Transcription, Translation, and Other Enzymatic Processes during Cell-Free Expression Using PERSIA

Scott Wick and Peter A. Carr

Part II: Applications of Cell-Free Expression Systems

11. Cell-Free Noncanonical Redox Cofactor Systems

William B. Black and Han Li

12. Cell-Free Protein Synthesis for High-Throughput Biosynthetic Pathway Prototyping

Blake J. Rasor, Bastian Vögeli, Michael C. Jewett, and Ashty S. Karim

13. Metabolomics Analysis of Cell-Free Expression Systems Using Gas Chromatography-Mass Spectrometry

April M. Miguez, Yan Zhang, and Mark P. Styczynski

14. Liposome Preparation by 3D-Printed Microcapillary-Based Apparatus

Orion M. Venero, Wakana Sato, Joseph M. Heili, Christopher Deich, and Katarzyna P. Adamala

15. Microfluidic Production of Porous Polymer Cell-Mimics Capable of Gene Expression

Imre Banlaki, François-Xavier Lehr, and Henrike Niederholtmeyer