Skip to main content
bioRxiv
  • Home
  • About
  • Submit
  • ALERTS / RSS
Advanced Search
Contradictory Results

“Cheater” particles render the VEGAS platform unsuitable for mammalian directed evolution

View ORCID ProfileChristopher E. Denes, View ORCID ProfileAlexander J. Cole, Minh Thuan Nguyen Tran, View ORCID ProfileMohd Khairul Nizam Mohd Khalid, View ORCID ProfileAlex W. Hewitt, View ORCID ProfileDaniel Hesselson, View ORCID ProfileG. Gregory Neely
doi: https://doi.org/10.1101/2022.06.07.495067
Christopher E. Denes
1The Dr. John and Anne Chong Lab for Functional Genomics, Charles Perkins Centre and School of Life & Environmental Sciences, The University of Sydney, Sydney, NSW, 2006, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Christopher E. Denes
Alexander J. Cole
2Centenary Institute and Faculty of Medicine and Health, The University of Sydney, Sydney, NSW, 2006, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Alexander J. Cole
Minh Thuan Nguyen Tran
3Menzies Institute for Medical Research, School of Medicine, University of Tasmania, Tasmania, 7000, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Mohd Khairul Nizam Mohd Khalid
3Menzies Institute for Medical Research, School of Medicine, University of Tasmania, Tasmania, 7000, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Mohd Khairul Nizam Mohd Khalid
Alex W. Hewitt
3Menzies Institute for Medical Research, School of Medicine, University of Tasmania, Tasmania, 7000, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Alex W. Hewitt
Daniel Hesselson
2Centenary Institute and Faculty of Medicine and Health, The University of Sydney, Sydney, NSW, 2006, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Daniel Hesselson
  • For correspondence: d.hesselson@centenary.org.au greg.neely@sydney.edu.au
G. Gregory Neely
1The Dr. John and Anne Chong Lab for Functional Genomics, Charles Perkins Centre and School of Life & Environmental Sciences, The University of Sydney, Sydney, NSW, 2006, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for G. Gregory Neely
  • For correspondence: d.hesselson@centenary.org.au greg.neely@sydney.edu.au
  • Abstract
  • Full Text
  • Info/History
  • Metrics
  • Preview PDF
Loading

Abstract

Directed evolution uses cycles of gene diversification and selection to generate proteins with novel properties. While traditionally directed evolution is performed in prokaryotic systems, recently a mammalian directed evolution system (viral evolution of genetically actuating sequences, or "VEGAS") has been described. Here we report that the VEGAS system has major limitations precluding its use for directed evolution. The primary technical issue with the VEGAS system is an immediate contamination with "cheater" particles that bypass directed evolution circuits. By sequencing we find these cheater particles contain Sindbis structural genes instead of the intended directed evolution target transgene. These cheaters outcompete the VEGAS transgenes within 2 rounds of transduction but cannot themselves activate synthetic circuits that drive expression of Sindbis structural genes, preventing directed evolution campaigns. Similar results have been obtained in independent labs. Taken together, the VEGAS system does not work as described and, without significant redesign to suppress cheaters, cannot be used for mammalian directed evolution campaigns.

Competing Interest Statement

The authors have declared no competing interest.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. All rights reserved. No reuse allowed without permission.
Back to top
PreviousNext
Posted June 21, 2022.
Download PDF
Email

Thank you for your interest in spreading the word about bioRxiv.

NOTE: Your email address is requested solely to identify you as the sender of this article.

Enter multiple addresses on separate lines or separate them with commas.
“Cheater” particles render the VEGAS platform unsuitable for mammalian directed evolution
(Your Name) has forwarded a page to you from bioRxiv
(Your Name) thought you would like to see this page from the bioRxiv website.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
“Cheater” particles render the VEGAS platform unsuitable for mammalian directed evolution
Christopher E. Denes, Alexander J. Cole, Minh Thuan Nguyen Tran, Mohd Khairul Nizam Mohd Khalid, Alex W. Hewitt, Daniel Hesselson, G. Gregory Neely
bioRxiv 2022.06.07.495067; doi: https://doi.org/10.1101/2022.06.07.495067
Digg logo Reddit logo Twitter logo Facebook logo Google logo LinkedIn logo Mendeley logo
Citation Tools
“Cheater” particles render the VEGAS platform unsuitable for mammalian directed evolution
Christopher E. Denes, Alexander J. Cole, Minh Thuan Nguyen Tran, Mohd Khairul Nizam Mohd Khalid, Alex W. Hewitt, Daniel Hesselson, G. Gregory Neely
bioRxiv 2022.06.07.495067; doi: https://doi.org/10.1101/2022.06.07.495067

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
  • Tweet Widget
  • Facebook Like
  • Google Plus One

Subject Area

  • Synthetic Biology
Subject Areas
All Articles
  • Animal Behavior and Cognition (3586)
  • Biochemistry (7544)
  • Bioengineering (5495)
  • Bioinformatics (20729)
  • Biophysics (10294)
  • Cancer Biology (7950)
  • Cell Biology (11609)
  • Clinical Trials (138)
  • Developmental Biology (6586)
  • Ecology (10166)
  • Epidemiology (2065)
  • Evolutionary Biology (13578)
  • Genetics (9520)
  • Genomics (12817)
  • Immunology (7905)
  • Microbiology (19503)
  • Molecular Biology (7640)
  • Neuroscience (41980)
  • Paleontology (307)
  • Pathology (1254)
  • Pharmacology and Toxicology (2192)
  • Physiology (3259)
  • Plant Biology (7018)
  • Scientific Communication and Education (1293)
  • Synthetic Biology (1947)
  • Systems Biology (5418)
  • Zoology (1113)