Cargando…

Producing molecular biology reagents without purification

We recently developed ‘cellular’ reagents–lyophilized bacteria overexpressing proteins of interest–that can replace commercial pure enzymes in typical diagnostic and molecular biology reactions. To make cellular reagent technology widely accessible and amenable to local production with minimal instr...

Descripción completa

Detalles Bibliográficos
Autores principales: Bhadra, Sanchita, Nguyen, Vylan, Torres, Jose-Angel, Kar, Shaunak, Fadanka, Stéphane, Gandini, Chiara, Akligoh, Harry, Paik, Inyup, Maranhao, Andre C., Molloy, Jenny, Ellington, Andrew D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8168896/
https://www.ncbi.nlm.nih.gov/pubmed/34061896
http://dx.doi.org/10.1371/journal.pone.0252507
_version_ 1783701952330203136
author Bhadra, Sanchita
Nguyen, Vylan
Torres, Jose-Angel
Kar, Shaunak
Fadanka, Stéphane
Gandini, Chiara
Akligoh, Harry
Paik, Inyup
Maranhao, Andre C.
Molloy, Jenny
Ellington, Andrew D.
author_facet Bhadra, Sanchita
Nguyen, Vylan
Torres, Jose-Angel
Kar, Shaunak
Fadanka, Stéphane
Gandini, Chiara
Akligoh, Harry
Paik, Inyup
Maranhao, Andre C.
Molloy, Jenny
Ellington, Andrew D.
author_sort Bhadra, Sanchita
collection PubMed
description We recently developed ‘cellular’ reagents–lyophilized bacteria overexpressing proteins of interest–that can replace commercial pure enzymes in typical diagnostic and molecular biology reactions. To make cellular reagent technology widely accessible and amenable to local production with minimal instrumentation, we now report a significantly simplified method for preparing cellular reagents that requires only a common bacterial incubator to grow and subsequently dry enzyme-expressing bacteria at 37°C with the aid of inexpensive chemical desiccants. We demonstrate application of such dried cellular reagents in common molecular and synthetic biology processes, such as PCR, qPCR, reverse transcription, isothermal amplification, and Golden Gate DNA assembly, in building easy-to-use testing kits, and in rapid reagent production for meeting extraordinary diagnostic demands such as those being faced in the ongoing SARS-CoV-2 pandemic. Furthermore, we demonstrate feasibility of local production by successfully implementing this minimized procedure and preparing cellular reagents in several countries, including the United Kingdom, Cameroon, and Ghana. Our results demonstrate possibilities for readily scalable local and distributed reagent production, and further instantiate the opportunities available via synthetic biology in general.
format Online
Article
Text
id pubmed-8168896
institution National Center for Biotechnology Information
language English
publishDate 2021
publisher Public Library of Science
record_format MEDLINE/PubMed
spelling pubmed-81688962021-06-11 Producing molecular biology reagents without purification Bhadra, Sanchita Nguyen, Vylan Torres, Jose-Angel Kar, Shaunak Fadanka, Stéphane Gandini, Chiara Akligoh, Harry Paik, Inyup Maranhao, Andre C. Molloy, Jenny Ellington, Andrew D. PLoS One Research Article We recently developed ‘cellular’ reagents–lyophilized bacteria overexpressing proteins of interest–that can replace commercial pure enzymes in typical diagnostic and molecular biology reactions. To make cellular reagent technology widely accessible and amenable to local production with minimal instrumentation, we now report a significantly simplified method for preparing cellular reagents that requires only a common bacterial incubator to grow and subsequently dry enzyme-expressing bacteria at 37°C with the aid of inexpensive chemical desiccants. We demonstrate application of such dried cellular reagents in common molecular and synthetic biology processes, such as PCR, qPCR, reverse transcription, isothermal amplification, and Golden Gate DNA assembly, in building easy-to-use testing kits, and in rapid reagent production for meeting extraordinary diagnostic demands such as those being faced in the ongoing SARS-CoV-2 pandemic. Furthermore, we demonstrate feasibility of local production by successfully implementing this minimized procedure and preparing cellular reagents in several countries, including the United Kingdom, Cameroon, and Ghana. Our results demonstrate possibilities for readily scalable local and distributed reagent production, and further instantiate the opportunities available via synthetic biology in general. Public Library of Science 2021-06-01 /pmc/articles/PMC8168896/ /pubmed/34061896 http://dx.doi.org/10.1371/journal.pone.0252507 Text en © 2021 Bhadra et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Bhadra, Sanchita
Nguyen, Vylan
Torres, Jose-Angel
Kar, Shaunak
Fadanka, Stéphane
Gandini, Chiara
Akligoh, Harry
Paik, Inyup
Maranhao, Andre C.
Molloy, Jenny
Ellington, Andrew D.
Producing molecular biology reagents without purification
title Producing molecular biology reagents without purification
title_full Producing molecular biology reagents without purification
title_fullStr Producing molecular biology reagents without purification
title_full_unstemmed Producing molecular biology reagents without purification
title_short Producing molecular biology reagents without purification
title_sort producing molecular biology reagents without purification
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8168896/
https://www.ncbi.nlm.nih.gov/pubmed/34061896
http://dx.doi.org/10.1371/journal.pone.0252507
work_keys_str_mv AT bhadrasanchita producingmolecularbiologyreagentswithoutpurification
AT nguyenvylan producingmolecularbiologyreagentswithoutpurification
AT torresjoseangel producingmolecularbiologyreagentswithoutpurification
AT karshaunak producingmolecularbiologyreagentswithoutpurification
AT fadankastephane producingmolecularbiologyreagentswithoutpurification
AT gandinichiara producingmolecularbiologyreagentswithoutpurification
AT akligohharry producingmolecularbiologyreagentswithoutpurification
AT paikinyup producingmolecularbiologyreagentswithoutpurification
AT maranhaoandrec producingmolecularbiologyreagentswithoutpurification
AT molloyjenny producingmolecularbiologyreagentswithoutpurification
AT ellingtonandrewd producingmolecularbiologyreagentswithoutpurification