Cargando…

BglBrick vectors and datasheets: A synthetic biology platform for gene expression

BACKGROUND: As engineered biological systems become more complex, it is increasingly common to express multiple operons from different plasmids and inducible expression systems within a single host cell. Optimizing such systems often requires screening combinations of origins of replication, express...

Descripción completa

Detalles Bibliográficos
Autores principales: Lee, Taek Soon, Krupa, Rachel A, Zhang, Fuzhong, Hajimorad, Meghdad, Holtz, William J, Prasad, Nilu, Lee, Sung Kuk, Keasling, Jay D
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3189095/
https://www.ncbi.nlm.nih.gov/pubmed/21933410
http://dx.doi.org/10.1186/1754-1611-5-12
_version_ 1782213424855384064
author Lee, Taek Soon
Krupa, Rachel A
Zhang, Fuzhong
Hajimorad, Meghdad
Holtz, William J
Prasad, Nilu
Lee, Sung Kuk
Keasling, Jay D
author_facet Lee, Taek Soon
Krupa, Rachel A
Zhang, Fuzhong
Hajimorad, Meghdad
Holtz, William J
Prasad, Nilu
Lee, Sung Kuk
Keasling, Jay D
author_sort Lee, Taek Soon
collection PubMed
description BACKGROUND: As engineered biological systems become more complex, it is increasingly common to express multiple operons from different plasmids and inducible expression systems within a single host cell. Optimizing such systems often requires screening combinations of origins of replication, expression systems, and antibiotic markers. This procedure is hampered by a lack of quantitative data on how these components behave when more than one origin of replication or expression system are used simultaneously. Additionally, this process can be time consuming as it often requires the creation of new vectors or cloning into existing but disparate vectors. RESULTS: Here, we report the development and characterization of a library of expression vectors compatible with the BglBrick standard (BBF RFC 21). We have designed and constructed 96 BglBrick-compatible plasmids with a combination of replication origins, antibiotic resistance genes, and inducible promoters. These plasmids were characterized over a range of inducer concentrations, in the presence of non-cognate inducer molecules, and with several growth media, and their characteristics were documented in a standard format datasheet. A three plasmid system was used to investigate the impact of multiple origins of replication on plasmid copy number. CONCLUSIONS: The standardized collection of vectors presented here allows the user to rapidly construct and test the expression of genes with various combinations of promoter strength, inducible expression system, copy number, and antibiotic resistance. The quantitative datasheets created for these vectors will increase the predictability of gene expression, especially when multiple plasmids and inducers are utilized.
format Online
Article
Text
id pubmed-3189095
institution National Center for Biotechnology Information
language English
publishDate 2011
publisher BioMed Central
record_format MEDLINE/PubMed
spelling pubmed-31890952011-10-08 BglBrick vectors and datasheets: A synthetic biology platform for gene expression Lee, Taek Soon Krupa, Rachel A Zhang, Fuzhong Hajimorad, Meghdad Holtz, William J Prasad, Nilu Lee, Sung Kuk Keasling, Jay D J Biol Eng Methodology BACKGROUND: As engineered biological systems become more complex, it is increasingly common to express multiple operons from different plasmids and inducible expression systems within a single host cell. Optimizing such systems often requires screening combinations of origins of replication, expression systems, and antibiotic markers. This procedure is hampered by a lack of quantitative data on how these components behave when more than one origin of replication or expression system are used simultaneously. Additionally, this process can be time consuming as it often requires the creation of new vectors or cloning into existing but disparate vectors. RESULTS: Here, we report the development and characterization of a library of expression vectors compatible with the BglBrick standard (BBF RFC 21). We have designed and constructed 96 BglBrick-compatible plasmids with a combination of replication origins, antibiotic resistance genes, and inducible promoters. These plasmids were characterized over a range of inducer concentrations, in the presence of non-cognate inducer molecules, and with several growth media, and their characteristics were documented in a standard format datasheet. A three plasmid system was used to investigate the impact of multiple origins of replication on plasmid copy number. CONCLUSIONS: The standardized collection of vectors presented here allows the user to rapidly construct and test the expression of genes with various combinations of promoter strength, inducible expression system, copy number, and antibiotic resistance. The quantitative datasheets created for these vectors will increase the predictability of gene expression, especially when multiple plasmids and inducers are utilized. BioMed Central 2011-09-20 /pmc/articles/PMC3189095/ /pubmed/21933410 http://dx.doi.org/10.1186/1754-1611-5-12 Text en Copyright ©2011 Lee et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Methodology
Lee, Taek Soon
Krupa, Rachel A
Zhang, Fuzhong
Hajimorad, Meghdad
Holtz, William J
Prasad, Nilu
Lee, Sung Kuk
Keasling, Jay D
BglBrick vectors and datasheets: A synthetic biology platform for gene expression
title BglBrick vectors and datasheets: A synthetic biology platform for gene expression
title_full BglBrick vectors and datasheets: A synthetic biology platform for gene expression
title_fullStr BglBrick vectors and datasheets: A synthetic biology platform for gene expression
title_full_unstemmed BglBrick vectors and datasheets: A synthetic biology platform for gene expression
title_short BglBrick vectors and datasheets: A synthetic biology platform for gene expression
title_sort bglbrick vectors and datasheets: a synthetic biology platform for gene expression
topic Methodology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3189095/
https://www.ncbi.nlm.nih.gov/pubmed/21933410
http://dx.doi.org/10.1186/1754-1611-5-12
work_keys_str_mv AT leetaeksoon bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT kruparachela bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT zhangfuzhong bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT hajimoradmeghdad bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT holtzwilliamj bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT prasadnilu bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT leesungkuk bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression
AT keaslingjayd bglbrickvectorsanddatasheetsasyntheticbiologyplatformforgeneexpression