Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits

A modified BioBrick™ assembly method was developed with higher fidelity than current protocols. The method utilizes a PCR reaction with a standard primer set to amplify the inserted part. Background colonies are reduced by a combination of dephosphorylation and digestion with DpnI restriction endonu...

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Detalles Bibliográficos
Autores principales: Speer, Michael A, Richard, Tom L
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2011
Materias:
Methodology
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3287150/
https://www.ncbi.nlm.nih.gov/pubmed/22176971
http://dx.doi.org/10.1186/1754-1611-5-17
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author Speer, Michael A
Richard, Tom L
author_facet Speer, Michael A
Richard, Tom L
author_sort Speer, Michael A
collection PubMed
description A modified BioBrick™ assembly method was developed with higher fidelity than current protocols. The method utilizes a PCR reaction with a standard primer set to amplify the inserted part. Background colonies are reduced by a combination of dephosphorylation and digestion with DpnI restriction endonuclease to reduce vector and insert background respectively. The molar ratio of the insert to vector in the ligation was also optimized, with the accuracy of the transformed construct approaching 100%.
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spelling pubmed-32871502012-02-28 Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits Speer, Michael A Richard, Tom L J Biol Eng Methodology A modified BioBrick™ assembly method was developed with higher fidelity than current protocols. The method utilizes a PCR reaction with a standard primer set to amplify the inserted part. Background colonies are reduced by a combination of dephosphorylation and digestion with DpnI restriction endonuclease to reduce vector and insert background respectively. The molar ratio of the insert to vector in the ligation was also optimized, with the accuracy of the transformed construct approaching 100%. BioMed Central 2011-12-16 /pmc/articles/PMC3287150/ /pubmed/22176971 http://dx.doi.org/10.1186/1754-1611-5-17 Text en Copyright ©2011 Speer and Richard; 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
Speer, Michael A
Richard, Tom L
Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits
title Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits
title_full Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits
title_fullStr Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits
title_full_unstemmed Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits
title_short Amplified insert assembly: an optimized approach to standard assembly of BioBrick(TM) genetic circuits
title_sort amplified insert assembly: an optimized approach to standard assembly of biobrick(tm) genetic circuits
topic Methodology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3287150/
https://www.ncbi.nlm.nih.gov/pubmed/22176971
http://dx.doi.org/10.1186/1754-1611-5-17
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