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
Descripción
Sumario: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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