Amplified insert assembly: an optimized approach to standard assembly of BioBrickTM genetic circuits

Journal of Biological Engineering

Tom L Richard¹  Michael A Speer¹ 

[1] Department of Agricultural and Biological Engineering, The Pennsylvania State University, 249 Agricultural Engineering Bldg., University Park, PA 16802, USA

关键词: DpnI;    assembly;    vector;    insert;    amplification;    PCR;    Synthetic biology;    BioBrick;   

Others  :  813551
DOI  :  10.1186/1754-1611-5-17

 received in 2011-05-03, accepted in 2011-12-16,  发布年份 2011

【 摘 要 】

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%.

【 授权许可】

   
2011 Speer and Richard; licensee BioMed Central Ltd.

【 图 表 】

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