C-Brick: A New Standard for Assembly of Biological Parts Using Cpf1

Abstract

So far, several DNA assembly standards have been developed, enabling scientists to conveniently share and modify characterized DNA parts. However, a majority of the restriction endonucleases used in these standards bear short recognition sites (e.g., 6 bps in BioBrick standard), which are widely distributed and need to be removed before further construction, causing much inconvenience. Although homing endonucleases, which recognize long DNA sequences, can be used for DNA assembly (e.g., iBrick standard), long scars will be left between parts, limiting their application. Here, we introduce a new DNA assembly standard, namely C-Brick, which employs the newly identified class 2 type V CRISPR-Cas systems protein Cpf1 endonuclease. C-Brick integrates both advantages of long recognition sites and short scars. With C-Brick standard, three chromoprotein cassettes were assembled and further expressed in Escherichia coli, producing colorful pigments. Moreover, C-Brick standard is also partially compatible with the BglBrick and BioBrick standards.