Difference between revisions of "Part:BBa K2020042"

Revision as of 20:49, 13 October 2016

Mj-tRNA with amber anticodon for incorporating ncAA in E.coli

For incorporating unnatural amino acids into a protein, a orthogonal tRNA:Synthetase-pair is needed which does not crossreact with the cognate tRNA:Synthetase-pairs. This tRNA can be assembled with a variety of synthetases into a plasmid to incorporate ncAA in E.coli in response to an amber stop codon

Usage and Biology

This tRNA derives from the wild type tyrosyl Methanococcus janaschii tRNA:Synthetase pair. It was proven to not crossreact with the cognate E.coli tRNA:synthetase-pairs (A Genetically Encoded Photocaged Tyrosine - Schultz et al, 2006).

The tRNA is used together with a tRNA-Synthetase. It has been proven to work with (enter links for parts)

• Y-RS
• oNBY-RS
• AzF
• CN-F synthetase
• Iodo-Y synthetase
• 5HT-P synthetase
• Nitro-Y synthetase
• Amino-Y synthetase

by [http://2014.igem.org/Team:Austin_Texas iGEM-Team Austin, Texas 2014].

[http://2016.igem.org/Team:Aachen iGEM-Team Aachen 2016] used the tRNA to successfully incorporate tyrosine, oNB-Y and DMNB-S in E.coli BL21 DE3 gold with their newly designed DMNBS-RS.

This tRNA has an amber anticodon for incorporating the ncAA in response to an amber codon. It has been used previously in an amberless E.coli strain as well as BL21 DE3 gold. Application of the tRNA is either the incorporation of the ncAA into a protein or usage with a reporter plasmid e.g. pRXG.

Sequence and Features

Functional Parameters