Restriction enzymes and Gibson Assembly® cloning reagents were obtained from New England Biolabs and polymerases were obtained from Life Technologies. TOP10 competent cells and InsP6 (also known as phytic acid) were obtained from Sigma-Aldrich. Other common chemicals, antibiotics, and growth media components were from Sigma-Aldrich or Thermo Fisher.
Bacterial expression vectors for production of recombinant proteins pMCSG7, pMCSG53, and pMCSG58 were obtained from Dr. Andrzej Joachimiak of the Midwest Center for Structural Genomics (MCSG). The vector pMCSG7 consists of a LIC site and encodes for an N-terminal TEV cleavable 6xHis-tagged protein [15 (link)]. The vector pMCSG53 is an upgraded version of pMCSG7 that also carries the rare codon tRNA genes argU and ileX [16 (link)]. The vector pMCSG58 is similar to pMCSG53 except it encodes for a C-terminal 6xHis-tagged protein [16 (link)]. Proteins were expressed in kanamycin-resistant BL21(DE3)MAGIC E. coli strain. The pMagic plasmid also provides a second copy of the rare codon tRNA gene argU ([17 (link)] and A. Joachimiak, personal communication).
Bacterial expression vectors for production of recombinant proteins pMCSG7, pMCSG53, and pMCSG58 were obtained from Dr. Andrzej Joachimiak of the Midwest Center for Structural Genomics (MCSG). The vector pMCSG7 consists of a LIC site and encodes for an N-terminal TEV cleavable 6xHis-tagged protein [15 (link)]. The vector pMCSG53 is an upgraded version of pMCSG7 that also carries the rare codon tRNA genes argU and ileX [16 (link)]. The vector pMCSG58 is similar to pMCSG53 except it encodes for a C-terminal 6xHis-tagged protein [16 (link)]. Proteins were expressed in kanamycin-resistant BL21(DE3)MAGIC E. coli strain. The pMagic plasmid also provides a second copy of the rare codon tRNA gene argU ([17 (link)] and A. Joachimiak, personal communication).
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