Part:BBa_K567016

metY-CGA

This biobrick is constructed by mutating the anticodon of tRNA^Met to TCG (base pairing codon CGA). This tRNA can transfer fMet to CGA when it is used as the start codon. KanR gene with start codon substituted for CGA is used to testify the function of metY-CGA. When this biobrick and metGM(BBa_K567014) or metGN(BBa_K567015) are co-transformed into the cell, the cells can survive on the LB Kan plate.

Construction of BBa_K567016

metY-CGA: We have cloned operon metY containing tRNA^Met from E.coli to pACYC184. The anticodon of tRNA^Met was mutated to TCG (base pairing codon CGA).

Characterization of BBa_K567016

When this part, MetRS PT7-metGN (BBa_K567015)(or MetRS PT7-metGM (BBa_K567014)) and KanR (BBa_K567020) are co-transformed into the cell, the cell is expected to survive Kanamycin.

Fig. Growth of ER2566 with a. metGN + metY-CGA, b. metGM + metY-CGA, c. + metGN, d. + metGM. Growth medium (left): LB Kana+Tet. Growth medium (right): LB Kana.

Cell growth shows that the cells show Kanamycin resistance only when both modified MetRS (metGN) and modified tRNA^Met(metY-CGA) are transformed into the cell, proving that metGN works well.

For more information concerning this part, please see [http://2011.igem.org/Team:SJTU-BioX-Shanghai 2011 SJTU-BioX-iGEM]

Related Biobrick:

PT7-metGM (BBa_K567014)

PT7-metGN (BBa_K567015)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 183
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 183
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 183
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 183
  
• 1000
  COMPATIBLE WITH RFC[1000]