Team:SJTU-BioX-Shanghai/Project/Subproject3
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We screened the MetRS obtained through error-prone PCR and obtained one target mutant. We have also tested the activity of the truncated MetRS PT7-''metG''N ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K567015 BBa_K567015])and found that the truncated MetRS acted as expected, losing specificity for tRNA<sup>Met</sup> anticodon while maintaining aminoacylation ability. | We screened the MetRS obtained through error-prone PCR and obtained one target mutant. We have also tested the activity of the truncated MetRS PT7-''metG''N ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K567015 BBa_K567015])and found that the truncated MetRS acted as expected, losing specificity for tRNA<sup>Met</sup> anticodon while maintaining aminoacylation ability. | ||
- | [[image:11SJTU-initial_codon_result.jpg|frame|center|''Fig 2.'' Growth of ER2566 with a. ''metG''N + ''metY''-CGA, b. ''metG''M + ''metY''-CGA, c. + ''metG''N, d. + ''metG''M. Growth medium (left): LB Kana+Tet. Growth medium (right): LB Kanamycin.]] | + | [[image:11SJTU-initial_codon_result.jpg|frame|center|''Fig 2.'' Growth of ER2566 with a. ''metG''N + ''metY''-CGA, b. ''metG''M + ''metY''-CGA, c. + ''metG''N, d. + ''metG''M. Growth medium (left): LB Kana+Tet. Growth medium (right): LB Kanamycin.'''Cells survived Kanamycin with our device. Without our device, cells cannot survive.''' ]] |
Cell growth shows that the cells have Kanamycin resistance only when both modified MetRS (''metG''N or ''metG''M) and modified tRNA<sup>Met</sup>(''metY''-CGA) are transformed into the cell, '''proving that tRNA ''metY''-CGA can transfer fMet to CGA when it is used as the start codon and that ''metG''N and ''metG''M work well. ''' | Cell growth shows that the cells have Kanamycin resistance only when both modified MetRS (''metG''N or ''metG''M) and modified tRNA<sup>Met</sup>(''metY''-CGA) are transformed into the cell, '''proving that tRNA ''metY''-CGA can transfer fMet to CGA when it is used as the start codon and that ''metG''N and ''metG''M work well. ''' |
Revision as of 23:35, 28 October 2011