Difference between revisions of "Part:BBa K567016"

Revision as of 02:29, 29 October 2011

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 Kana 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]

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K567016 short</partinfo>
-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. KanaR gene with start codon substituted for CGA is used to testify the function of ''metY''-CGA. When this biobrick and ''metG''M(BBa_K567014) or ''metG''N(BBa_K567015) are co-transformed into the cell, the cells can survive on the LB Kana plate.
+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 ''metG''M(BBa_K567014) or ''metG''N(BBa_K567015) are co-transformed into the cell, the cells can survive on the LB Kana plate.
@@ Line 12: / Line 12: @@
 ===Characterization of BBa_K567016===
-When this part, MetRS PT7-''metG''N (BBa_K567015)(or MetRS PT7-''metG''M (BBa_K567014)) and KanaR (BBa_K567020) are co-transformed into the cell, the cell is expected to survive Kana.
+When this part, MetRS PT7-''metG''N (BBa_K567015)(or MetRS PT7-''metG''M (BBa_K567014)) and KanR (BBa_K567020) are co-transformed into the cell, the cell is expected to survive Kanamycin.
 [[image:11SJTU-initial_codon_result.jpg|frame|center|Fig. 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 Kana.]]
-Cell growth shows that the cells show Kana resistance only when both modified MetRS (''metG''N) and modified tRNA<sup>Met</sup>(''metY''-CGA) are transformed into the cell, proving that ''metG''N works well.
+Cell growth shows that the cells show Kanamycin resistance only when both modified MetRS (''metG''N) and modified tRNA<sup>Met</sup>(''metY''-CGA) are transformed into the cell, proving that ''metG''N works well.