Coding
tetA(C)b

Part:BBa_J31006:Design

Designed by: Sabriya Rosemond, Erin Zwack   Group: iGEM06_Davidson   (2006-07-12)
Revision as of 20:57, 6 December 2006 by Kahaynes (Talk | contribs) (→‎Design Notes)

tetracycline resistance protein TetA(C) (backwards) [cf. BBa_J31007]


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 1043
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 897
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 343
    Illegal NgoMIV site found at 503
    Illegal NgoMIV site found at 871
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

This part was PCR amplified from pSB1AT3 using the following primers. The primers have non-annealing 5'- extensions that introduce a SpeI site to the left and an XbaI site to the right of the coding region (allowing BioBrick cloning in the reverse orientation). Primer annealing sites are shown in bold.
Forward: 5’ ATGCACTAGTATGAAATCTAACAATGCGCTCATC (SpeI site)
Reverse: 5’ GCATTCTAGATTAGGTCGAGGTGGCCCGGC (XbaI site)

The final part was cloned into vector pSB1A2. The Biobricks on this part are not wild type, but the cut sites are still viable.

Standard BioBrick Cloning Sites (Knight) 5'--GAATTC GCGGCCGC T TCTAGA G ----insert---- T ACTAGT A GCGGCCG CTGCAG--
3'--CTTAAG CGCCGGCG A AGATCT C -------------- A TGATCA T CGCCGGC GACGTC--
BBa_J31001 Cloning Sites 5'--GAATTC GCGGCCGC T TCTAGA * --Tet coding-- * ACTAGT A GCGGCCG CTGCAG--
3'--CTTAAG CGCCGGCG A AGATCT * -------------- * TGATCA T CGCCGGC GACGTC--

Prefix
There is no G spacer (*) between the XbaI and the Tet coding region.
Suffix
There is no T spacer (*) between the Tet coding region and the SpeI site.

Data

Once placed to the right of a reverse RBS, TetB conveys tetrcycline resistance to JM109 cells (see BBa_S03532).

Source

The tetracycline resistance coding region was PCR amplified from pSB1AT3.

References