Translational_Unit

Part:BBa_K1465228:Design

Designed by: iGEM-Team Bielefeld 2014   Group: iGEM14_Bielefeld-CeBiTec   (2014-10-05)


Sedoheptulose 1,7-bisphosphatase (glpX) from Bacillus methanolicus


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

  • Amplification of the pSB1C3-Backbone (BBa_J04450) using the specific Gibson-Primer [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#fw-SBPase-pSB1C3 fw-SBPase-pSB1C3] and [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#rv-SBPase-BBa_B0034-pSB1C3 rv-SBPase-BBa_B0034-pSB1C3]:
fw-SBPase-pSB1C3 (57 bp)

5'-AATTTATTGAAGCTTATCATTGATACTAGTAGCGGCCGCTGCAGTCCGGCAAAAAAG-3'

rv-SBPase-BBa_B0034-pSB1C3 (65 bp)

5'-TTTTCGCTTTTCAATTCCCTCATTTTCTCCTCTTTCTCTAGAAGCGGCCGCGAATTCCAGAAATC-3'
  • Amplification of the front part of the Sedoheptulose-1,7-Bisphosphatase (glpX) for the removal of the illegal XbaI restriction site. This part contains the RBS BBa_B0034 and the front part of the Sedoheptulose-1,7-Bisphosphatase from Bacillus metanolicus. For the amplification the specific Gibson-Primer [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#fw-pSB1C3-BBa_B0034-SBPase fw-pSB1C3-BBa_B0034-SBPase] and [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#rv-SBPase-Schnittstelle rv-SBPase-XbaI] were used:
fw-pSB1C3-BBa_B0034-SBPase (58 bp)

5'-GAATTCGCGGCCGCTTCTAGAGAAAGAGGAGAAAATGAGGGAATTGAAAAGCGAAAAG-3'

rv-SBPase-XbaI (46 bp)

5'-TTTTCTGTTAAAGATGCGTCCAAATTTACGCAGCCTTTTGCTTTTG-3'
  • Amplification of the back part of the Sedoheptulose-1,7-Bisphosphatase (glpX) for the removal of the illegal XbaI restriction site. For the amplification the specific Gibson-Primer [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#fw-SBPase-Schnittstelle fw-SBPase-XbaI] and [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Primer#rv-pSB1C3-SBPase rv-pSB1C3-SBPase] were used:
fw-SBPase-XbaI (56 bp)

5'-AAAAGCAAAAGGCTGCGTAAATTTGGACGCATCTTTAACAGAAAATATGAAATCAG-3'

rv-pSB1C3-SBPase (56 bp)

5'-GACTGCAGCGGCCGCTACTAGTATCAATGATAAGCTTCAATAAATTGGTATTTTAC-3'
  • [http://2014.igem.org/Team:Bielefeld-CeBiTec/Notebook/Protocols#Gibson Gibson-Assembly] of the pSB1C3-Backbone and the coding sequences of the Sedoheptulose-1,7-Bisphosphatase SBPase from Bacillus metanolicus containing also the RBS BBa_B0034.

Source

  • [http://www.ncbi.nlm.nih.gov/gene/?term=glpX%20Bacillus%20methanolicus Bacillus methanolicus MGA3]: glpX class II FBPase

References