Coding

Part:BBa_K1151001

Designed by: Davide Magrž   Group: iGEM13_UniSalento_Lecce   (2013-09-21)

Histidine-rich metal-binding protein

So called for emphasize its origins in Helicobacter pylori and its avidity for nickel. Consisting of 60 aminoacids, the protein is rich in histidine (28 residues, 46.7 %) and contains short repeating motifs, it exists as an equilibration of multimeric forms in solution, with 20-mers (approx. 136 kDa) being the predominant species. Hpn can bind tightly and reversibly up to five Ni2+ ions per each monomer of 7 kDa in a pH-dependent manner (pH 7.4 ). In H. pylori play an important role in storing nickel required to the survival of the bacterium.

Boiling prep and digestion with restriction enzymes

Once inserted the gene coding for Hpn in a PSB1C3 plasmid (digestion and ligation), we transformed DH5a cells with the construct and then we recovered the ligated plasmid by boiling prep. We then proceeded with the digestion with EcoRI and PstI to check.


BoilingUnisalento.jpg

Figure 1: Digestion result of boiling prep plasmids.

PCR with VF2 and VR2 primers

                                    PCRVF21Unisalento.jpg

Figure 2: PCR results.


Sequence and Features


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]


[edit]
Categories
Parameters
n/aHistidine-rich metal-binding protein