Part:BBa_K1648004

Insertion Kit for Fusing Protein of Interest to Magnetosome Membrane

Construct Map for K1648004

Consist of a gene (mamC) from magnetosome island of Magnetospirillum gryphiswaldense genome, a promotor & RBS(J13002) in front of multiple restriction sites and a double terminator(B0015) at the back of multiple restriction sites.

Different genes can be added into this part at multiple restriction sites, so that different proteins will be expressed on the magnetosome after transformation of the part into A. vinelandii genome via random integration.

A GFP-nanobody(K1648005) is added to this part in order to express GFP-nanobody on the magnetosome. This new part is K1648006

This Part were verified by double digestion (Fig. 1) and sequencing.

Fig 1. The photo of 1% agarose gel electrophoresis showing PCR products of operons in Magnetosome Island (MAI). L: DNA ladder. Lane 1: mamHIEJKLMN. Lane 2: mamOPQRBSTU. Lane 3: mamPQRBSTU. Lane 4: mamYXZftsZ-like. Lane 5: mamGFDC. Lane 6: mamGFD. Lane 7: mms6.

RFP is inserted in Insertion Kit for biobrick standard. The Insertion kit with RFP is transformed in E. coli (DH5 alpha). Expression of RFP in Insertion is shown(Fig 2.).

Fig 2. Insertion kit with RFP(BBa_K1648004) is expressed in E. coli (DH5 alpha)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2759
  Illegal XbaI site found at 2765
  Illegal SpeI site found at 1
  Illegal PstI site found at 7
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2759
  Illegal SpeI site found at 1
  Illegal PstI site found at 7
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2759
  Illegal XhoI site found at 1162
  Illegal XhoI site found at 2188
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2759
  Illegal XbaI site found at 2765
  Illegal SpeI site found at 1
  Illegal PstI site found at 7
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal EcoRI site found at 2759
  Illegal XbaI site found at 2765
  Illegal SpeI site found at 1
  Illegal PstI site found at 7
  Illegal NgoMIV site found at 2546
  Illegal NgoMIV site found at 2636
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Plasmid lacks a prefix.
  Plasmid lacks a suffix.
  Illegal BsaI site found at 2611