Part:BBa_K3042005

Dino III GFP Plasmid

A dinoflagellate optimized reporter plasmid. DinoIII is a dinoflagellate expression system that is constructed on the basis of genomic and transcriptomic data from many dinoflagellates. DinoIII GFP is a dinoflagellate optimized reporter plasmid which was created by the University of Connecticut. This part can be used as a reporter via fluorescence. When the transformed cells fluoresce green, this indicates the expression of the GFP (Specher, 2019). This part has been successfully transformed into O.Marina by Senjie Lab at University of Connecticut. iGEM GSU wants to replicate their results with this part as well as into Symbiodinium microadriaticum, another dinoflagellate.

*All sequences/info are from snapgene* Features: RNA elements Sprecher, Brittany & Zhang, Huan & Lin, Senjie. (2019). Nuclear gene transformation in a dinoflagellate. 10.1101/602821.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 682
  Illegal NheI site found at 1781
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 1201
  Illegal XhoI site found at 1102
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1763
  

AmpR promoter- Ampicillin resistance promoter AmpR- Product: Beta lactamase Confers resistance to ampicillin and related antibiotics

Ori- High copy number ColE1/pMB1/pBR322/pUC origin of replication Cap binding site-Cap binding activates transcription in the presence of cAMP

Lac operator- The lac repressor binds the lac operator to block transcription in E. Coli. This inhibition can be relieved by adding lactose or IPTG Lac promoter- Promoter for E. Coli lac operon Terminator- part k3042003 Cycle 3 GFP- Mammalian codon-optimized cycle 3 GFP (Crameri et al., 1996) Promoter region- part K3042002