Part:BBa_K3739043

J23100-B0030-his-GFP-B0010

This part contains a Vibrio natriegens-optimized GFP coding sequence, and the GFP is fused at N-terminal with his-tag in order to let us purify the protein. We use BBa_K3739043 to construct the expression system and help prove the function of some signal peptide.

Biology

GFP

Protein tags are peptides genetically fused to the proteins of interest. There are many different kinds of tags developed for for different purposes. For labeling experiments, normally the proteins of interest are tagged directly by fluorescent tags. Green fluorescent protein (GFP) make it easier to study protein localization, interactions and dynamics when fusing with other peptides and proteins.

Usage

Here, we used BBa_K3739003 to construct the expression system and obtained the composite part BBa_K3739043. We transformed the constructed plasmid on the expression vector pET-28a (+) into V. natriegens to extract the protein. This protein work together with those from BBa_K3739038 and BBa_K3739041 to verify the function of our secretory peptides Aly01 and LMT.

Characterization

1.Colony PCR We construct his-GFP CDS with J23100 promotor and B0030 RBS, the transformed cells are selected by colony PCR. The experiment result is showed in Fig. 1.

Fig. 1. DNA gel electrophoresis of colony PCR product of J23100-B0030-his-GFP-pSB1C3
Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 139