Part:BBa_K2201404

Reverse Sequence of I746909

This part is the reverse sequence of BBa_I746909. I746909 was build to charachterize superfolder GFP under the control of the T7 promotor by the Cambridge iGEM team 2008.

Usage and Biology

GFP is a fluorescent protein. In this part it is under the control of the T7 promotor which is recognized by the T7 RNA polymerase. When combining this part with the T7 RNA polymerase the GFP will be expressed and can be detected. This mechanism can be used for detection methods. We provide this part in reverse order for possible integration into a positive selection plasmid for aminoacyl-tRNA synthetase library selection.

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
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI.rc site found at 62
  

Functional Parameters

We investigated if reversing the sequence of I746909 does influence the fluorescence efficiency compared to the original part. Both constructs were tested in the E. coli strain BL21. After both cultures grew O/N the OD₆₀₀ and the fluorescence was measured. Figure 1 shows the Emission ploted against the OD.

Figure 1: Fluorescence efficiency of forward and reverse pT7-GFP. GFP forward (GFPn, red) and GFP reverse (GFPr, green) wer both tested in BL21. The emission is ploted against the OD. The results are based on oone biological replicate. R² is shown in the legend.

It can be seen that the fluorescence of the original part (BBa_I746909) is more intense.