Difference between revisions of "Part:BBa K2117005"
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Excitation: 488 nm
 
Excitation: 488 nm
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===Experimental data===
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The two parts (TEF1 and hrGFP) was assembled with the pSB1A8YL backbone (BBa_K2117009) using standard 3A assembly and transformed into E. coli. The construct was confirmed using restriction analysis (See Figure 1), PCR and sequencing (data not shown). The construct was then transformed into Y. lipolytica PO1f along with the pSB1A8YL plasmid without inserts as negative control.
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*Insert gel picture of restriction analysis*
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The transformation cultures were grown in 30 C in 48 hours and analysed by Confocal Laser Fluorescent microscopy. The control showed some autofluorescence, but the transformant with the construct gave a clear GFP signal (See Figure 2).
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<partinfo>BBa_K2117005 parameters</partinfo>
 
<partinfo>BBa_K2117005 parameters</partinfo>
 
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Revision as of 16:13, 19 October 2016


Device encoding TEF1 and hrGFP for expression in Yarrowia lipolytica

This is a composite part consisting of the TEF1 promoter (BBa_K2117000) and hrGFP (BBa_K2117003).

This part encodes the humanized Renilla reniformis green fluorescent protein (hrGFP) codon-optimized for Yarrowia lipolytica and the native Y. lipolytica constitutive promoter TEF1.

Studies have shown the hrGFP to be functional in Y. lipolytica under the expression of the TEF1 promoter measured by flow cytometry. *insert reference*


Usage and Biology

The green fluorescent protein originates from the sea pansy, Renilla reniformis. This part can be used as a reporter protein for protein expression in the yeast chassis, Y. lipolytica. It is compatible with all the iGEM Biobrick standards.

Emission: 500-600 nm

Excitation: 488 nm

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 BsaI.rc site found at 2


Functional Parameters

Emission: 500-600 nm

Excitation: 488 nm

Experimental data

The two parts (TEF1 and hrGFP) was assembled with the pSB1A8YL backbone (BBa_K2117009) using standard 3A assembly and transformed into E. coli. The construct was confirmed using restriction analysis (See Figure 1), PCR and sequencing (data not shown). The construct was then transformed into Y. lipolytica PO1f along with the pSB1A8YL plasmid without inserts as negative control.

  • Insert gel picture of restriction analysis*

The transformation cultures were grown in 30 C in 48 hours and analysed by Confocal Laser Fluorescent microscopy. The control showed some autofluorescence, but the transformant with the construct gave a clear GFP signal (See Figure 2).