Difference between revisions of "Part:BBa K678002"
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<partinfo>BBa_K678002 short</partinfo>
 
<partinfo>BBa_K678002 short</partinfo>
  
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Vector expressing eGFP targeted to the peroxisomes with the targeting sequence S-K-L often referred to as peroxisomal targeting sequence 1 (PTS1) from the cytomegalovirus (CMV) promoter with the BGH polyA tail and with a hygromycin resistance cassette.
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This is a device intended for expression of green fluorescent protein (GFP) in the peroxisomes of mammalian cells.
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The expression of GFP is under the control of the strong constitutive cytomegalovirus (CMV) promoter. The targeting sequence S-K-L often referred to as peroxisomal targeting sequence 1 (PTS1) is directly fused to the C-terminal of GFP, this sequence should ensure the localization of GFP to the peroxisomes of the cell.  
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===Usage and Biology===
 
===Usage and Biology===
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We used this device to target enhanced GFP to the peroxisomes of the mammalian cell line U-2 OS (Figure 1).
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We used this device to target enhanced GFP to the peroxisomes of the mammalian cell line U-2 OS (Figure 1). Although the microscopy shows GFP distributed in points which most likely are the peroxisomes of the cells, further analysis are necessary to confirm that GFP is in fact is targeted to the peroxisomes.
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[[Image:Gfppts.png|left|400px|thumb|<b>DTU-Denmark-2 2011</b> Figure 1: eGFP targeted to the peroxisomes of the mammalian cell line U-2 OS.]]
 
[[Image:Gfppts.png|left|400px|thumb|<b>DTU-Denmark-2 2011</b> Figure 1: eGFP targeted to the peroxisomes of the mammalian cell line U-2 OS.]]

Revision as of 10:18, 27 September 2011

eGFP-PTS1 for mammalian cells

This is a device intended for expression of green fluorescent protein (GFP) in the peroxisomes of mammalian cells. The expression of GFP is under the control of the strong constitutive cytomegalovirus (CMV) promoter. The targeting sequence S-K-L often referred to as peroxisomal targeting sequence 1 (PTS1) is directly fused to the C-terminal of GFP, this sequence should ensure the localization of GFP to the peroxisomes of the cell.


Usage and Biology

We used this device to target enhanced GFP to the peroxisomes of the mammalian cell line U-2 OS (Figure 1). Although the microscopy shows GFP distributed in points which most likely are the peroxisomes of the cells, further analysis are necessary to confirm that GFP is in fact is targeted to the peroxisomes.


DTU-Denmark-2 2011 Figure 1: eGFP targeted to the peroxisomes of the mammalian cell line U-2 OS.
DTU-Denmark-2 2011 Figure 2: he mammalian cell line U-2 OS transfected with device BBa_K6788002 on a plasmid and a plasmid expressing Cyan fluorescent protein.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 2233
    Illegal SpeI site found at 49
    Illegal PstI site found at 2323
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2233
    Illegal SpeI site found at 49
    Illegal PstI site found at 2323
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2233
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 2233
    Illegal SpeI site found at 49
    Illegal PstI site found at 2323
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 2233
    Illegal SpeI site found at 49
    Illegal PstI site found at 2323
    Illegal NgoMIV site found at 3088
  • 1000
    COMPATIBLE WITH RFC[1000]