Difference between revisions of "Part:BBa K1049002:Experinece"
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Our team KIT-Kyoto 2013 constructed this part for the purpose of measurement. T7 promoter is an IPTG-inducible promoter. We added 20uL IPTG (100mM) to our genetically modified E.coli after cultivation at 28 and 37 degree C. 2 hours after, we extracted soluble proteins from it by using FastBreak™ Cell Lysis Reagent and did SDS-polyacrylamide gel electrophoresis.
 
Our team KIT-Kyoto 2013 constructed this part for the purpose of measurement. T7 promoter is an IPTG-inducible promoter. We added 20uL IPTG (100mM) to our genetically modified E.coli after cultivation at 28 and 37 degree C. 2 hours after, we extracted soluble proteins from it by using FastBreak™ Cell Lysis Reagent and did SDS-polyacrylamide gel electrophoresis.
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[[File:ATF2.png|500px|thumb|left|SDS-PAGE]]
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[[File:ATF2.png|500px|thumb|left|Figure.1 SDS-PAGE Starting from the left, marker, candidate 1 at 28 degree C, candidate 1 + IPTG at 28 degree C, candidate 2 at 28 degree C, candidate 2 + IPTG at 28 degree C, marker, candidate 1 at 37 degree C, candidate 1 + IPTG at 37 degree C, candidate 2 at 37 degree C, candidate 2 + IPTG at 37 degree C.
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]]
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ATF2 gene encodes AATase, which is about 62kDa. The consumption of protein marker is like this.
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Myosin                                200kDa
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β‐Galactosidase                  120kDa
 +
Bovine Serum Albumin        95kDa
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Glutamine dehydrogenase    68kDa
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Ovalbumin                          50kDa
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Carbonic Anhydrase            36kDa
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Myoglobin                            27kDa
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Lysozyme                            20kDa
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Aprotinin                              10kDa
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 +
You can find the band at lanes which are added IPTG just beneath the band of 68kDa.

Revision as of 02:35, 20 September 2013

Our team KIT-Kyoto 2013 constructed this part for the purpose of measurement. T7 promoter is an IPTG-inducible promoter. We added 20uL IPTG (100mM) to our genetically modified E.coli after cultivation at 28 and 37 degree C. 2 hours after, we extracted soluble proteins from it by using FastBreak™ Cell Lysis Reagent and did SDS-polyacrylamide gel electrophoresis.

Figure.1 SDS-PAGE Starting from the left, marker, candidate 1 at 28 degree C, candidate 1 + IPTG at 28 degree C, candidate 2 at 28 degree C, candidate 2 + IPTG at 28 degree C, marker, candidate 1 at 37 degree C, candidate 1 + IPTG at 37 degree C, candidate 2 at 37 degree C, candidate 2 + IPTG at 37 degree C.

ATF2 gene encodes AATase, which is about 62kDa. The consumption of protein marker is like this.

Myosin 200kDa β‐Galactosidase 120kDa Bovine Serum Albumin 95kDa Glutamine dehydrogenase 68kDa Ovalbumin 50kDa Carbonic Anhydrase 36kDa Myoglobin 27kDa Lysozyme 20kDa Aprotinin 10kDa

You can find the band at lanes which are added IPTG just beneath the band of 68kDa.