Difference between revisions of "Part:BBa K2205004"
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Sarcosine Oxidase under the control of an IPTG-inducible T7 promoter. Catalyses sarcosine to yield glycine, formaldehyde and hydrogen peroxide. | Sarcosine Oxidase under the control of an IPTG-inducible T7 promoter. Catalyses sarcosine to yield glycine, formaldehyde and hydrogen peroxide. | ||
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| + | ==Induction with IPTG== | ||
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| + | [[File:T--Newcastle--Correct sox protein gel.png|400px|]] | ||
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| + | '''Sarcosine Oxidase Expression:''' Sarcosine Oxidase expression was induced by adding 40 µl of 100 mM IPTG. Lane 1: 6 µl ladder, Lane 2: 10 µl sfGFP, Lane 3: BL21-DE3, Lane 4: 10µl SOX 1, Lane 5: 10 µl SOX 2, Lane 6: 10 µl SOX 3, Lane 7: 10 µl SOX 4, Lane 8: 10 µl SOX 5, Lane 9: 10 µl SOX 6, Lane 10: 6 µl ladder. | ||
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| + | ==Formaldehyde Production== | ||
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| + | [[File:T--Newcastle--SOX testing.JPG|400px|]] | ||
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| + | '''Testing of Sarcosine Oxidase with Sarcosine:''' Lysed cell cultures induced with 40 µl of a 100 mM stock of IPTG tested with formaldehyde strips. From left to right: SOX+0 µl sarcosine, SOX+IPTG+0 µl sarcosine, WT+0 µl sarcosine, WT+IPTG+0 µl sarcosine, SOX+50 µl sarcosine, SOX+IPTG+50 µl sarcosine, WT+50 µl sarcosine, WT+IPTG+50 µl sarcosine, SOX+200 µl sarcosine, SOX+IPTG+200 µl, WT+200 µl sarcosine, WT+IPTG+200 µl. | ||
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| + | This shows sarcosine oxidase works as expected, however there is leaky expression as formaldehyde is produced when no IPTG is added. | ||
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==Usage and Biology== | ==Usage and Biology== | ||
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Latest revision as of 20:14, 27 October 2017
T7-Sarcosine Oxidase
Sarcosine Oxidase under the control of an IPTG-inducible T7 promoter. Catalyses sarcosine to yield glycine, formaldehyde and hydrogen peroxide.
Induction with IPTG
Sarcosine Oxidase Expression: Sarcosine Oxidase expression was induced by adding 40 µl of 100 mM IPTG. Lane 1: 6 µl ladder, Lane 2: 10 µl sfGFP, Lane 3: BL21-DE3, Lane 4: 10µl SOX 1, Lane 5: 10 µl SOX 2, Lane 6: 10 µl SOX 3, Lane 7: 10 µl SOX 4, Lane 8: 10 µl SOX 5, Lane 9: 10 µl SOX 6, Lane 10: 6 µl ladder.
Formaldehyde Production
Testing of Sarcosine Oxidase with Sarcosine: Lysed cell cultures induced with 40 µl of a 100 mM stock of IPTG tested with formaldehyde strips. From left to right: SOX+0 µl sarcosine, SOX+IPTG+0 µl sarcosine, WT+0 µl sarcosine, WT+IPTG+0 µl sarcosine, SOX+50 µl sarcosine, SOX+IPTG+50 µl sarcosine, WT+50 µl sarcosine, WT+IPTG+50 µl sarcosine, SOX+200 µl sarcosine, SOX+IPTG+200 µl, WT+200 µl sarcosine, WT+IPTG+200 µl.
This shows sarcosine oxidase works as expected, however there is leaky expression as formaldehyde is produced when no IPTG is added.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]