Difference between revisions of "Part:BBa K3146010"
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and means were used to construct the plot. | and means were used to construct the plot. | ||
− | [[T--Westminster UK--todetable.png|404px|thumb|center|Table1]]. | + | [[File:T--Westminster UK--todegraph.png|404px|thumb|center|Figure 1]] |
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+ | [[File:T--Westminster UK--todetable.png|404px|thumb|center|Table1]]. | ||
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Even though addition of a different RBS and Promoter to the TodE coding sequence could not be | Even though addition of a different RBS and Promoter to the TodE coding sequence could not be | ||
directly compared to the previous years results we believe the expression of the enzyme was | directly compared to the previous years results we believe the expression of the enzyme was | ||
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different strain of E. Coli top 10 that generally has lower recombinant protein expression than E coli | different strain of E. Coli top 10 that generally has lower recombinant protein expression than E coli | ||
DH-a that was used last year. As well as being able to obtain similar results using 4 hour bacterial | DH-a that was used last year. As well as being able to obtain similar results using 4 hour bacterial | ||
− | stock. Raw data can be seen in table 1 above and plot A.V.agains time can be seen in | + | stock. Raw data can be seen in table 1 above and plot A.V.agains time can be seen in Figure 1. |
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Latest revision as of 03:21, 22 October 2019
TodE with promoter and RBS
TodE improved from the previous BioBrick, BBa_K2626000.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 405
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 455
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 480
TodE 3-methylcatechol 2,3-dioxygenase enzyme was ligated to BBa_J23100 constitutive promoter,
BBa_B0034 strong Ribosome Binding Site in order to improve expression. Heat shock
transformation protocol was applied, colonies allowed to grow for 24 hours. 4 hour bacterial stock
inoculated with single colony was lysed and 2mM of 3-methylcatechol was added to the solution.
E. Coli TOP10 and 2mM 3-methylcatechol was used as control. AV at 384 was monitored for 3
hours. Measurements were take in triplicates using Nanodrop 200 Thermo scientific every hour
and means were used to construct the plot.
Even though addition of a different RBS and Promoter to the TodE coding sequence could not be
directly compared to the previous years results we believe the expression of the enzyme was
improved. This statement could be supported due to very similar results being obtained in a
different strain of E. Coli top 10 that generally has lower recombinant protein expression than E coli
DH-a that was used last year. As well as being able to obtain similar results using 4 hour bacterial
stock. Raw data can be seen in table 1 above and plot A.V.agains time can be seen in Figure 1.