Difference between revisions of "Part:BBa M50005:Experience"
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===User Reviews=== | ===User Reviews=== | ||
<I>Stanford BioE44 "Team WERK" 12.10.2016</I> | <I>Stanford BioE44 "Team WERK" 12.10.2016</I> | ||
| − | <BR>We transformed E. coli with both BBa_M50004 and BBa_M50005 in an attempt to replicate the first step of A. borkumensis' alkane degradation in different bacteria. Under proper induction (rhamnose), our Western blot protocol revealed high levels of alkG production. However, pH testing failed to confirm the cooperative function of alkB1 and alkG, with transformed bacteria in alkane-rich conditions behaving relatively indistinguishably from those in standard conditions. This may have been due to a failure in double transformation, with the two proteins being expressed at disproportionate levels. <BR><BR> | + | <BR>We transformed E. coli with both BBa_M50004 and BBa_M50005 in an attempt to replicate the first step of A. borkumensis' alkane degradation in different bacteria. Under proper induction (rhamnose), our Western blot protocol revealed high levels of alkG production. However, pH testing failed to confirm the cooperative function of alkB1 and alkG, with transformed bacteria in alkane-rich conditions behaving relatively indistinguishably from those in standard conditions. This may have been due to a failure in double transformation, with the two proteins being expressed at disproportionate levels. |
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| + | [[File:His_2.jpg]] | ||
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| + | <BR> The above image shows the results of our Western Blot protocol (using an anti-His antibody). Moderate levels of alkG expression are evident in all transformed bacteria; expression is most prevalent in singly-transformed bacteria (the highlighted region). | ||
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| + | [[File:Alkane Sum.jpg]] | ||
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| + | <BR> The above graph shows the results of our pH testing; each dataset represents a different instance of transformed E. coli. Measurements were normalized against the initial pH of each bacteria culture. Under proper collaborative function of the alkG and alkB1 enzymes, pH ought to have decreased over time relative to the control; however, no concrete results were observed. | ||
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===Stanford Location=== | ===Stanford Location=== | ||
| − | Plasmid name: | + | Plasmid name: ABO_2708 <BR>DNA2.0 Gene #: pD861-CH<BR>Organism: E. coli<BR>Device type: Actuator<BR>Glycerol stock barcode: 0133021780<BR>Box label: BioE44 F16 |
Latest revision as of 22:31, 11 December 2016
This experience page is provided so that any user may enter their experience using this part.
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how you used this part and how it worked out.
Applications of BBa_M50005
BBa_M50005 contains the alkG protein (ABO_2708) originating in the bacterium Alcanivorax borkumensis. It is meant to be used in conjunction with BBa_M50004 (alkB1 protein, ABO_2707) and other enzymes from A. borkumensis to degrade alkanes measuring up to 32 carbons as part of A. borkumensis' digestive process.
User Reviews
Stanford BioE44 "Team WERK" 12.10.2016
We transformed E. coli with both BBa_M50004 and BBa_M50005 in an attempt to replicate the first step of A. borkumensis' alkane degradation in different bacteria. Under proper induction (rhamnose), our Western blot protocol revealed high levels of alkG production. However, pH testing failed to confirm the cooperative function of alkB1 and alkG, with transformed bacteria in alkane-rich conditions behaving relatively indistinguishably from those in standard conditions. This may have been due to a failure in double transformation, with the two proteins being expressed at disproportionate levels.
The above image shows the results of our Western Blot protocol (using an anti-His antibody). Moderate levels of alkG expression are evident in all transformed bacteria; expression is most prevalent in singly-transformed bacteria (the highlighted region).
The above graph shows the results of our pH testing; each dataset represents a different instance of transformed E. coli. Measurements were normalized against the initial pH of each bacteria culture. Under proper collaborative function of the alkG and alkB1 enzymes, pH ought to have decreased over time relative to the control; however, no concrete results were observed.
UNIQ44a75bd3e340f023-partinfo-00000000-QINU UNIQ44a75bd3e340f023-partinfo-00000001-QINU
Stanford Location
Plasmid name: ABO_2708
DNA2.0 Gene #: pD861-CH
Organism: E. coli
Device type: Actuator
Glycerol stock barcode: 0133021780
Box label: BioE44 F16