Difference between revisions of "Part:BBa M50004: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 (IPTG), our Western blot protocol revealed moderate levels of alkB1 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>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 (IPTG), our Western blot protocol revealed moderate levels of alkB1 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:FLAG_2.jpg]] | ||
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| + | <BR> The above image shows the results of our Western Blot protocol (using an anti-FLAG antibody). Low levels of alkB1 expression from singly-transformed bacteria are visible in the highlighted region; however, doubly-transformed bacteria expressed hardly any alkB1 production. | ||
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[[File:Alkane Sum.jpg]] | [[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_2707 <BR>DNA2.0 Gene #: pD444-CF<BR>Organism: E. coli<BR>Device type: Actuator<BR>Glycerol stock barcode: 0133027153<BR>Box label: BioE44 F16 |
Latest revision as of 22:33, 11 December 2016
This experience page is provided so that any user may enter their experience using this part.
Please enter
how you used this part and how it worked out.
Applications of BBa_M50004
BBa_M50004 contains the alkB1 protein (ABO_2707) originating in the bacterium Alcanivorax borkumensis. It is meant to be used in conjunction with BBa_M50005 (alkG protein, ABO_2708) 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 (IPTG), our Western blot protocol revealed moderate levels of alkB1 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-FLAG antibody). Low levels of alkB1 expression from singly-transformed bacteria are visible in the highlighted region; however, doubly-transformed bacteria expressed hardly any alkB1 production.
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.
UNIQ1da9ae6f1838a24e-partinfo-00000000-QINU UNIQ1da9ae6f1838a24e-partinfo-00000001-QINU
Stanford Location
Plasmid name: ABO_2707
DNA2.0 Gene #: pD444-CF
Organism: E. coli
Device type: Actuator
Glycerol stock barcode: 0133027153
Box label: BioE44 F16