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Team:Groningen/international cooperation
From 2012.igem.org
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The iGEM team LMU Munich 2012 has developed a plasmid backbone that is suitable for gene expression in <i>B. subtilis</i> <b>(BBa_K823023)</b>. | The iGEM team LMU Munich 2012 has developed a plasmid backbone that is suitable for gene expression in <i>B. subtilis</i> <b>(BBa_K823023)</b>. | ||
| - | This plasmid backbone contains an integration locus in amyE, which means after being transformed into <i>B. subtilis</i>, | + | This plasmid backbone contains an integration locus in amyE, which means that after being transformed into <i>B. subtilis</i>, |
| - | the cloned insert will be integrated with <i>B. subtilis</i> genome at the amyE locus. <i>B. subtilis</i> is known for its | + | the cloned insert will be integrated with the <i>B. subtilis</i> genome at the amyE locus. <i>B. subtilis</i> is known for its |
starch degrading ability and amyE is a gene responsible for the starch degradation phenotype in <i>B.subtilis</i> by regulating a-amylase production. | starch degrading ability and amyE is a gene responsible for the starch degradation phenotype in <i>B.subtilis</i> by regulating a-amylase production. | ||
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We characterized this plasmid backbone by doing a simple growth experiment on a starch agar plate. <i>B. subtilis</i> | We characterized this plasmid backbone by doing a simple growth experiment on a starch agar plate. <i>B. subtilis</i> | ||
strains (168 wildtype, pSac-Cm, and K823023, clockwise respectively) were inoculated in LB agar+0.4% starch and incubated overnight | strains (168 wildtype, pSac-Cm, and K823023, clockwise respectively) were inoculated in LB agar+0.4% starch and incubated overnight | ||
| - | in 37<sup>o</sup>C (left picture). After the addition of 0.1% iodine, | + | in 37<sup>o</sup>C (left picture). After the addition of 0.1% iodine, a clear zone was formed around <i>B. subtilis</i> |
wildtype and <i>B. subtilis</i> strain containing pSac-Cm. There was no clear zone observed around | wildtype and <i>B. subtilis</i> strain containing pSac-Cm. There was no clear zone observed around | ||
<i>B. subtilis</i> strain containing backbone K823023 (right picture). | <i>B. subtilis</i> strain containing backbone K823023 (right picture). | ||
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At the beginning of August we received the chromoproteins amilCP (blue), amilGFP (yellow) from | At the beginning of August we received the chromoproteins amilCP (blue), amilGFP (yellow) from | ||
<a class="inlink" href="https://2011.igem.org/Team:Uppsala-Sweden">iGEM Uppsala 2011</a>. | <a class="inlink" href="https://2011.igem.org/Team:Uppsala-Sweden">iGEM Uppsala 2011</a>. | ||
| - | We were able to transform their BBa_K592009 and BBa_K592010 behind our identified promoters into <i>E.coli</i> | + | We were able to transform their BBa_K592009 and BBa_K592010 behind our identified promoters into <i>E.coli</i> |
| - | + | and managed to turn <i>B.subtilis</i> into blue and yellow, which has never been done before! | |
For the latest results, check our <a class="inlink" href="https://2012.igem.org/Team:Groningen/pigmentproduction">Pigment page</a>. | For the latest results, check our <a class="inlink" href="https://2012.igem.org/Team:Groningen/pigmentproduction">Pigment page</a>. | ||
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Emily Hicks from the winning Calgary iGEM team in 2011, and now member of iGEM Calgary 2012, has studied in Groningen during her exchange period. | Emily Hicks from the winning Calgary iGEM team in 2011, and now member of iGEM Calgary 2012, has studied in Groningen during her exchange period. | ||
She was very interested in her iGEM Groningen colleagues; therefore we asked her to give a lecture about her previous iGEM experience. | She was very interested in her iGEM Groningen colleagues; therefore we asked her to give a lecture about her previous iGEM experience. | ||
| - | We were really motivated by Emily's | + | We were really motivated by Emily's suggestions concerning medal requirements, information about wiki etc. Thank you Emily! |
</p> | </p> | ||
<br> | <br> | ||
Revision as of 22:35, 26 September 2012
The iGEM team LMU Munich 2012 has developed a plasmid backbone that is suitable for gene expression in B. subtilis (BBa_K823023).
This plasmid backbone contains an integration locus in amyE, which means that after being transformed into B. subtilis,
the cloned insert will be integrated with the B. subtilis genome at the amyE locus. B. subtilis is known for its
starch degrading ability and amyE is a gene responsible for the starch degradation phenotype in B.subtilis by regulating a-amylase production.
|
|
We characterized this plasmid backbone by doing a simple growth experiment on a starch agar plate. B. subtilis
strains (168 wildtype, pSac-Cm, and K823023, clockwise respectively) were inoculated in LB agar+0.4% starch and incubated overnight
in 37oC (left picture). After the addition of 0.1% iodine, a clear zone was formed around B. subtilis
wildtype and B. subtilis strain containing pSac-Cm. There was no clear zone observed around
B. subtilis strain containing backbone K823023 (right picture).
The inability of the B. subtilis strain containing BBa_K823023 to degrade starch has demonstrated that the amyE locus in
the B. subtilis genome had been replaced. Apart from this characterization experiment, our team has been using BBa_K823023
as a plasmid backbone for our volatile detection device in B. subtilis
sboA-lycopene.
- Nicholson, W.L, Park, Y.K, Henkin, T.M, Won, M., Weickert, M.J, Gaskell, J.A, Chambliss, G.H. 2006. Catabolite repression-resistant mutations of the Bacillus subtilis alpha-amylase promoter affect transcription levels and are in an operator-like sequence. J Mol Biol. 1987 Dec 20;198(4):609-18.
- Zeigler, Daniel R. 2002. Bacillus Genetic Stock Center Catalog of Strains, Seventh Edition, Volume 4: Integration Vectors for Gram-Positive Organisms. The Bacillus Genetic Stock Center. Department of Biochemistry, The Ohio State University. 484 West Twelfth Avenue,Columbus, Ohio 43210.
We participated in the survey of the iGEM TU Munich and are very proud that we got their original Bavarian Collaboration-Medal.
|
At the beginning of August we received the chromoproteins amilCP (blue), amilGFP (yellow) from
iGEM Uppsala 2011.
We were able to transform their BBa_K592009 and BBa_K592010 behind our identified promoters into E.coli
and managed to turn B.subtilis into blue and yellow, which has never been done before!
For the latest results, check our Pigment page.
|
All of the Dutch iGEM teams collaborated to create an iGEM presence at the
Discovery Festival on Friday 28 September
in Amsterdam, Rotterdam and Eindhoven. We will aim to get the public in touch with Synthetic Biology in a playful way.
Besides these Dutch iGEM meetings and the Discovery Festival, we also collaborated with Wageningen and Amsterdam on a molecular level.
Many times our transformation of E.coli failed, so we ran out of the backbone biobricks for B. subtilis. Luckily Wageningen
and Amsterdam were so kind to provide us with some of their bricks. Thank you guys!
Emily Hicks from the winning Calgary iGEM team in 2011, and now member of iGEM Calgary 2012, has studied in Groningen during her exchange period.
She was very interested in her iGEM Groningen colleagues; therefore we asked her to give a lecture about her previous iGEM experience.
We were really motivated by Emily's suggestions concerning medal requirements, information about wiki etc. Thank you Emily!
Prof. Papin, supervisor to the iGEM Virginia team and supervising author to the idFBA article, graciously shared his knowledge and scripts relating to
idFBA.
Our sponsors:
