Difference between revisions of "Part:BBa K1189032:Experience"
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This experience page is provided so that any user may enter their experience using this part.<BR>Please enter | This experience page is provided so that any user may enter their experience using this part.<BR>Please enter | ||
how you used this part and how it worked out. | how you used this part and how it worked out. | ||
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| + | <html> | ||
| + | <h1>Purpose</h1> | ||
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| + | <p>The <a href="http://2013.igem.org/Team:Calgary">Calgary iGEM 2013</a> team sought to detect pathogenic enterohemorrhagic <i>E. coli</i> (EHEC) DNA using pairs of TALEs designed to pathogenic <i>E. coli</i> markers. This TALE was designed based on a sequence alignment of the <i>stx2</i> gene found in EHEC organisms. It binds to a 19 bp region of the toxin, will detect EHEC strains including <i>E. coli O157:H7</i>, as well as several other EHEC strains. This TALE, along with the complementary <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K1189033">BBa_K1189033</a>, will eventually be integrated into the final <a href="http://2013.igem.org/Team:Calgary/Project/OurSensor/Prototype">lateral flow strip FerriTALE prototype</a>.</p> | ||
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| + | <h1>Design details</h1> | ||
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| + | <p>The sequence includes a KasI restriction enzyme cut site so that other TALEs may be substituted with the C-terminal linker and K coil. This is one element in which the 2013 Calgary team attempted to make their DNA detection system modular and useful for detection of other DNA sequences based on the selection of an appropriate TALE. The target site was specifically selected to increase binding strength of the TALE to DNA as per Meckler et al. (2013). Please see the <a href="http://2013.igem.org/Team:Calgary/Project/OurSensor/Detector#Engineered TALEs">iGEM Calgary 2013 Wiki</a> for more details about how these TALEs were engineered for optimal binding.</a> | ||
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| + | <figure> | ||
| + | <img src="https://static.igem.org/mediawiki/2013/5/50/UCalgary2013TRTwotargetTALE.png" alt="EHEC TALE 2 SOE PCR Product" width="600" height="300"> | ||
| + | <figcaption> | ||
| + | <p><b>Figure 1.</b> Sequence alignment used to select a pair of TALEs to detect a large group of EHEC.</p> | ||
| + | </figcaption> | ||
| + | </figure> | ||
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| + | <h1>Progress</h1> | ||
| + | <p>The Calgary team has tested their <a href="http://2013.igem.org/Team:Calgary/Project/OurSensor">DNA detection system </a>using proof of concept TALEs designed by the <a href="http://2012.igem.org/Team:Slovenia">2012 Slovenia iGEM team</a>. They have assembled BBa_K1189033 as shown in Figure 2 and are working on expressing, testing, and integrating these protein components into the final system.</p> | ||
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| + | <h1>References</h1> | ||
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| + | <p>Meckler, J.F., Bhakta, M.S., Kim, M.S., Ovadia, R., Habrian, C.H., Zykovich, A., ... Baldwin, E.P. (2013). Quantitative analysis of TALE-DNA interactions suggests polarity effects. Nucleic Acids Research, 41(7), 4118–28. doi:10.1093/nar/gkt085</p> | ||
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| + | <br></br> | ||
| + | </html> | ||
===Applications of BBa_K1189032=== | ===Applications of BBa_K1189032=== | ||
Latest revision as of 10:49, 28 October 2013
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.
Purpose
The Calgary iGEM 2013 team sought to detect pathogenic enterohemorrhagic E. coli (EHEC) DNA using pairs of TALEs designed to pathogenic E. coli markers. This TALE was designed based on a sequence alignment of the stx2 gene found in EHEC organisms. It binds to a 19 bp region of the toxin, will detect EHEC strains including E. coli O157:H7, as well as several other EHEC strains. This TALE, along with the complementary BBa_K1189033, will eventually be integrated into the final lateral flow strip FerriTALE prototype.
Design details
The sequence includes a KasI restriction enzyme cut site so that other TALEs may be substituted with the C-terminal linker and K coil. This is one element in which the 2013 Calgary team attempted to make their DNA detection system modular and useful for detection of other DNA sequences based on the selection of an appropriate TALE. The target site was specifically selected to increase binding strength of the TALE to DNA as per Meckler et al. (2013). Please see the iGEM Calgary 2013 Wiki for more details about how these TALEs were engineered for optimal binding.
Figure 1. Sequence alignment used to select a pair of TALEs to detect a large group of EHEC.
Progress
The Calgary team has tested their DNA detection system using proof of concept TALEs designed by the 2012 Slovenia iGEM team. They have assembled BBa_K1189033 as shown in Figure 2 and are working on expressing, testing, and integrating these protein components into the final system.
References
Meckler, J.F., Bhakta, M.S., Kim, M.S., Ovadia, R., Habrian, C.H., Zykovich, A., ... Baldwin, E.P. (2013). Quantitative analysis of TALE-DNA interactions suggests polarity effects. Nucleic Acids Research, 41(7), 4118–28. doi:10.1093/nar/gkt085
Applications of BBa_K1189032
User Reviews
UNIQ086cc995662699d2-partinfo-00000001-QINU UNIQ086cc995662699d2-partinfo-00000002-QINU