Difference between revisions of "Part:BBa K2984050"
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This vector is a part of the <a href="https://2019.igem.org/Team:Humboldt_Berlin/Part_Collection">Chlamy-HUB-Collection</a>. This part is composed of the PsaD promoter with an expression enhancing intron <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984046">PsaDintron</a>, the <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984045">bleomycin resistance</a> with self cleaving peptide scp, the secretion signal <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984000">ARS</a>, the <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K3002037">MHETase enzyme</a>, kindly provided by the TU Kaiserslautern 2019 team, the secretion enhancing <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984015">SP20</a> glycomodule, and the <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984021">Rbcs2 terminator</a>. | This vector is a part of the <a href="https://2019.igem.org/Team:Humboldt_Berlin/Part_Collection">Chlamy-HUB-Collection</a>. This part is composed of the PsaD promoter with an expression enhancing intron <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984046">PsaDintron</a>, the <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984045">bleomycin resistance</a> with self cleaving peptide scp, the secretion signal <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984000">ARS</a>, the <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K3002037">MHETase enzyme</a>, kindly provided by the TU Kaiserslautern 2019 team, the secretion enhancing <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984015">SP20</a> glycomodule, and the <a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K2984021">Rbcs2 terminator</a>. | ||
| − | The part can be used to express and secrete the MHETase enzyme in C. reinhardtii. The bleomycin resistance leads to a higher expression of the enzyme, due to its working mechanism. For every two bleomycin molecules, the bleomycin resistance gene must be expressed once. The resistance works in a ratio of 1:2 to the antibiotic. By having the resistance in the same open reading frame, the enzyme is also expressed, leading to a higher expression of the enzyme. | + | The part can be used to express and secrete the MHETase enzyme in C. reinhardtii. The bleomycin resistance leads to a higher expression of the enzyme, due to its working mechanism. For every two bleomycin molecules, the bleomycin resistance gene must be expressed once. The resistance works in a ratio of 1:2 to the antibiotic (Hayes et al., 1990). By having the resistance in the same open reading frame, the enzyme is also expressed, leading to a higher expression of the enzyme (Lumbreras et al. 1998). |
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<img src="https://2019.igem.org/wiki/images/3/3b/T--Humboldt_Berlin--Konstruk_15.jpg" alt="colonies_total" width="500"> | <img src="https://2019.igem.org/wiki/images/3/3b/T--Humboldt_Berlin--Konstruk_15.jpg" alt="colonies_total" width="500"> | ||
| − | <figcaption>Fig.1 - Image of a successful transformation in E.coli after ligation of the construct. The construct can then be isolated from E. coli to be transformed in C. reinhardtii</figcaption> | + | <figcaption>Fig.1 - Image of a successful transformation in <i>E. coli</i> after ligation of the construct. The construct can then be isolated from <i>E. coli</i> to be transformed in <i>C. reinhardtii</i>.</figcaption> |
</figure> | </figure> | ||
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<partinfo>BBa_K2984050 parameters</partinfo> | <partinfo>BBa_K2984050 parameters</partinfo> | ||
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| + | |||
| + | ==References== | ||
| + | <ol> | ||
| + | <li> | ||
| + | Lumbreras, V., Stevens, D. R., & Purton, S. (1998). Efficient foreign gene expression in <i>Chlamydomonas reinhardtii</i> mediated by an endogenous intron. The Plant Journal, 14(4), 441-447. | ||
| + | </li> | ||
| + | <li> | ||
| + | Hayes, J. D., & Wolf, C. R. (1990). Molecular mechanisms of drug resistance. Biochemical Journal, 272(2), 281. | ||
| + | </li> | ||
| + | </ol> | ||
Latest revision as of 23:01, 19 October 2019
L1c-PsaDintron-bleRscp-ARS-MHETase-SP20-RbcS2
This vector is a part of the Chlamy-HUB-Collection. This part is composed of the PsaD promoter with an expression enhancing intron PsaDintron, the bleomycin resistance with self cleaving peptide scp, the secretion signal ARS, the MHETase enzyme, kindly provided by the TU Kaiserslautern 2019 team, the secretion enhancing SP20 glycomodule, and the Rbcs2 terminator. The part can be used to express and secrete the MHETase enzyme in C. reinhardtii. The bleomycin resistance leads to a higher expression of the enzyme, due to its working mechanism. For every two bleomycin molecules, the bleomycin resistance gene must be expressed once. The resistance works in a ratio of 1:2 to the antibiotic (Hayes et al., 1990). By having the resistance in the same open reading frame, the enzyme is also expressed, leading to a higher expression of the enzyme (Lumbreras et al. 1998).
Sequence and Features
Assembly Compatibility:
- 10INCOMPATIBLE WITH RFC[10]Illegal PstI site found at 2614
Illegal PstI site found at 2938
Illegal PstI site found at 3281
Illegal PstI site found at 4091 - 12INCOMPATIBLE WITH RFC[12]Illegal PstI site found at 2614
Illegal PstI site found at 2938
Illegal PstI site found at 3281
Illegal PstI site found at 4091
Illegal NotI site found at 2949 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 3859
Illegal BamHI site found at 4 - 23INCOMPATIBLE WITH RFC[23]Illegal PstI site found at 2614
Illegal PstI site found at 2938
Illegal PstI site found at 3281
Illegal PstI site found at 4091 - 25INCOMPATIBLE WITH RFC[25]Illegal PstI site found at 2614
Illegal PstI site found at 2938
Illegal PstI site found at 3281
Illegal PstI site found at 4091
Illegal NgoMIV site found at 1607
Illegal NgoMIV site found at 2547
Illegal NgoMIV site found at 3008
Illegal NgoMIV site found at 3038
Illegal NgoMIV site found at 3353
Illegal NgoMIV site found at 3371
Illegal NgoMIV site found at 3407 - 1000COMPATIBLE WITH RFC[1000]
Characterization
References
- Lumbreras, V., Stevens, D. R., & Purton, S. (1998). Efficient foreign gene expression in Chlamydomonas reinhardtii mediated by an endogenous intron. The Plant Journal, 14(4), 441-447.
- Hayes, J. D., & Wolf, C. R. (1990). Molecular mechanisms of drug resistance. Biochemical Journal, 272(2), 281.