Difference between revisions of "Part:BBa K5477027"
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<partinfo>BBa_K5477027 short</partinfo> | <partinfo>BBa_K5477027 short</partinfo> | ||
− | The pRET2-LexA-ERα(LBD) composite part integrates the LexA DNA-binding domain (DBD) with the ligand-binding domain (LBD) of Estrogen Receptor Alpha (ERα), under the control of the pRET2 promoter. This | + | The pRET2-LexA-ERα(LBD) composite part integrates the LexA DNA-binding domain (DBD) with the ligand-binding domain (LBD) of Estrogen Receptor Alpha (ERα) [https://parts.igem.org/Part:BBa_K5477013| BBa_K5477013], under the control of the pRET2 promoter [https://parts.igem.org/Part:BBa_K5477000| BBa_K5477000]. This allows for the precise regulation of gene expression in response to estrogen or estrogen-like molecules, such as bisphenol A (BPA) (1) (2) (3) (4). The pRET2 promoter controls the expression of this fusion protein, ensuring that the LexA-ERα(LBD) is expressed under specific conditions determined by the regulatory elements of the pRET2 sequence. Upon presence of BPA, the LexA-ERα(LBD) binds specifically to LexA operator sequences (Lex6Op) which is a part of our reporter module which initiates transcription of NanoLuc. |
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+ | The composite part was cloned using the method of USER-cloning into YCp-L. The YCp-L plasmid is a centromeric plasmid that carries a LEU2 marker for leucine selection in yeast. Centromeric plasmids like YCp-L are low-copy vectors, typically maintained at one to two copies per cell due to the presence of a CEN sequence. This composite part was used in our device: | ||
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<partinfo>BBa_K5477027 parameters</partinfo> | <partinfo>BBa_K5477027 parameters</partinfo> | ||
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+ | ===References=== | ||
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+ | 1. Hafezi SA, Abdel-Rahman WM. The Endocrine Disruptor Bisphenol A (BPA) Exerts a Wide Range of Effects in Carcinogenesis and Response to Therapy. Curr Mol Pharmacol. 2019;12(3):230-238. doi: 10.2174/1874467212666190306164507. PMID: 30848227; PMCID: PMC6864600. | ||
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+ | 2. Lo, Elena & Piparo, Elena & Siragusa, Lydia & Raymond, Frédéric & Passeri, Giovanna & Cruciani, Gabriele & Schilter, Benoit. (2019). Bisphenol A Binding Promiscuity: A Virtual Journey through the Universe of Proteins 1. | ||
+ | |||
+ | 3. Park, Choa & Song, Heewon & Choi, Junyeong & Sim, Seunghye & Kojima, Hiroyuki & Park, Joonwoo & Iida, Mitsuru & Lee, Youngjoo. (2020). The mixture effects of bisphenol derivatives on estrogen receptor and androgen receptor. Environmental Pollution. 260. 114036. 10.1016/j.envpol.2020.114036. | ||
+ | |||
+ | 4. Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x |
Revision as of 20:00, 26 September 2024
pRET2-LexA-ERα(LBD) - receptor module
The pRET2-LexA-ERα(LBD) composite part integrates the LexA DNA-binding domain (DBD) with the ligand-binding domain (LBD) of Estrogen Receptor Alpha (ERα) BBa_K5477013, under the control of the pRET2 promoter BBa_K5477000. This allows for the precise regulation of gene expression in response to estrogen or estrogen-like molecules, such as bisphenol A (BPA) (1) (2) (3) (4). The pRET2 promoter controls the expression of this fusion protein, ensuring that the LexA-ERα(LBD) is expressed under specific conditions determined by the regulatory elements of the pRET2 sequence. Upon presence of BPA, the LexA-ERα(LBD) binds specifically to LexA operator sequences (Lex6Op) which is a part of our reporter module which initiates transcription of NanoLuc.
The composite part was cloned using the method of USER-cloning into YCp-L. The YCp-L plasmid is a centromeric plasmid that carries a LEU2 marker for leucine selection in yeast. Centromeric plasmids like YCp-L are low-copy vectors, typically maintained at one to two copies per cell due to the presence of a CEN sequence. This composite part was used in our device:
Sequence and Features
- 10INCOMPATIBLE WITH RFC[10]Illegal XbaI site found at 1606
Illegal PstI site found at 1790
Illegal PstI site found at 1961 - 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 182
Illegal PstI site found at 1790
Illegal PstI site found at 1961 - 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 1740
- 23INCOMPATIBLE WITH RFC[23]Illegal XbaI site found at 1606
Illegal PstI site found at 1790
Illegal PstI site found at 1961 - 25INCOMPATIBLE WITH RFC[25]Illegal XbaI site found at 1606
Illegal PstI site found at 1790
Illegal PstI site found at 1961
Illegal AgeI site found at 37 - 1000COMPATIBLE WITH RFC[1000]
References
1. Hafezi SA, Abdel-Rahman WM. The Endocrine Disruptor Bisphenol A (BPA) Exerts a Wide Range of Effects in Carcinogenesis and Response to Therapy. Curr Mol Pharmacol. 2019;12(3):230-238. doi: 10.2174/1874467212666190306164507. PMID: 30848227; PMCID: PMC6864600.
2. Lo, Elena & Piparo, Elena & Siragusa, Lydia & Raymond, Frédéric & Passeri, Giovanna & Cruciani, Gabriele & Schilter, Benoit. (2019). Bisphenol A Binding Promiscuity: A Virtual Journey through the Universe of Proteins 1.
3. Park, Choa & Song, Heewon & Choi, Junyeong & Sim, Seunghye & Kojima, Hiroyuki & Park, Joonwoo & Iida, Mitsuru & Lee, Youngjoo. (2020). The mixture effects of bisphenol derivatives on estrogen receptor and androgen receptor. Environmental Pollution. 260. 114036. 10.1016/j.envpol.2020.114036.
4. Zhou, T., Liang, Z. & Marchisio, M.A. Engineering a two-gene system to operate as a highly sensitive biosensor or a sharp switch upon induction with β-estradiol. Sci Rep 12, 21791 (2022). https://doi.org/10.1038/s41598-022-26195-x