Difference between revisions of "Part:BBa K2151200"
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<h1>'''Fudan 2022 iGEM team'''</h1> | <h1>'''Fudan 2022 iGEM team'''</h1> | ||
| − | As many researches indicate, the major problem of polycistronic vectors, which contain two or more target genes under one promoter, is the much lower expression of the downstream genes compared with that of the first gene next to the promoter<ref>Kim, K. J., Kim, H. E., Lee, K. H., Han, W., Yi, M. J., Jeong, J., & Oh, B. H. (2004). Two-promoter vector is highly efficient for overproduction of protein complexes. Protein science : a publication of the Protein Society, 13(6), 1698–1703.</ref>. | + | As many researches indicate, the major problem of polycistronic vectors, which contain two or more target genes under one promoter, is the much lower expression of the downstream genes compared with that of the first gene next to the promoter<ref>Kim, K. J., Kim, H. E., Lee, K. H., Han, W., Yi, M. J., Jeong, J., & Oh, B. H. (2004). Two-promoter vector is highly efficient for overproduction of protein complexes. Protein science : a publication of the Protein Society, 13(6), 1698–1703.</ref>. Instead of assembling CDSs sequentially, we construct a ribozyme-assisted polycistronic co-expression system (pRAP) by inserting ribozyme sequences between crtEBIY. In the pRAP system, the RNA sequences of hammerhead ribozyme conduct self-cleaving, and the polycistronic mRNA transcript is thus co-transcriptionally converted into individual mono-cistrons ''in vivo''. Self-interaction of the polycistron can be nullified and each cistron can initiate translation with comparable efficiency. Besides, we can precisely manage this co-expression system by adjusting the RBS strength of individual mono-cistrons. |
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<h2>'''References'''</h2> | <h2>'''References'''</h2> | ||
<references /> | <references /> | ||
Revision as of 08:02, 12 October 2022
crtEBIY
. This biobrick was created through standard biobrick assembly of K118014(RBS+crtE), K118006(RBS+crtB), K118005(RBS+crtI) and K118013(crtY). These genes are a part of the carotenoid biosynthesis pathway and together, this biobrick converts converts colourless farnesyl pyrophosphate to orange beta-carotene Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 1974
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 1510
Illegal NgoMIV site found at 1640
Illegal AgeI site found at 725 - 1000COMPATIBLE WITH RFC[1000]
Fudan 2022 iGEM team
As many researches indicate, the major problem of polycistronic vectors, which contain two or more target genes under one promoter, is the much lower expression of the downstream genes compared with that of the first gene next to the promoter[1]. Instead of assembling CDSs sequentially, we construct a ribozyme-assisted polycistronic co-expression system (pRAP) by inserting ribozyme sequences between crtEBIY. In the pRAP system, the RNA sequences of hammerhead ribozyme conduct self-cleaving, and the polycistronic mRNA transcript is thus co-transcriptionally converted into individual mono-cistrons in vivo. Self-interaction of the polycistron can be nullified and each cistron can initiate translation with comparable efficiency. Besides, we can precisely manage this co-expression system by adjusting the RBS strength of individual mono-cistrons.
References
- ↑ Kim, K. J., Kim, H. E., Lee, K. H., Han, W., Yi, M. J., Jeong, J., & Oh, B. H. (2004). Two-promoter vector is highly efficient for overproduction of protein complexes. Protein science : a publication of the Protein Society, 13(6), 1698–1703.