Difference between revisions of "Part:BBa K4321009:Design"
(→References) |
|||
| Line 7: | Line 7: | ||
===Design Notes=== | ===Design Notes=== | ||
| + | We designed our cassettes with flanking BsaI restriction sites to clone our cassettes downstream of a strong artificial promoter named Pgrac that was contained within our E.coli to Bacillus subtilis shuttle plasmid, pCG004. This cassette however, does not contain these restriction sites or the Pgrac feature so that future iGEM teams may clone this cassette into their desired plasmid under the control of their chosen promoter. | ||
| + | |||
Cyt2Ba was design upstream of a RBS, a report fluorescent protein variant BFP (BBa_K4321001), and a Bacillus subtilis specific terminator. For our project we designed this cassette with flanking BsaI sites (included in cassette), for insertion into pCG004. However, desired restriction sites can be added to both ends of the cassette for desired plasmid insertion. Following the digestion with BsaI the last two bases will be lost to yield a 1682 bp fragment. | Cyt2Ba was design upstream of a RBS, a report fluorescent protein variant BFP (BBa_K4321001), and a Bacillus subtilis specific terminator. For our project we designed this cassette with flanking BsaI sites (included in cassette), for insertion into pCG004. However, desired restriction sites can be added to both ends of the cassette for desired plasmid insertion. Following the digestion with BsaI the last two bases will be lost to yield a 1682 bp fragment. | ||
Revision as of 16:05, 11 October 2022
Cyt2Ba Cassette
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
We designed our cassettes with flanking BsaI restriction sites to clone our cassettes downstream of a strong artificial promoter named Pgrac that was contained within our E.coli to Bacillus subtilis shuttle plasmid, pCG004. This cassette however, does not contain these restriction sites or the Pgrac feature so that future iGEM teams may clone this cassette into their desired plasmid under the control of their chosen promoter.
Cyt2Ba was design upstream of a RBS, a report fluorescent protein variant BFP (BBa_K4321001), and a Bacillus subtilis specific terminator. For our project we designed this cassette with flanking BsaI sites (included in cassette), for insertion into pCG004. However, desired restriction sites can be added to both ends of the cassette for desired plasmid insertion. Following the digestion with BsaI the last two bases will be lost to yield a 1682 bp fragment.
Source
The cassette was synthesized by IDT. Information on the origin of each part can be found on their respective part pages: Cyt2Ba - BBa_K4321000, RBS - BBa_B0034, BFP - BBa_K4321001, Terminator - BBa_K4321008.
References
Cohen, S., Dym, O., Albeck, S., Ben-Dov, E., Cahan, R., Firer, M., & Zaritsky, A. (2008). High-resolution crystal structure of activated Cyt2Ba monomer from Bacillus thuringiensis subsp. israelensis. Journal of molecular biology, 380(5), 820–827. https://doi.org/10.1016/j.jmb.2008.05.010
Soberón, M., López-Díaz, J. A., & Bravo, A. (2013). Cyt toxins produced by bacillus thuringiensis: A protein fold conserved in several pathogenic microorganisms. Peptides, 41, 87–93. https://doi.org/10.1016/j.peptides.2012.05.023
Valtierra-de-Luis, D., Villanueva, M., Berry, C., & Caballero, P. (2020). Potential for bacillus thuringiensis and other bacterial toxins as biological control agents to combat dipteran pests of medical and agronomic importance. Toxins, 12(12), 773. https://doi.org/10.3390/toxins12120773
Wang, FF., Qu, SX., Lin, JS. et al. Identification of Cyt2Ba from a New Strain of Bacillus thuringiensis and Its Toxicity in Bradysia difformis. Curr Microbiol 77, 2859–2866 (2020). https://doi.org/10.1007/s00284-020-02018-y