Team:Aix-Marseille/Part Collection
Part Collection
Our collection of parts is designed to allow the creation of proteins, or phage-like particles, that target a wide range of different Gram-negative bacteria. Each part corresponds to the domains 1 (D1) and 2 (D2) from the p3 protein of filamentous phages targeting the different organisms. The parts in the collection are expected to be used as fusion proteins and so conform to the [http://parts.igem.org/Assembly_standard_25 Rfc25] standard. To have more detail about the design of these parts, and how we used them to make phage-like particles in our project, you can check out our design page.
This part collection contains the following biobricks all in [http://parts.igem.org/Assembly_standard_25 Rfc25] format:
- p3_E.coli - [http://parts.igem.org/Part:BBa_K2255008 BBa_K2255008]
- p3_N.gonorrhoeae - [http://parts.igem.org/Part:BBa_K2255009 BBa_K2255009]
- p3_P.aeruginosa - [http://parts.igem.org/Part:BBa_K2255010 BBa_K2255010]
- p3_R.solanacearum_RSM1 - [http://parts.igem.org/Part:BBa_K2255011 BBa_K2255011]
- p3_R.solanacearum_RSS1 - [http://parts.igem.org/Part:BBa_K2255012 BBa_K2255012]
- p3_V.Cholerae_CTXΦ - [http://parts.igem.org/Part:BBa_K2255013 BBa_K2255013]
- p3_V.Cholerae_fs2 - [http://parts.igem.org/Part:BBa_K2255014 BBa_K2255014]
- p3_V.Cholerea_VGJΦ - [http://parts.igem.org/Part:BBa_K2255015 BBa_K2255015]
- p3_X.campestris - [http://parts.igem.org/Part:BBa_K2255016 BBa_K2255016]
- p3_X.fastidiosa - [http://parts.igem.org/Part:BBa_K2255017 BBa_K2255017]
- p3_X.fuscans - [http://parts.igem.org/Part:BBa_K2255018 BBa_K2255018]
Usage
These biobricks will be used to engineer phage-like particles targeting different bacteria. Check out our design page to know more about their design.
Design
The domain 3 (D3) and the signal sequence are the best conserved parts of the innophage attachment protein p3. Using a global protein sequence alignment (Needleman-Wunsch and MUSCLE alignments), using two or three sequence at a time, we were able create a good alignment. We used this to determine the domain 1 (D1) and domain 2 (D2) sequence for the attachment proteins of each phages.
We were able to find these domains because they are separated by flexible sequences, often composed of Glycine and Serine [1]. Then we then retrotranslated these protein sequences to obtain gene sequences and we used the iDT tool to optimise this for production in E.coli.
| Pathogene | Filamentous phage | GI | Part ID |
|---|---|---|---|
| Escherichia coli | M13 (fd,ff)[2] | 927334 | [http://parts.igem.org/Part:BBa_K2255008 BBa_K2255008] |
| Neisseria gonorrheae | NgoΦ6[3] | 1260906 | [http://parts.igem.org/Part:BBa_K2255009 BBa_K2255009] |
| Pseudomonas aeruginosa | Pf3[4] | 215374 | [http://parts.igem.org/Part:BBa_K2255010 BBa_K2255010] |
| Ralstonia solanacearum | RSM1Φ[5] | 5179368 | [http://parts.igem.org/Part:BBa_K2255011 BBa_K2255011] |
| RSS1Φ[5] | 4525385 | [http://parts.igem.org/Part:BBa_K2255012 BBa_K2255012] | |
| Vibrio Cholerea | CTXΦ[6] | 26673076 | [http://parts.igem.org/Part:BBa_K2255013 BBa_K2255013] |
| VFJΦ(fs2)[7] | 1261866 | [http://parts.igem.org/Part:BBa_K2255014 BBa_K2255014] | |
| VGJΦ[8] | 1260523 | [http://parts.igem.org/Part:BBa_K2255015 BBa_K2255015] | |
| Xanthomonas campestris | ΦLf[9] | 3730653 | [http://parts.igem.org/Part:BBa_K2255016 BBa_K2255016] |
| Xylella fastidiosa | XfasM23[10] | 6203562 | [http://parts.igem.org/Part:BBa_K2255017 BBa_K2255017] |
| Xanthomonas fucans | XacF1[11] | 17150318 | [http://parts.igem.org/Part:BBa_K2255018 BBa_K2255018] |
- ↑ Heilpern, A. J. & Waldor, M. K. pIIICTX, a predicted CTXphi minor coat protein, can expand the host range of coliphage fd to include Vibrio cholerae. J. Bacteriol. 185, 1037–1044 (2003).
- ↑ Smeal, S. W., Schmitt, M. A., Pereira, R. R., Prasad, A. & Fisk, J. D. Simulation of the M13 life cycle I: Assembly of a genetically-structured deterministic chemical kinetic simulation. Virology 500, 259–274 (2017).
- ↑ Piekarowicz, A. et al. Neisseria gonorrhoeae Filamentous Phage NgoΦ6 Is Capable of Infecting a Variety of Gram-Negative Bacteria. J Virol 88, 1002–1010 (2014).
- ↑ Luiten, R. G., Schoenmakers, J. G. & Konings, R. N. The major coat protein gene of the filamentous Pseudomonas aeruginosa phage Pf3: absence of an N-terminal leader signal sequence. Nucleic Acids Res 11, 8073–8085 (1983).
- ↑ 5.0 5.1 T, K. et al. Genomic characterization of the filamentous integrative bacteriophages {phi}RSS1 and {phi}RSM1, which infect Ralstonia solanacearum., Genomic Characterization of the Filamentous Integrative Bacteriophages φRSS1 and φRSM1, Which Infect Ralstonia solanacearum. J Bacteriol 189, 189, 5792, 5792–5802 (2007).
- ↑ Heilpern, A. J. & Waldor, M. K. pIIICTX, a predicted CTXphi minor coat protein, can expand the host range of coliphage fd to include Vibrio cholerae. J. Bacteriol. 185, 1037–1044 (2003).
- ↑ Ikema, M. & Honma, Y. A novel filamentous phage, fs-2, of Vibrio cholerae O139. Microbiology 144, 1901–1906 (1998).
- ↑ Campos, J. et al. VGJφ, a Novel Filamentous Phage of Vibrio cholerae, Integrates into the Same Chromosomal Site as CTXφ. J. Bacteriol. 185, 5685–5696 (2003).
- ↑ Tseng, Y.-H., Lo, M.-C., Lin, K.-C., Pan, C.-C. & Chang, R.-Y. Characterization of filamentous bacteriophage ΦLf from Xanthomonas campestris pv. campestris. Journal of general virology 71, 1881–1884 (1990).
- ↑ Chen, J. & Civerolo, E. L. Morphological evidence for phages in Xylella fastidiosa. Virology Journal 5, 75 (2008).
- ↑ Ahmad, A. A., Askora, A., Kawasaki, T., Fujie, M. & Yamada, T. The filamentous phage XacF1 causes loss of virulence in Xanthomonas axonopodis pv. citri, the causative agent of citrus canker disease. Front. Microbiol. 5, (2014).
