Composite

Part:BBa_K3504022

Designed by: Ahmed Wael   Group: iGEM20_AFCM-Egypt   (2020-10-25)
Revision as of 18:02, 26 October 2020 by Ahmed Wael (Talk | contribs)


Alphavirus replicon NSPs- SFV

NOTICE: Parts in our range for this season have been created as a part of our Phase I design of our project. These parts HAVE NOT been tested or characterized in the lab due to COVID-19-related precautionary measures. We have enriched our new parts pages with data from literature and results from our modeling and simulations. If you are intending on using this part or others in our range, please keep in mind these limitations and update these parts with data from your experimentation. Feel free to reach us at: igem.afcm@gmail.com for further inquiries.


Part Description

A composite of parts (BBa_K3504013,BBa_K3504014,BBa_K3504015,BBa_K3504016) and CMV promoter which form ,as a unit, the main constituent of alphavirus replicon of semiliki forest virus which as a whole can give the circuit self-replicating ability

Usage

Alphavirus genomes encode four non-basic proteins, nsP1–4, which are translated from the genomic RNA and interact with host factors to form replicative enzyme complexes. These non-structural proteins are translated as a polyprotein that self-cleaves into separate proteins, which assemble into a replicase complex. The replicase then directs replication and amplification of our vaccine inside the Myocytes.

Characterization

We have made simulations using mathematical modelling techniques to characterize the increase in expression when using replicons over traditional methods while also providing simulations that Characterize the function of replicon by eliciting an increased response in both Dendritic Cell population and T-Helper Population.

We also provide Functional characterization of replicons from literature. As This figure shows HIVA-specific T-cell responses after a single immunization with clinical-grade plasmid DNA vaccines between DREP.HIVA and pTHr.HIVA in individual mice immunized by 10 μg of them all of which complies with our mathematical modelling & simulations

Figure 3. Functional characterization of replicons from literature. This figure shows HIVA-specific T-cell responses after a single immunization with clinical-grade plasmid DNA vaccines between DREP.HIVA and pTHr.HIVA in individual mice immunized by 10 μg of them.
Figure 4. Mathematical modelling simulation of Number of positive strand RNA in traditional vaccination presented by the graph to the left vs with the use of self amplifying replicon on the right.
Figure 5. Mathematical modelling simulation of T-helper cells population response according to logfc in response to DREP vaccine on the left vs traditional DNA vaccine on the right.
Figure 6. Mathematical modelling simulation of Dendritic Cells population response according to logfc in response to DREP vaccine on the left vs traditional DNA vaccine on the right.






































References

Maruggi, Giulietta, et al. “Engineered Alphavirus Replicon Vaccines Based on Known Attenuated Viral Mutants Show Limited Effects on Immunogenicity.” Virology, vol. 447, no. 1–2, Dec. 2013, pp. 254–264, 10.1016/j.virol.2013.07.021. Accessed 25 Sept. 2020. Nordström, E. K., Forsell, M. N., Barnfield, C., Bonin, E., Hanke, T., Sundström, M., ... & Liljeström, P. (2005). Enhanced immunogenicity using an alphavirus replicon DNA vaccine against human immunodeficiency virus type 1. Journal of general virology, 86(2), 349-354. openarchive.ki.se/xmlui/bitstream/handle/10616/42305/Thesis_Maria_Lisa_Knudsen.pdf;sequence=3

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 2766
    Illegal EcoRI site found at 4344
    Illegal XbaI site found at 7239
    Illegal PstI site found at 1018
    Illegal PstI site found at 3378
    Illegal PstI site found at 4392
    Illegal PstI site found at 5045
    Illegal PstI site found at 5110
    Illegal PstI site found at 6970
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 2766
    Illegal EcoRI site found at 4344
    Illegal NheI site found at 1224
    Illegal NheI site found at 3065
    Illegal NheI site found at 6732
    Illegal PstI site found at 1018
    Illegal PstI site found at 3378
    Illegal PstI site found at 4392
    Illegal PstI site found at 5045
    Illegal PstI site found at 5110
    Illegal PstI site found at 6970
    Illegal NotI site found at 6736
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 2766
    Illegal EcoRI site found at 4344
    Illegal BglII site found at 7314
    Illegal XhoI site found at 5897
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 2766
    Illegal EcoRI site found at 4344
    Illegal XbaI site found at 7239
    Illegal PstI site found at 1018
    Illegal PstI site found at 3378
    Illegal PstI site found at 4392
    Illegal PstI site found at 5045
    Illegal PstI site found at 5110
    Illegal PstI site found at 6970
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 2766
    Illegal EcoRI site found at 4344
    Illegal XbaI site found at 7239
    Illegal PstI site found at 1018
    Illegal PstI site found at 3378
    Illegal PstI site found at 4392
    Illegal PstI site found at 5045
    Illegal PstI site found at 5110
    Illegal PstI site found at 6970
    Illegal NgoMIV site found at 2211
    Illegal NgoMIV site found at 2448
    Illegal NgoMIV site found at 3558
    Illegal NgoMIV site found at 4295
    Illegal NgoMIV site found at 4305
    Illegal NgoMIV site found at 5971
    Illegal AgeI site found at 5965
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 3115
    Illegal BsaI.rc site found at 4598
    Illegal BsaI.rc site found at 7229


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Parameters
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