Attributions and References

iGEM Toronto would like to acknowledge all the hardwork each member puts into this project. We would like to thank people outside of our team who helped us along the way.

iGEM Toronto Team Member Attributions

The whole team performed background research, project scoping, and goal settings together at the beginning of the project.

Lead Team

  • Project Administration: managing and coordinating the project activities, planning, and execution

    • Wet Lab

    The whole lab designed and constructed experiments. Throughout the summer, all wet lab members performed experiments in the University of Toronto IBME lab. During our meetings, we cleaned, discussed, and interpreted our data. Based on the results, we decided on our next steps.

    • Writing:
      • Results summary and interpretation: Ruochen Liu
      • Safety: Sathwik, Ally Cheung
      • Experimental story and protocol rationale: Jaden Bhogal, Amogh Manivannan
      • Fluorescence LAMP Data Analysis: Zihou Ye

    Dry Lab

    • Primer Designs: Ruochen Liu, Alston Lo, Maggie Chen, Marry Xuan
    • Primer Scorer: Dev Patel, Rodrigo Reyes, Victoria Gao, Maggie Chen, Alston Lo
    • Writing:
      • Difficulties and solutions: Maggie Chen
      • What makes a good primer set: Mohammad
      • Reference materials: Victoria Gao
      • Primer scorer: Victoria Gao
      • Workflow: Rodrigo Reyes

    Hardware

    The hardware team brainstormed ideas together to decide how to tackle the problem in the given timeframe. After that, the hardware team was splited into three groups to engineer the sample extraction tool, reagents mixer and the portable heater.

    • Sample extraction Tool: Blythe, Robert, Kyrylo, Miranda
    • Reagents mixer: Blythe, Robert, Miranda
    • Portable Heater: Vishwa, Kyrylo, Tuo, Miranda
    • Writing:
      • The dipstick experiment: Tuo, Miranda
      • Printing and testing the DNA extraction tool: Blythe Huang, Robert, Miranda
      • Designing, building, and testing the reagent mixer: Vishwa, Blythe Huang, Robert
      • How does the heater work: Vishwa, Miranda, Kyrylo
      • Designing, building, and testing the sample extraction tool: Kyrylo
      • Workflow: Tuo

    Human Practice

    • Preparing and/or implementing tools/activities to engage the broader community: Geumbin, Liliane, Sean
    • Reaching out to stakeholders and communicating with them: Blythe, Michelle, Liliane, Sean
    • Writing:
      • Education, Communication and Public Engagement: Geumbin, Liliane, Sean
      • Stakeholders: Blythe, Michelle, Liliane, Sean

    iGEM Deliverables

  • Safety Form: Ally, Sathwik, Sean
  • Judging Form: Zihou Ye, Sean Yam, Maggie Chen, Ally
  • Parts and writing about the Parts: Marry Xuan
  • Title and Abstract: Wanda, Ally, Rodrigo, Sean
  • Developing Wiki: Brendan, Ally, Dev, Tuo, Miranda, Paul

    • Acknowledgements

    Thank you to all the people involved in helping make iGEM Toronto successful!

    Project support and advice


    Keith Pardee, PhD

    Associate Professor at the Faculty of Pharmacy, University of Toronto

    Dr. Pardee meets with iGEM Toronto regularly from the beginning of the project. He refined our ideas with his expertise in molecular diagnostics. He shared the painpoints that his research lab went through and taught us how to avoid them. He recommended us to look into LAMP for DNA detection.


    Jennifer Doucet, PhD

    Postdoctoral Fellow at the Pardee's Lab, University of Toronto

    Jennifer supervised and supported iGEM Toronto's fluorescence LAMP using qPCR machine experiments and the freeze-dried LAMP reaction experiments. She was incredibly helpful in helping us understand how to intrepret and report our lab results. She advised us on troubleshooting LAMP experiments and provided us a lot of suggestions on how to minimize contaminations.


    Marcus Dillon, PhD

    Associate Professor at the Department of Ecology & Evolutionary Biology, University of Toronto

    Dr. Dillon was the first person pointed out that our initial proposed solution had added complexity with the toehold switch. He recommended us to look into DNA detection methods. He was extremely insightful in primer designs. He advised us to research and focus on DNA regions that are highly conserved and divergent in other species. We learnt about the trade off between selectivity and specificity from Dr. Dillon.


    A graduate student at the Pardee Lab, PhD

    Faculty of Pharmacy, University of Toronto

    She was incredibly helpful in helping us understand how to intrepret and report our lab results. She advised us on troubleshooting LAMP experiments and provided us directions on how to design LAMP experiments at the beginning of the project.

    Lab Support


    Gary Hoang

    Teaching Lab & Design Studio Technician at Institute of Biomedical Engineering, Univeristy of Toronto

    Gary provided us with lab space, common lab reagents and supplies. He ensured our lab members adhere to the biosafety standards of the University of Toronto Environmental Health and Safety Department. He provided us advises on the protocol and lab techniques. He was extremely patient with the questions our team had with our experiments. iGEM Toronto is very thankful for his support in the lab over the summer.


    Yan Wang, PhD

    Assistant Professor in the Department of Biological Sciences at University of Toronto Scarborough

    Dr. Wang provided us with his lab space, common lab reagents, fungal sample and supplies. He supervised us on the testing of dipstick DNA extraction method using PCR and gel electrophoresis along side with Phd student Huimei Yang.

    Human Practices support

    We would like to acknowledge the people we interviewed, who provided a lot of insights on the human practice. See Human Practices for details.


  • Tim Payne
  • A forester in Michigan
  • Dr. Sandy Smith
  • Mackenzie DiGasparro
  • Let's Talk Science - University of Toronto Branch
  • University of Toronto Engineering Student Society
  • iGEM Aboa
  • iGEM Trigate
  • iGEM HKUST


    • References

    Wet Lab

    Notomi, T., Okayama, H., Masubuchi, H., Yonekawa, T., Watanabe, K., Amino, N., & Hase, T. (2000). Loop-mediated isothermal amplification of DNA. Nucleic acids research, 28(12), E63. https://doi.org/10.1093/nar/28.12.e63

    Nagamine, K., Hase, T., & Totomi, T. (2002). Accelerated reaction by loop-mediated isothermal amplification using loop primers. Molecular and Cellular Probes, 16, 223–229. 10.1006/mcpr.2002.0415

    Pardee, K., Green, A. A., Takahashi, M. K., Braff, D., Lambert, G., Lee, J. W., Ferrante, T., et al. (2016). Rapid, low-cost detection of Zika virus using programmable biomolecular components. Cell, 165(5), 1255–1266. https://doi.org/10.1016/j.cell.2016.04.059

    Tanner, N. A., Zhang, Y., & Evans, T. C., Jr (2015). Visual detection of isothermal nucleic acid amplification using pH-sensitive dyes. BioTechniques, 58(2), 59–68. https://doi.org/10.2144/000114253

    Zanoli, L. M., & Spoto, G. (2013). Isothermal amplification methods for the detection of nucleic acids in microfluidic devices. Biosensors, 3(1), 18–43. https://doi.org/10.3390/bios3010018

    Lu, S., Duplat, D., Benitez-Bolivar, P., Leo´n, C., Villota, S. D., Veloz-Villavicencio, E., et al. (2022). Multicenter international assessment of a SARSCoV-2 RT-LAMP test for point of care clinical application. PLoS ONE, 17(5): e0268340. https://doi.org/10.1371/journal.pone.0268340

    Pardee, K., Green, A. A., Ferrante, T., Cameron, D. E., DaleyKeyser, A., Yin, P., & Collins, J. J. (2014). Paper-based synthetic gene networks. Cell, 159(4), 940–954. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4243060/pdf/nihms634354.pdf

    Reboud, J., Xu, G., Garrett, A., Adriko, M., Yang, Z., Tukahebwa, E. M., Rowell, C., & Cooper, J. M. (2019). Paper-based microfluidics for DNA diagnostics of malaria in low resource underserved rural communities. Proceedings of the National Academy of Sciences of the United States of America, 116(11), 4834–4842. https://doi.org/10.1073/pnas.1812296116.

    Wang, J., Dextre, A., Fascual-Garrigos, A, et al. (2021). Fabrication of a paper-based colorimetric molecular test for SARS-CoV-2. MethodsX, 8, 101586. https://doi.org/10.1016/j.mex.2021.101586

    Ye, J., Coulouris, G., Zaretskaya, I., Cutcutache, I., Rozen, S., & Madden, T. L. (2012). Primer-BLAST: a tool to design target-specific primers for polymerase chain reaction. BMC bioinformatics, 13, 134. https://doi.org/10.1186/1471-2105-13-134

    NCBI search for C. douglasii: https://www.ncbi.nlm.nih.gov/nuccore/?term=Ceratocystis+douglasii

    Modelling

    Blaser, Simon et al. "A Loop-Mediated Isothermal Amplification (LAMP) Assay For Rapid Identification Of ≪Em≫Bemisia Tabaci≪/Em≫". Journal Of Visualized Experiments, no. 140, 2018. Myjove Corporation, https://doi.org/10.3791/58502.

    Eiken Chemical Co., Ltd. “A Guide to LAMP primer designing (PrimerExplorer V5).” 15 Oct. 2019

    Jia, Ben et al. “GLAPD: Whole Genome Based LAMP Primer Design for a Set of Target Genomes.” Frontiers in microbiology vol. 10 2860. 13 Dec. 2019, doi:10.3389/fmicb.2019.02860

    LucigenVideo. Loop-Mediated Isothermal Amplification (LAMP): Primer Design And Assay Optimization. 2018, https://www.youtube.com/watch?v=GJkvQqDufh0.

    Metsky, Hayden C., et al. “Designing Sensitive Viral Diagnostics with Machine Learning.” Nature Biotechnology, vol. 40, no. 7, 2022, pp. 1123–1131., https://doi.org/10.1038/s41587-022-01213-5.

    Schoch, Conrad L. et al. "Nuclear Ribosomal Internal Transcribed Spacer (ITS) Region As A Universal DNA Barcode Marker For Fungi". Proceedings Of The National Academy Of Sciences, vol 109, no. 16, 2012, pp. 6241-6246. Proceedings Of The National Academy Of Sciences, https://doi.org/10.1073/pnas.1117018109.

    Si Ammour, Melissa et al. "Use Of LAMP For Assessing Botrytis Cinerea Colonization Of Bunch Trash And Latent Infection Of Berries In Grapevines". Plants, vol 9, no. 11, 2020, p. 1538. MDPI AG, https://doi.org/10.3390/plants9111538.

    Silva Zatti, Matheus et al. "Isothermal Nucleic Acid Amplification Techniques For Detection And Identification Of Pathogenic Fungi: A Review". Mycoses, vol 63, no. 10, 2020, pp. 1006-1020. Wiley, https://doi.org/10.1111/myc.13140.

    Wu, C. P. et al. "Rapid And Accurate Detection Of Ceratocystis Fagacearum From Stained Wood And Soil By Nested And Real-Time PCR". *Forest Pathology*, vol 41, no. 1, 2011, pp. 15-21. *Wiley*, https://doi.org/10.1111/j.1439-0329.2009.00628.x.

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