Team:Judd UK/Pages/Description
Description
Over 2 billion people worldwide in both MEDCs and LEDCs are affected by anaemia; mainly infants and pregnant women. Iron deficiency symptoms include lethargy, heart conditions, pale skin, reduced cognitive ability and even death. Additionally, many countries give out iron tablets prophylactically to pregnant women and to children, whether they are anaemic or not (because anaemia is such an issue). Side effects of iron overdose are similarly harmful so to be able to home-test and know that iron supplements are not required would be extremely beneficial.
Our iGEM project is to create a home testing kit for detecting iron levels in saliva, which is strongly correlated with iron levels in the blood, to warn people with abnormal concentrations of this essential mineral. The kits will be in the form of slips of paper, which are cell-free systems. These include all the necessary cellular components in cell-free extracts and our genetic construct which are embedded into the paper and freeze-dried. They are easily stored and distributed at a stable room temperature and when necessary can be activated by rehydration. The current method for measuring iron is a full blood count which is expensive, time-consuming and often not available in less economically developed countries. Our test will be cheap and easy to use, making it a more efficient method of testing for iron.
In order to react to different iron levels, our construct will be promoted by the AceB gene, which can be regulated by FUR, a transcriptional repressor of genes involved in iron homeostasis. In the presence of Fe2+ ions, this protein dimerises which allows linkage to a FUR binding site to inhibit mRNA transcription of the downstream gene. To ensure an increase in iron leads to an increase in transcription we will use an inverter, which involves placing a LacI coding region downstream of the FUR regulated promoter. A second plasmid has a promoter that in the presence of LacI inhibits transcription of the chromoprotein coding region downstream. Therefore, in the presence of no iron the iron sensitive promoter allows LacI to be produced, which inhibits transcription of the amilCP chromoprotein on the second plasmid. However, in the presence of iron the iron sensitive promoter is inhibited so LacI is not produced and amilCP can be transcribed to produce a vibrant blue colour in response to iron. Different levels of iron will result in different levels of transcription of the chromoprotein so our device will be semi-quantitative. <a href="https://2017.igem.org/Team:Judd_UK/Achievements">Achievements</a>
References
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[1] Chandran, D. et al. “Mathematical Modeling and Synthetic Biology.” Drug discovery today. Disease models 5.4 (2008): 299–309
[2] Hamed, Mazen Y., J. B. Neilands, and V. Huynh. "Binding of the ferric uptake regulation repressor protein (Fur) to Mn (II), Fe (II), Co (II), and Cu (II) ions as co-repressors: Electronic absorption, equilibrium, and 57 Fe Mössbauer studies." Journal of inorganic biochemistry 50.3 (1993): 193-210
[3] Anne Bagg and J. B. Neilands “Molecular Mechanism of Regulation of Siderophore-Mediated Iron Assimilation.” Microbiological Reviews. Dec. 1987, p. 509-518.
[4] Becskei A and Serrano L "Genetic network driven control of PHBV copolymer composition", J Biotechnol 122(1):99-121, 2006
[5] Deng, Zengqin, et al. "Mechanistic insights into metal ion activation and operator recognition by the ferric uptake regulator." Nature communications 6 (2015).
[6] Basu S et al "A synthetic multicellular system for programmed pattern formation", Nature 434:1130-1134, 2005
[7] Eyal Karzbrun et al “Coarse-Grained Dynamics of Protein Synthesis in a Cell-Free System” Physical Review Letters, Jan 28 2011, PACS numbers: 87.18.Vf
[8] Semsey et al “The effect of LacI autoregulation on the performance of the lactose utilization system in Escherichia coli” 2013 Jul; 41(13): 6381–6390
[9]Selinger, D. W. "Global RNA Half-Life Analysis In Escherichia Coli Reveals Positional Patterns Of Transcript Degradation". Genome Research 13.2 (2003): 216-223. Web. 18 Oct. 2016.
[10] H. Bremer and P. P. Dennis, in Escherichia Coli and Salmonella: Cellular and Molecular Biology, edited by F. C. Neidhardt et al. (ASM Press, Washington, DC, 1996), pp. 1553–1569, 2nd ed.
[11]<a href="https://www.abbexa.com/ecoli-ferric-uptake-regulator-protein-recombinant?gclid=Cj0KCQjw95vPBRDVARIsAKvPd3J1bjFD-mag6B0xR1MofivOa6KgOigQoMOsmw8TbRZZDVBj-_C8WjAaApo5EALw_wcB">https://www.abbexa.com/ecoli-ferric-uptake-regulator-protein-recombinant?gclid=Cj0KCQjw95vPBRDVARIsAKvPd3J1bjFD-mag6B0xR1MofivOa6KgOigQoMOsmw8TbRZZDVBj-_C8WjAaApo5EALw_wcB</a>
[12] Setty Y et al "Detailed map of a cis-regulatory input function", P Natl Acad Sci USA 100(13):7702-7707, 2003
[13] <a href="http://www.nature.com/nature/journal/v274/n5673/abs/274765a0.html">http://www.nature.com/nature/journal/v274/n5673/abs/274765a0.html</a>
[14] <a href="http://parts.igem.org/Part:BBa_K592009">http://parts.igem.org/Part:BBa_K592009</a>
[15] <a href="http://www.uniprot.org/uniprot/P03023">http://www.uniprot.org/uniprot/P03023</a>
[16] <a href="http://parts.igem.org/Part:BBa_K1163003">http://parts.igem.org/Part:BBa_K1163003</a>
[18] <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3711431/#gkt351-B31">https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3711431/#gkt351-B31</a>
[19] <a href="https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3435124/table/t1-mjhid-4-1-e2012051/">https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3435124/table/t1-mjhid-4-1-e2012051/</a>
[20] O.L. Miller, Jr., B. A. Hamkalo, and C. A. Thomas, Jr. “Visualization of Bacterial Genes in Action” Science 169, 392, (1970)
[21] J. Shin abd V. Noireaux, J. Biol. “Study of messenger RNA inactivation and protein degradation in an Escherichia coli cell-free expression system” Eng. 4, 9 (2010)
[22] J. A. Megerle, G. Fritz, U. Gerland, K. Jung, and J. O. Ra¨dler, Biophys. J. 95, 2103 (2008)
[23] <a href="https://slack-redir.net/link?url=http%3A%2F%2Fpubs.acs.org%2Fdoi%2Fabs%2F10.1021%2Fja954077c">https://slack-redir.net/link?url=http%3A%2F%2Fpubs.acs.org%2Fdoi%2Fabs%2F10.1021%2Fja954077c</a>
[24] <a href="http://pubs.acs.org/doi/abs/10.1021/ja00230a041">http://pubs.acs.org/doi/abs/10.1021/ja00230a041</a>
-- Duran Canatan and Sevgi Kosaci Akdeniz; “Iron and Ferritin Levels in Saliva of Patients with Thalassemia and Iron Deficiency Anemia”; Mediterr J Hematol Infect Dis. 2012; 4(1): e2012051.
- -- Vinayak Kumar Mantu, Ruchi Mitra, Jeewan Kumar Mitra; “A comparative study of salivary and serum iron in normal and anaemic female patients in Udaipur city, Rajasthan, India”; Archives of Oral Research, v. 9, n. 3, p. 250-254, Sept./Dec. 2013
- -- Jagannathan N, Thiruvengadam C, Ramani P, Premkumar P, Natesan A, Sherlin HJ; “Salivary ferritin as a predictive marker of iron deficiency anemia in children”; J Clin Pediatr Dent. 2012 Fall;37(1):25-30.
- -- Marzieh Aghazadeh, Hosein Eslami, Ahmad Pirzadeh, Zahra Aghazadeh, Hadiseh Dousti, Parisa Falsafi; “Evaluation of Correlation Between Saliva And Serum Ferretin Level In Patients With Iron Deficiency Anemia And Comparison Group”; IOSR Journal Of Humanities And Social Science (IOSR-JHSS) Volume 21, Issue 6, Ver. 5(June. 2016) PP 54-59.
-- Mishra OP, Agarwal KN, Agarwal RM; “Salivary iron status in children with iron deficiency and iron overload”; J Trop Pediatr. 1992 Apr;38(2):64-7.
- -- Evry 2013 Team; Iron Sensitive Promoter; <a href="http://parts.igem.org/Part:BBa_K1163103">http://parts.igem.org/Part:BBa_K1163103</a> (accessed 22-5-17)
- -- “FerritinCHECK Iron Deficiency Test”; <a href="http://www.stressnomore.co.uk/ferritincheck-iron-deficiency-test-11802.html">www.stressnomore.co.uk/ferritincheck-iron-deficiency-test-11802.html</a> (accessed 22-5-17)
- -- iGEM11_Uppsala-Sweden; “amilCP, blue chromoprotein”; http://parts.igem.org/Part:BBa_K592009 ; 2011-09-18
- -- BioTek; “Quantitation of Green Fluorescent Protein in Microplates using the FL600”; <a href="http://www.biotek.com/resources/application-notes/green-fluorescent-protein-quantification-in-microplates/">www.biotek.com/resources/application-notes/green-fluorescent-protein-quantification-in-microplates/</a>; Rev. 2-20-01
- -- World Health Organisation; “THE GLOBAL PREVALENCE OF ANAEMIA IN 2011”; <a href="http://apps.who.int/iris/bitstream/10665/177094/1/9789241564960_eng.pdf">http://apps.who.int/iris/bitstream/10665/177094/1/9789241564960_eng.pdf</a>; ISBN 978 92 4 156496 0 (NLM classification: WH 155)
- -- Anemia / Iron Deficiency Statistics; <a href="http://www.statisticbrain.com/anemia-iron-deficiency-statistics">http://www.statisticbrain.com/anemia-iron-deficiency-statistics</a> (accessed 14-6-17)
- -- Evry 2013 Team; Notebook; <a href="https://2013.igem.org/Team:Evry/Notebook">https://2013.igem.org/Team:Evry/Notebook</a> (accessed 18-6-17)
- -- N. Kalantari, S. Ghaffari; “Evaluation of Toxicity of Heavy Metals for Escherichia coli Growth”; Iranian Journal of Environmental Health Science & Engineering, 2008, Vol. 5, No. 3, pp. 173-178
