Bio detection of anoxia in Lake Xochimilco using modified Escherichia - PowerPoint PPT Presentation

Bio detection of anoxia in Lake Xochimilco using modified Escherichia coli by GenetiX

Instituto Tecnológico y de Estudios Superiores de Monterrey, Campus Ciudad de México

Our Team

Background  Xochimilco: • Source of water for the Metropolitan Area • Highly touristic and agricultural element • World Heritage Site • Local endemic species in danger of extinction • Highly polluted with indutrial residues • Critical levels of d O 2 below minimun requirements • Artificial eutrophication

What is Eutrophication? 6 mg/L Water Less than Xochimilco 3 mg/L Movement Dissolved of water Oxygen Temperature Salinity Factors Pressure Photosynthesis Respiration Eutrophication

Eutrophication Increased Photosynthesis Nutrient Fertilizers Nitrogen and Indicators Chlorophyl Phosphorus Blocking of light to littoral zone Parameter Oligotrophic Eutrophic Average total phosphorus 8.0 84.4 Microbial Average total nitrogen 661 1875 decomposition Average Chlorophyll a 1.7 14.3 Depletion of Chlorophyll a. peak concentration 4.2 42.6 dissolved oxygen Hypoxia Less than 2.8mg/L

Objective  Detection of anormal ferric and anoxia/hipoxia levels on water samples from the Xochimilco Lake by using biosensors 2 ppm>[O 2 ] 0.5ppm>[Fe] O 2 Prom Fe Prom 0.5ppm<[Fe] 2 ppm<[O 2 ]

Topic  Biosensors: • Fast response in time • Fast and continuous measurement • High specificity (shape-specific recognition) • Simplicity in its use • Capability of measuring concentrations ranging from 10 -18 to 10 -19 M, so we need low sample requirements • Real time measurements

Justification • Acceptable levels of disolved iron in water: 0.5 ppm • Iron promoter minimal iron dissolution concentration: 1 ppm • Oxygen Promoter Works in d O 2 concentrations over 0.2 ppm • d O 2 levels in Xochimilco vary around 3 ppm • Minimal requirement of d O 2 for wild-life sustainance 6 ppm • Detection of quantitative values of GFP and RFP by Flow Cytometry

Our Biosensor

Methodology & Results A B + Bacteria Transformation Heat 20 hours Shock 37° C A B A B Transformed Bacteria Concentrated DNA A A B B

Methodology & Results A B Check +enzymes Concentrated DNA +Enzymes Expected Pattern B A A Cut Bands Ligation B Agarose Gel

Methodology & Results

EcoRI Pstl Oxygen Promoter + G.F.P Iron Promoter + G.F.P Oxygen Promoter + mR.F.P.

Discussions

Conclusions Construction of biosensors was not completed due to what appear to be flawed ligations and/or low efficiency competent cells.

Future Work  Completion of the biosensors  Proof of concept growing bacteria in absence and presence of stimuli  Quantitative measurement of disolved O ₂ and Fe in various concentrations for construction of a photoluminescent scale.  Application of biosensors to a range of samples from specific points distributed all along Xochimilco lake.

Posada Tec 2013

Travesuras 2014

Synthetic Biology Conference

Thanks  ITESM CCM  Hidrosina  GoNext Project  Tecnologico de Monterrey campus Mty.  Nautilus  Tolerantum  QFB. Isabel Coria

Special Thanks  M.Sc. Lamberto Álvarez-Palacios  Luis Vaca Dominguez M.D. Ph. D.  Alicia Sampieri García Ph. D.