bio detection of anoxia in lake xochimilco using modified

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 dO2 below minimun requirements

• Artificial eutrophication

What is Eutrophication?

Dissolved Oxygen

Less than 3 mg/L

Movement of water

Temperature

Salinity

Pressure

Photosynthesis

Respiration

Eutrophication

Factors

Water

Xochimilco

6 mg/L

Eutrophication

Increased Photosynthesis

Nutrient Fertilizers

Nitrogen and Phosphorus

Blocking of light to littoral zone

Microbial decomposition

Depletion of dissolved oxygen

Hypoxia Less than 2.8mg/L

Indicators Chlorophyl

Parameter Oligotrophic Eutrophic

Average total phosphorus 8.0 84.4

Average total nitrogen 661 1875

Average Chlorophyll a 1.7 14.3

Chlorophyll a. peak concentration 4.2 42.6

Objective

Detection of anormal ferric and anoxia/hipoxia

levels on water samples from the Xochimilco Lake by

using biosensors

O2Prom

2 ppm>[O2]

2 ppm<[O2]

Fe Prom

0.5ppm>[Fe]

0.5ppm<[Fe]

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 dO2 concentrations

over 0.2 ppm

• dO2 levels in Xochimilco vary around 3 ppm

• Minimal requirement of dO2 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

20 hours

37° C

A B A B

A

B

Concentrated DNA

A

B

Transformed Bacteria

Heat Shock

Methodology & Results A B

Concentrated DNA

Check +enzymes

+Enzymes

Expected Pattern

Agarose Gel

Cut Bands A

B Ligation

A B

Methodology & Results

Oxygen Promoter + G.F.P

Iron Promoter + G.F.P

Oxygen Promoter + mR.F.P.

EcoRI Pstl

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 O₂ 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

Luis Vaca Dominguez M.D. Ph. D.

Alicia Sampieri García Ph. D.

M.Sc. Lamberto Álvarez-Palacios