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Portable Infrared System for Amplifying DNA and Conducting Field Nucleic Acid Testing PRESENTER: DILLON MIR

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Portable Infrared System for Amplifying DNA and Conducting Field Nucleic Acid TestingPRESENTER: DILLON MIR

Hypothesis

The use of infrared light to heat PCR tubes in order to create a simplified and cost effective method of DNA amplification.

Hypothesis questions

Can we create a model of PCR tube heating using infrared lamps?

Can we create a heating device that can be bought for a low cost or easily made?

Can we make this device easy to use?

Research methods

First tests were run with water in order to create the heating model without using an excess of the somewhat expensive DNA helicase.

Tests were run starting at room temperature (~24°C). Lowered or elevated temperatures causes a difference time to heat the tube.

PCR tube heated to about 65 degrees Celsius.

The heating and cooling models of different temperatures were created in order to find a time and voltage the would not damage the DNA sample.

Allowed to cool to room temperature before another experiment was run.

The first prototype

Infrared lamps

PCR tube

Breadboard

Aluminum block

Heating Equations and derivation

 

Heating Equations and derivation

The heat rate accumulation is influenced by the heat flux of the infrared light and the cooling heat flux rate due to convection.

Heating Equations and derivation

The equation was simplified using the below constants.

Heating Equations and derivation

Integrating both sides of the equation yields

Solving for the integration constant

Initial Conditions

@t=0 T=To

Heating Equations and derivation

Cooling Equations and derivation

For periods of cooling, the infrared light is not heating the system anymore.

This reduce the equation just to the cooling rate due to convection.

Cooling Equations and derivation

The value for beta was reduced using the relation between area and volume for a cylinder.

 

Cooling Equations and derivation

The equation variables are separated and are solved by integrating both sides.

Cooling Equations and derivation

Initial Conditions

@t=0 T=Ts

 

Cooling Equations and derivation

After the integration constant is solved for, it is put back into the equation.

For elevated temperatures

The initial conditions change when working with elevated temperatures.

Initial Conditions

@t=0 T=Te

 

For elevated temperatures

The constant is then put back into the equation.

Add new term

or

For elevated temperatures

Prototype 2

The next prototype we created was a “Sun of Vergina” design.

Holds up to 6 PCR tubes for multiple tests at once.

Made of clay with silver paint for reflection.

Prototype 2

BreadboardInfrared lamps

PCR tubes2nd prototype

Future prototype

Created in Solidworks.

Once designed it can be sent to a machine shop to be created.

Easy to produce more with the same specifications.

DNA amplification procedures

DNA amplification process:

1. Combine necessary components together in a PCR tube.

2. Heat PCR tube for 30 minutes.

3. Remove liquid from PCR tube and place in glass slide.

4. turn out light (except blue side LED).

5. Observe drop (green color in drop is amplified DNA).

DNA amplification

Materials Quantity (µL)

Water 14

10X Annealing Buffer 2.5

MgSO4 1

NaCl 2

IsoAmp dNTP Solution 2

DNA template 2.5

Forward Primer 1

Reverse Primer 1

IsoAmp Enzyme Mix 2

DNA amplification

Blue light makes for easier analysis using image software.

Green color inside drop is the amplified (copied) DNA.

Higher concentrations of amplified DNA will produce a more visible shade of green.

Results

The “data” values are the temperature that was measured with respect to time at different voltages.

The “prediction” values are based on the heating model that was created from the collected data.

Conclusions

Using infrared lamps to heat PCR tubes was successful.

Heating and cooling models of a PCR tube with reasonable accuracy were created.

A multi-tube DNA amplification platform was successfully designed and tested.

Acknowledgements

Dr. Antonio Garcia

Instructor/Advisor

Professor Karmella Haynes

Provided DNA helicase samples

Angel Lastra

Equations and derivations