Difference between revisions of "Team:JMU Wuerzburg/Description"
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| − | <h3>Test Tonic – a rapid test system for the malaria-causing parasite Plasmodium</h3> | + | <h3>Test Tonic – a rapid test system for the malaria-causing parasite <span style="font-style: italic;">Plasmodium</span></h3> |
<div> | <div> | ||
| − | Malaria is | + | Malaria is a widely spread infectious disease that causes 400.000 |
| + | deaths every year and affects more than 200 million people in total according to the WHO`s Worlds Malaria Report 2017. | ||
| + | The disease is caused by different species of the protozoan parasite Plasmodium. | ||
| + | For a successful therapy of Malaria, a fast and sensitive detection of the species affecting the patient is crucial. | ||
| + | <sup><a href="#desc_refs">1</a></sup> | ||
<br><br> | <br><br> | ||
| − | Our aim is to construct a test system that is capable of detecting the DNA of Plasmodium. We venture to create a rapid, easily usable and cheap diagnostic device for large area application. To reach this goal, we design primers and probes by analysing published sequences of the human pathogenic Plasmodium species with bioinformatic tools. We reach out to create one primer-probe pair that can detect Plasmodium in general and specific primer | + | Our aim is to construct a test system that is capable of detecting the DNA of Plasmodium. |
| + | We venture to create a rapid, easily usable and cheap diagnostic | ||
| + | device for large area application. | ||
| + | To reach this goal, we design primers and probes by analysing published sequences | ||
| + | of the human pathogenic<span style="font-style: italic;"> Plasmodium species </span>with bioinformatic tools. | ||
| + | We reach out to create one primer-probe pair that can detect<span style="font-style: italic;"> Plasmodium </span>in general | ||
| + | and specific primer-probe pairs for each species that causes malaria in humans. | ||
<br><br> | <br><br> | ||
| − | + | For the amplification of characteristic parts of the<span style="font-style: italic;"> Plasmodium </span>genome we developed two pairs of primers for each species. | |
| + | One of these indicates if a patient is affected of<span style="font-style: italic;"> Plasmodium </span>in general. | ||
| + | The other one specifically detects the species <span style="font-style: italic;">Plasmodium falciparum</span>. | ||
<br><br> | <br><br> | ||
| − | + | After identification of fitting sequences, we evaluate and optimize our primers by running qPCR assays with E. | |
| + | coli containing a plasmid with a short, synthetic, non-pathogenic sequence of the<span style="font-style: italic;"> Plasmodium </span>genome as a positive control. | ||
| + | Additionally we conduct qPCR assays with genomic DNA of cultured<span style="font-style: italic;"> Plasmodium </span>parasites. | ||
| + | We would also like to use inactivated patient-derived samples to confirm selectivity and specificity of our test which requires formal permission by the ethics Committee. | ||
<br><br> | <br><br> | ||
| − | After creating a fundamental model with a multiplex qPCR we elaborate a way to apply our detection system to Recombinase Polymerase Amplification (RPA). RPA is a promising alternative for qPCR to isothermally amplify our target sequences in a short period of time. | + | To perform the amplification directions in a user-friendly way we designed a hardware model. |
| + | The reactions steps are conducted in a simple test tube with separated chambers to isolate different reactions. | ||
| + | A step motor and an Arduino manage the movement and mixing of the reaction fluids. | ||
| + | To avoid the need of expensive thermocyclers, the amplification is conducted isothermally by RPA (Recombinase Polymerase Amplification), which is capable of amplifying very low initial concentrations of DNA within around 20 minutes<sup><a href="#desc_refs">2</a></sup>. The amplified DNA product is later detected in a lateral flow assay. | ||
| + | The presence of<span style="font-style: italic;"> Plasmodium </span>DNA and specifically <span style="font-style: italic;">Plasmodium falciparum</span> DNA is indicated through a change of color. | ||
| + | |||
| + | <br><br> | ||
| + | After creating a fundamental model with a multiplex qPCR we elaborate a way to apply our detection system to Recombinase Polymerase Amplification (RPA) <sup><a href="#desc_refs">3</a></sup>. | ||
| + | RPA is a promising alternative for qPCR to isothermally amplify our target sequences in a short period of time. | ||
| + | It makes such a test system cheap and avoids the need of an expensive thermocycler. | ||
| + | This results in many advantages for the application in travelling situations and in areas without proper infrastructure and energy supply. | ||
| + | |||
| + | <hr style="width: 50%; margin-bottom: 0;"> | ||
| + | <h5 id="collabs_refs">List of References</h5> | ||
| + | <sup>1</sup> <a href="http://www.who.int/malaria/areas/treatment/overview/en/">http://www.who.int/malaria/areas/treatment/overview/en/</a><br> | ||
| + | <sup>2</sup> <a href="https://www.ncbi.nlm.nih.gov/pubmed/27160000">https://www.ncbi.nlm.nih.gov/pubmed/27160000</a><br> | ||
| + | <sup>3</sup> <a href="https://reader.elsevier.com/reader/sd/pii/S0165993617302583?token=AE0F18E7C2136EF7A427BAE926122837FF1300E42C71ECC9B6E963D576D6DA841786904CC29F16458D3472BC66EF6B7F">https://reader.elsevier.com/reader/sd/pii/S0165993617302583?token=AE0F18E7C2136EF7A427BAE926122837FF1300E42C71ECC9B6E963D576D6DA841786904CC29F16458D3472BC66EF6B7F</a> | ||
</div> | </div> | ||
</div> | </div> | ||
</html> | </html> | ||
Revision as of 00:09, 17 October 2018
Test Tonic – a rapid test system for the malaria-causing parasite Plasmodium
Malaria is a widely spread infectious disease that causes 400.000
deaths every year and affects more than 200 million people in total according to the WHO`s Worlds Malaria Report 2017.
The disease is caused by different species of the protozoan parasite Plasmodium.
For a successful therapy of Malaria, a fast and sensitive detection of the species affecting the patient is crucial.
1
Our aim is to construct a test system that is capable of detecting the DNA of Plasmodium. We venture to create a rapid, easily usable and cheap diagnostic device for large area application. To reach this goal, we design primers and probes by analysing published sequences of the human pathogenic Plasmodium species with bioinformatic tools. We reach out to create one primer-probe pair that can detect Plasmodium in general and specific primer-probe pairs for each species that causes malaria in humans.
For the amplification of characteristic parts of the Plasmodium genome we developed two pairs of primers for each species. One of these indicates if a patient is affected of Plasmodium in general. The other one specifically detects the species Plasmodium falciparum.
After identification of fitting sequences, we evaluate and optimize our primers by running qPCR assays with E. coli containing a plasmid with a short, synthetic, non-pathogenic sequence of the Plasmodium genome as a positive control. Additionally we conduct qPCR assays with genomic DNA of cultured Plasmodium parasites. We would also like to use inactivated patient-derived samples to confirm selectivity and specificity of our test which requires formal permission by the ethics Committee.
To perform the amplification directions in a user-friendly way we designed a hardware model. The reactions steps are conducted in a simple test tube with separated chambers to isolate different reactions. A step motor and an Arduino manage the movement and mixing of the reaction fluids. To avoid the need of expensive thermocyclers, the amplification is conducted isothermally by RPA (Recombinase Polymerase Amplification), which is capable of amplifying very low initial concentrations of DNA within around 20 minutes2. The amplified DNA product is later detected in a lateral flow assay. The presence of Plasmodium DNA and specifically Plasmodium falciparum DNA is indicated through a change of color.
After creating a fundamental model with a multiplex qPCR we elaborate a way to apply our detection system to Recombinase Polymerase Amplification (RPA) 3. RPA is a promising alternative for qPCR to isothermally amplify our target sequences in a short period of time. It makes such a test system cheap and avoids the need of an expensive thermocycler. This results in many advantages for the application in travelling situations and in areas without proper infrastructure and energy supply.
2 https://www.ncbi.nlm.nih.gov/pubmed/27160000
3 https://reader.elsevier.com/reader/sd/pii/S0165993617302583?token=AE0F18E7C2136EF7A427BAE926122837FF1300E42C71ECC9B6E963D576D6DA841786904CC29F16458D3472BC66EF6B7F
Our aim is to construct a test system that is capable of detecting the DNA of Plasmodium. We venture to create a rapid, easily usable and cheap diagnostic device for large area application. To reach this goal, we design primers and probes by analysing published sequences of the human pathogenic Plasmodium species with bioinformatic tools. We reach out to create one primer-probe pair that can detect Plasmodium in general and specific primer-probe pairs for each species that causes malaria in humans.
For the amplification of characteristic parts of the Plasmodium genome we developed two pairs of primers for each species. One of these indicates if a patient is affected of Plasmodium in general. The other one specifically detects the species Plasmodium falciparum.
After identification of fitting sequences, we evaluate and optimize our primers by running qPCR assays with E. coli containing a plasmid with a short, synthetic, non-pathogenic sequence of the Plasmodium genome as a positive control. Additionally we conduct qPCR assays with genomic DNA of cultured Plasmodium parasites. We would also like to use inactivated patient-derived samples to confirm selectivity and specificity of our test which requires formal permission by the ethics Committee.
To perform the amplification directions in a user-friendly way we designed a hardware model. The reactions steps are conducted in a simple test tube with separated chambers to isolate different reactions. A step motor and an Arduino manage the movement and mixing of the reaction fluids. To avoid the need of expensive thermocyclers, the amplification is conducted isothermally by RPA (Recombinase Polymerase Amplification), which is capable of amplifying very low initial concentrations of DNA within around 20 minutes2. The amplified DNA product is later detected in a lateral flow assay. The presence of Plasmodium DNA and specifically Plasmodium falciparum DNA is indicated through a change of color.
After creating a fundamental model with a multiplex qPCR we elaborate a way to apply our detection system to Recombinase Polymerase Amplification (RPA) 3. RPA is a promising alternative for qPCR to isothermally amplify our target sequences in a short period of time. It makes such a test system cheap and avoids the need of an expensive thermocycler. This results in many advantages for the application in travelling situations and in areas without proper infrastructure and energy supply.
List of References
1 http://www.who.int/malaria/areas/treatment/overview/en/2 https://www.ncbi.nlm.nih.gov/pubmed/27160000
3 https://reader.elsevier.com/reader/sd/pii/S0165993617302583?token=AE0F18E7C2136EF7A427BAE926122837FF1300E42C71ECC9B6E963D576D6DA841786904CC29F16458D3472BC66EF6B7F