Revision as of 22:45, 31 October 2017

Experiments

Project Management

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The Idea

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The test will be based on a newly developed method called toehold switch sensors. This method has already been successfully applied to the diagnosis of the Zika virus [3]. We are therefore convinced that toehold switch sensors will also be useful in detecting T. solium. In order to develop our test, we first need to achieve two things: The test will be based on a newly developed method called toehold switch sensors. This method has already been successfully applied to the diagnosis of the Zika virus [3]. We are therefore convinced that toehold switch sensors will also be useful in detecting T. solium. In order to develop our test, we first need to achieve two things:

Summary: Lorem ipsum dolor sit amet, consetetur sadipscing elitr, sed diam nonumy eirmod tempor invidunt ut labore et dolore magna aliquyam erat, sed diam voluptua. At vero eos et accusam et justo duo dolores et ea rebum.

The Experiments

Production of Toehold Switch Sensors

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Toehold Switch Sensor Test

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Application

Überschrift

The synthetic biology experiments described here are particularly important for participation in the iGEM competition. Simultaneously, a second team will work on the recovery of T. solium-RNA from patient stool samples in order to make our test applicable for medical purposes. We are currently forming the necessary partnerships for this goal: A working group in Nairobi has already committed to providing us with relevant material. However, these experiments will be conducted in a properly equipped laboratory, independent from the previously described experiments.

Experiments: Overview

Transcriptome analysis of T. solium
1. RNA Isolation
2. RNA sequencing
Bioinformatics
Switch Synthesis
1. Extension PCR
2. Nested PCR
3. Colony PCR
4. Gel Electrophoresis
5. DNA Clean-Up
Toehold Switch Sensor Test – Testing Pipeline
1. High troughput screening in liquid medium
2. Reliable cell free expression on cellulose membranes
Optimizing the Screening Pipeline
Other Experiments
1. 1. Preparation of nucleic acids (DNA preparation, Restriction digest/ DNA ligation, in vitro transcription, RNA purification)
2. 2. Preparation of sensors for iGEM Submission (Mutagenesis & Cloning)

Please find detailed protocols under the "Protocol" Section

The bioinformatics behind our switch design

In silico design

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The switch synthesis

Extension PCR

Toehold switches were constructed by two PCR-based steps, in which the recognition and hairpin region was attached to a LacZ reporter element. The construction was amplified from a circular DNA Template.

Primer P1 and P2 are specifically designed for different Toehold switches.( Abb. 1) These include the Recognition-Structure, the characteristic Hairpin, Ribosome binding site and Linker. The Switches were constructed by ligating them to the lacZ reporter gene.

A specific third step was used to purify the preceding amplifications and also attach a T7 Promotor region.

Nested PCR

Nested PCR is a method for speeding up the process of switch generation by combining the two steps of our extension-PCR in one cycling step. In order to extent the whole switch length without interruption, purification of P1-product and second PCR Set-Up. After consumption of P1-forward primer the already added P2 Primer takes over the reaction.

Our experiments showed the suitability of nested PCR for the creation of diagnost-x toehold switches even though different to the situation described by Schuelke (1) our forward primers have equal overlaps and therefore are not suitable for modulating the annealing temperature in order to start the primers separately. Reactions with different relation in the amounts of P1 and P2 forward primer proved the relation of P1/P2=1/4 as the most reliable in sequence stability.

Colony PCR

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

DNA Clean Up (via column & gel extraction)

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Gel electrophoresis

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The Toehold Switch Sensor Test

Standard screening in liquid phase

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

CFE on membrane trials (BSA vs Whatman membrane)

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Taenia solium model

RNA Library

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Optimizing of our screening pipeline

Primer efficiency analysis & PCR characterization (P2 vs P3)

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Optimizing CFE

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Other Experiments

TA Cloning

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Fusion Cloning

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Mutagenesis

Überschrift

The synthetic biology experiments described here are particularly important for participation in the iGEM competition. Simultaneously, a second team will work on the recovery of T. solium-RNA from patient stool samples in order to make our test applicable for medical purposes. We are currently forming the necessary partnerships for this goal: A working group in Nairobi has already committed to providing us with relevant material. However, these experiments will be conducted in a properly equipped laboratory, independent from the previously described experiments.

The Idea

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The test will be based on a newly developed method called toehold switch sensors. This method has already been successfully applied to the diagnosis of the Zika virus [3]. We are therefore convinced that toehold switch sensors will also be useful in detecting T. solium. In order to develop our test, we first need to achieve two things: The test will be based on a newly developed method called toehold switch sensors. This method has already been successfully applied to the diagnosis of the Zika virus [3]. We are therefore convinced that toehold switch sensors will also be useful in detecting T. solium. In order to develop our test, we first need to achieve two things:

Summary: Lorem ipsum dolor sit amet, consetetur sadipscing elitr, sed diam nonumy eirmod tempor invidunt ut labore et dolore magna aliquyam erat, sed diam voluptua. At vero eos et accusam et justo duo dolores et ea rebum.

The Experiments

Production of Toehold Switch Sensors

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Toehold Switch Sensor Test

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Application

Überschrift

The synthetic biology experiments described here are particularly important for participation in the iGEM competition. Simultaneously, a second team will work on the recovery of T. solium-RNA from patient stool samples in order to make our test applicable for medical purposes. We are currently forming the necessary partnerships for this goal: A working group in Nairobi has already committed to providing us with relevant material. However, these experiments will be conducted in a properly equipped laboratory, independent from the previously described experiments.

Protocols

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The Idea

The Tapeworm Taenia solium

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

The test will be based on a newly developed method called toehold switch sensors. This method has already been successfully applied to the diagnosis of the Zika virus [3]. We are therefore convinced that toehold switch sensors will also be useful in detecting T. solium. In order to develop our test, we first need to achieve two things: The test will be based on a newly developed method called toehold switch sensors. This method has already been successfully applied to the diagnosis of the Zika virus [3]. We are therefore convinced that toehold switch sensors will also be useful in detecting T. solium. In order to develop our test, we first need to achieve two things:

Summary: Lorem ipsum dolor sit amet, consetetur sadipscing elitr, sed diam nonumy eirmod tempor invidunt ut labore et dolore magna aliquyam erat, sed diam voluptua. At vero eos et accusam et justo duo dolores et ea rebum.

The Experiments

Production of Toehold Switch Sensors

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Toehold Switch Sensor Test

Taenia solium, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point  [1]. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis.

Application

Überschrift

The synthetic biology experiments described here are particularly important for participation in the iGEM competition. Simultaneously, a second team will work on the recovery of T. solium-RNA from patient stool samples in order to make our test applicable for medical purposes. We are currently forming the necessary partnerships for this goal: A working group in Nairobi has already committed to providing us with relevant material. However, these experiments will be conducted in a properly equipped laboratory, independent from the previously described experiments.

@@ Line 29: / Line 29: @@
          <div class="col-4 text-center"><a href="#" type="button" class="btn btn-primary igem_but btn-lg" id="01">Project Management</a></div>
          <div class="col-4 text-center"><a href="#" type="button" class="btn btn-primary igem_but btn-lg" id="02">Experiments – Overview</a></div>
-         <div class="col-4 text-center"><a href="#" type="button" class="btn btn-primary igem_but btn-lg" id="03">Protocols</a></div>
+         <div class="col-4 text-left"><a href="#" type="button" class="btn btn-primary igem_but btn-lg" id="03">Protocols</a></div>
      </div>
@@ Line 133: / Line 133: @@
 <!-- Experiment Overview -->
-<div id="experi" style="display: none;">
+<div id="experi" style="display: none;"><div id="inhaltsverzeichnis">
-<!-- Experiments -->
+<div class="row">
-    <!-- Schema 1 -->
+    <div class="col-12">
-    <div class="row">
+        <h3 class="text-center igem_blue mt-4"><strong>Experiments: Overview</strong></h3>
-        <div class="col-12">
-            <h3 class="text-center igem_blue mt-4"><strong>The preliminary work</strong></h3>
-        </div>
      </div>
-    <div class="row mt-3">
+</div>
-        <div class="col-6">
+<div class="row">
-            <h5 class="text-left igem_blue pb-1">RNA isolation </h5>
+    <div class="col-6">
-             <p class="text-justify"><em>Taenia solium</em>, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point <a class="igem_blue" href="#"> [1]</a>. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis. </p>
+    <ul>
-         </div>
+        <li><a class="igem_blue" href="#transkriptom">Transcriptome analysis of T. solium</a>
-        <div class="col-6">
+            <ol>
-             <img class="img-fluid" src="https://placekitten.com/550/400">
+            <li>RNA Isolation</li>
-        </div>
+             <li>RNA sequencing</li>
-    </div>
+        </ol></li>
-    <div class="row mt-5">
+        <li><a class="igem_blue" href="#bioinformatics">Bioinformatics</a></li>
-         <div class="col-6">
+         <li><a class="igem_blue" href="#switch_synthese">Switch Synthesis</a>
-            <img class="img-fluid" src="https://placekitten.com/550/400">
+             <ol>
-         </div>
+            <li>Extension PCR</li>
-         <div class="col-6">
+            <li>Nested PCR</li>
-            <h5 class="text-left igem_blue">RNA sequencing </h5>
+            <li>Colony PCR</li>
-            <p class="text-justify"><em>Taenia solium</em>, otherwise known as pork tapeworm, is a parasite with potentially serious health effects in humans. It infects millions of people worldwide, leading to severe brain diseases, blindness, epilepsy, and death. Countries where where pork production and consumption are coupled with poor hygiene are the worst affected, including large areas of Latin America, Sub-Saharan Africa, and South- and East-Asia, where over 14% of people have experienced an infection at some point <a class="igem_blue" href="#"> [1]</a>. However, a clear picture of how many people are currently infected by T. solium does not exist, due to the current difficulty in making a diagnosis. </p>
+            <li>Gel Electrophoresis</li>
-        </div>
+            <li>DNA Clean-Up</li>
+         </ol></li>
+        <li><a class="igem_blue" href="#switch_testing">Toehold Switch Sensor Test – Testing Pipeline</a>
+            <ol>
+            <li>High troughput screening in liquid medium</li>
+            <li>Reliable cell free expression on cellulose membranes</li>
+         </ol></li>
+         <li><a class="igem_blue" href="#screening_opti">Optimizing the Screening Pipeline</a></li>
+        <li><a class="igem_blue" href="#andere_experimente">Other Experiments</a>
+            <ol>
+            <li>1. Preparation of nucleic acids (DNA preparation, Restriction digest/ DNA ligation, in vitro transcription, RNA purification)</li>
+            <li>2. Preparation of sensors for iGEM Submission (Mutagenesis & Cloning)</li>
+        </ol></li>
+        <p class="text-center"><strong>Please find detailed protocols under the "Protocol" Section</strong></p>
+    </ul>
      </div>
+    <div class="col-6"></div>
+</div>
+</div>
      <!-- Seperator -->

Difference between revisions of "Team:Berlin diagnostX/Experiments"

Revision as of 22:45, 31 October 2017

Experiments

Project Management

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Idea

The Tapeworm Taenia solium

The Experiments

Production of Toehold Switch Sensors

Toehold Switch Sensor Test

Application

Überschrift

Experiments: Overview

The bioinformatics behind our switch design

In silico design

The switch synthesis

Extension PCR

Nested PCR

Colony PCR

DNA Clean Up (via column & gel extraction)

Gel electrophoresis

The Toehold Switch Sensor Test

Standard screening in liquid phase

CFE on membrane trials (BSA vs Whatman membrane)

Taenia solium model

RNA Library

Optimizing of our screening pipeline

Primer efficiency analysis & PCR characterization (P2 vs P3)

Optimizing CFE

Other Experiments

TA Cloning

Fusion Cloning

Mutagenesis

Überschrift

The Idea

The Tapeworm Taenia solium

The Experiments

Production of Toehold Switch Sensors

Toehold Switch Sensor Test

Application

Überschrift

Protocols

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Idea

The Tapeworm Taenia solium

The Experiments

Production of Toehold Switch Sensors

Toehold Switch Sensor Test

Application

Überschrift

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Tapeworm Taenia solium

The Tapeworm Taenia solium