Difference between revisions of "Team:SMS Shenzhen/Experiments"

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     <h1>I Abstract</h1>
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     <h1>I Overview of Experiment Design Ideas</h1>
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     <p style="font-size: 18px"> The goal of this experiment was to measure the physical properties of dextran, sucrose and glucose. Measuring the physical properties of these three sugars is fundamental in our experiment. we assume that glucose will be stained by our stains. Then, dextran and sucrose will not decompose. We set the three sugars into different concentrations and then put them in a water bath. Finally, the absorbance of solution is finally measured by the microplate reader. </p>
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     <p style="font-size: 18px">There are three crucial aspects we can focus on to prove the plausibility of our design concept, including the preliminary characteristic determination of participant sugars, the enzyme activities of the two target enzymes in realistic conditions of oral cavities, and the optic observations about the enzyme effectiveness on real scale teeth.</p>
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<h1>II Experiment Design</h1>
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<b style="font-size: 20px">Experiment 1: Preliminary Experiments: </b>
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    <h1> II Background and Hypothesis</h1>
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<p style="font-size: 18px">We characterize the stability of reaction participant sugers in various of realistic thermal and pH conditions</p>
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    <p style="font-size: 18px"> 1. Our colorant is called DNS which will react with the reducing sugar in a 100 ℃ water bath;  </p>
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<a href="https://2018.igem.org/Team:SMS_Shenzhen/Experiments/PriliminaryExperiments">
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    <p style="font-size: 18px"> 2. Glucose is found in our literature to be a reducing sugar. We assume that DNS colorant reaction, the higher the concentration, the higher the absorbance of the solution after the reaction (OD540);  </p>
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<b style="font-size: 18px">Click here for more details</b>
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    <p style="font-size: 18px"> 3. Dextran and sucrose are not reducing sugars, DNS does not react with them, so their concentrations are not related to absorbance.  </p>
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<h1> </h1>
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<b style="font-size: 20px">Experiment 2: Enzyme Activity Measurements</b>
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<p style="font-size: 18px">We measure the activity of two target enzymes - DexA and FruA</p>
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<a href="https://2018.igem.org/Team:SMS_Shenzhen/Experiments/EnzymeActivity">
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<b style="font-size: 18px">Click here for more details</b>
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<h1> </h1>
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<b style="font-size: 20px">Experiment 3: Optic Observation of Enzyme Effectiveness on Teeth sample</b>
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<p style="font-size: 18px">We demonstrated how our project will function on real scale teeth by using colour developing agent of dental plaque and image analysis</p>
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<a href="https://2018.igem.org/Team:SMS_Shenzhen/Experiments/OpticObservation">
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    <h1> III Purpose</h1>
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<b style="font-size: 18px">Click here for more details</b>
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    <p style="font-size: 18px"> 1. Construct the standard curve between the concentration of glucose and its absorbance.  </p>
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</a>
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    <p style="font-size: 18px"> 2. Considering sucrose and dextran, in the polymer state, are not the reducing sugar and do not react with DNS. But their monsters will react with them. So we measured two other factors that could cause their decomposition, pH and temperature. Taking into account the effects of the external environment on Dextran and sucrose in the real reaction, we control the variables separately and design experiments on the pH and hydration of dextran under thermal conditions.  </p>
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<h1>III Modeling</h1>
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    <h1>IV Experiment Protocol</h1>
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<b style="font-size: 20px">General references</b>
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    <h1> </h1>
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<p style="font-size: 18px">Here will be the references</p>
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    <h2>Experiment: glucose standard curve</h2>
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    <p style="font-size: 18px"> 1. Configure 1 mg/ml standard glucose solution, converse it to 250 mg/250 ml (adding 0.25 g glucose with distilled water to the constant volume of the 250ml volumetric flask)  </p>
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    <p style="font-size: 18px"> 2. Use the solution in the first step as stock liquor, take 0, 0.2, 0.4, 0.6, 0.8, 1.0 ml from it into the 15 ml test tube adding distilled water to 1ml.  </p>
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    <p style="font-size: 18px"> 3. Add 2ml DNS into each tube.  </p>
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    <p style="font-size: 18px"> 4. After adequate mixing, put each tube in 100 °C water bath for two minutes.  </p>
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    <p style="font-size: 18px">  5. After cooling, add distilled water to the solution to 15ml. </p>
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    <p style="font-size: 18px"> 6. Take 100ul from each tube into the labelling board, note down the positions of solutions in different concentrations.  </p>
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    <p style="font-size: 18px"> 7. Use microplate reader to measure solutions’ absorbance at 540 nm wave length and use distilled water as the control group.  </p>
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    <p style="font-size: 18px"> 8. The resulted data were photographed and kept as excel in USB flash disk.  </p>
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    <h1> </h1>
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    <h1> </h1>
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    <h1> </h1>
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    <h2>Experiment: preparation of the measurement of dextran thermal stability and acid stability</h2>
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    <h3>· Acid stability preparation</h3>
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    <p style="font-size: 18px">1. Take 1g dextran and dissolve it into 12.5ml Tris-HCl buffer and 12.5ml DD water.  </p>
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    <p style="font-size: 18px">2. Divide the solution into three parts and mark clearly (acid/ alkaline/ neutral +dex).</p>
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    <p style="font-size: 18px">  3. A small beaker can be used to add three drops of concentrated sulphuric acid into the portion marked "acid" and two grains sodium hydroxide into the portion marked “alkali”. Mix each portion uniformly and leave the “neutral” portion unchanged.  </p>
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    <p style="font-size: 18px"> 4. Take 100ul from each portion into a 2 ml tube and mark it (acid/ alkaline/ neutral+dex). Let them stand on the finger tube frame.  </p>
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    <h2>Experiment: preparation for measurement of sucrose thermal stability and acid stability
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    <h3>· Acid stability preparation</h3>
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    <p style="font-size: 18px">  </p>
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Revision as of 23:29, 17 October 2018

Title

Title

I Abstract

The goal of this experiment was to measure the physical properties of dextran, sucrose and glucose. Measuring the physical properties of these three sugars is fundamental in our experiment. we assume that glucose will be stained by our stains. Then, dextran and sucrose will not decompose. We set the three sugars into different concentrations and then put them in a water bath. Finally, the absorbance of solution is finally measured by the microplate reader.

II Background and Hypothesis

1. Our colorant is called DNS which will react with the reducing sugar in a 100 ℃ water bath;

2. Glucose is found in our literature to be a reducing sugar. We assume that DNS colorant reaction, the higher the concentration, the higher the absorbance of the solution after the reaction (OD540);

3. Dextran and sucrose are not reducing sugars, DNS does not react with them, so their concentrations are not related to absorbance.

III Purpose

1. Construct the standard curve between the concentration of glucose and its absorbance.

2. Considering sucrose and dextran, in the polymer state, are not the reducing sugar and do not react with DNS. But their monsters will react with them. So we measured two other factors that could cause their decomposition, pH and temperature. Taking into account the effects of the external environment on Dextran and sucrose in the real reaction, we control the variables separately and design experiments on the pH and hydration of dextran under thermal conditions.

IV Experiment Protocol

Experiment: glucose standard curve

1. Configure 1 mg/ml standard glucose solution, converse it to 250 mg/250 ml (adding 0.25 g glucose with distilled water to the constant volume of the 250ml volumetric flask)

2. Use the solution in the first step as stock liquor, take 0, 0.2, 0.4, 0.6, 0.8, 1.0 ml from it into the 15 ml test tube adding distilled water to 1ml.

3. Add 2ml DNS into each tube.

4. After adequate mixing, put each tube in 100 °C water bath for two minutes.

5. After cooling, add distilled water to the solution to 15ml.

6. Take 100ul from each tube into the labelling board, note down the positions of solutions in different concentrations.

7. Use microplate reader to measure solutions’ absorbance at 540 nm wave length and use distilled water as the control group.

8. The resulted data were photographed and kept as excel in USB flash disk.

Experiment: preparation of the measurement of dextran thermal stability and acid stability

· Acid stability preparation

1. Take 1g dextran and dissolve it into 12.5ml Tris-HCl buffer and 12.5ml DD water.

2. Divide the solution into three parts and mark clearly (acid/ alkaline/ neutral +dex).

3. A small beaker can be used to add three drops of concentrated sulphuric acid into the portion marked "acid" and two grains sodium hydroxide into the portion marked “alkali”. Mix each portion uniformly and leave the “neutral” portion unchanged.

4. Take 100ul from each portion into a 2 ml tube and mark it (acid/ alkaline/ neutral+dex). Let them stand on the finger tube frame.

Experiment: preparation for measurement of sucrose thermal stability and acid stability

· Acid stability preparation