Difference between revisions of "Team:NCTU Formosa/Composite Part"

 
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           <div class="node">
 
           <div class="node">
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           <div class="topic"><p class="text_color">Omega-hexatoxin-Hv1a</p></div>
+
           <div class="topic"><p class="text_color">Composite Parts</p></div>
 
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           <div class="topic"><p class="text_color">U2-segestritoxin-Sf1a</p></div>
+
           <div class="topic"><p class="text_color">PANTIDE without Lectin</p></div>
 
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           <div class="topic"><p class="text_color">Orally Active Insecticidal Peptide</p></div>
+
           <div class="topic"><p class="text_color">PANTIDE with Lectin</p></div>
 
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           <div class="topic"><p class="text_color">Omega-hexatoxin-Hv1a(lectin)</p></div>
+
           <div class="topic"><p class="text_color">PANTIDE with GS Linker</p></div>
 
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<!--圖片-->
 
<!--圖片-->
 
   <div class="img-container">
 
   <div class="img-container">
−
         <img src="https://static.igem.org/mediawiki/2016/7/79/Parts.png" class="main-img" width="100%">
+
         <img src="https://static.igem.org/mediawiki/2016/9/98/NCTU_COMPOSITE_BIG_PICTURE.png
 +
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 +
<!--------------------wiki content----------------------->
 +
<section class="content_container">
 +
<div>
 +
<p class="title">Composite Parts</p>
 +
<p class="content">We have known the function of the three toxic peptides of PANTIDE, Omega-hexatoxin-Hv1a, µ-segestritoxin-Sf1a, and Orally Active Insecticidal Peptide(OAIP) respectively and they are easy to express with the help of the bacteria <i>E. coli</i> BL21 Rosetta Gami strain. So, we then start to create BioBricks of these three toxins which we have obtained the nucleotide sequences. We use a strong T7 promoter that is suitable with the strain of <i>E. coli</i> we chose and also powerful at expressing proteins with multiple disulfide bonds, and we add His-tag for protein purification.</p>
  
−
<!--wiki content-->
+
<p class="title">PANTIDE without Lectin</p>
 +
<p class="content"> PANTIDE can be a single toxic peptide of the three toxins we chose, Hv1a, Sf1a, and OAIP respectively, which derive from the natural predators spiders. </p>
  
 +
<!----------Hv1a-------->
 +
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974011" class="biobrick"><p class="content-1">BBa_K1974011</p></a>
  
−
<section class="content_container">
+
<div>
−
  <div>
+
      <img src="https://static.igem.org/mediawiki/2016/f/f7/NCTU_FORMOSA_H1.png" class="picture">
−
    <p class="title">Omega-hexatoxin-Hv1a<p>
+
</div>
−
    <p class="content">T7 Promoter+B0034+Hv1a+linker+6xHistag</p>
+
−
    <p class="content-1">Introduction: </p>
+
−
        <div>
+
−
            <img src="a">
+
−
        </div>
+
−
    <p class="content">By ligating the IPTG induced promoter T7 (BBa_ I712074), strong ribosome binding site (BBa_B0034), hv1a, linker, and the 6xHistag (BBa_ K1223006), we can express Hv1a, the toxin by IPTG induction.</p>
+
−
    <p class="content">This year we create a revolutionary system that integrates biological pesticides, an automatic detector, a sprinkler, and IoT. We made a database that contains most of the spider toxins and selected the target toxins by programming. Omega-hexatoxin-Hv1a is coded for the venom of a spider, Hadronyche versuta. It is under the control of the strong T7 promoter. A 6xHistag is added for further protein purification.</p>
+
−
    <p class="content-1">Mechanism of Hv1a</p>
+
−
    <p class="content">According to the reference, Omega-hexatoxin-Hv1a has a structure called ICK(inhibitor cysteine knot).[1] This kind of structure contains three disulfide bonds and beta-sheet. With this structure, Hv1a can resist the high temperature, acid base solution and the digest juice of insect gut. Hv1a can bind on insect voltage-gated Calcium channels (CaV1) in the central nervous system, making it paralyze and die eventually.</p>
+
  
−
        <div>
+
<p class="content">
−
            <img src="b">
+
T7 promoter+RBS+Hv1a+linker+His-tag+terminator
−
            <img src="c">
+
<br>PANTIDE that contains the gene coding for Hv1a linked with His-tag.
−
            <img src="d">
+
</p>
−
        </div>
+
  
−
    <p class="content-1">Features of Hv1a</p>
+
<!---------Sf1a----------->
−
    <p class="content">1. Non-toxic: Omega-hexatoxin-Hv1a is non-toxic to mammals and Hymenoptera (bees). Since the structure of the target ion channel is different, omega-hexatoxin-Hv1a does not harm mammals and bees. So it is safe to use it as a biological pesticide.</p>
+
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974012" class="biobrick"><p class="content-1">BBa_K1974012</p></a>
−
    <p class="content">2. Biodegradable: Omega-hexatoxin-Hv1a is a polypeptide so it must degrade over time. After degradation, the toxin will become nutrition in the soil.</p>
+
−
    <p class="content">3. Species-specific: According to reference, Omega-hexatoxin-Hv1a has specificity to Lepidopteran (moths), Dipteran (flies) and Orthopteran (grasshoppers).</p>
+
−
    <p class="content">4. Eco-friendly: Compare with chemical pesticides, Omega-hexatoxin-Hv1a will not remain in soil and water so that it will not pollute the environment and won’t harm the ecosystem. </p>
+
−
    <p class="content">Altogether, using Hv1a is totally an environmentally friendly way for solving harmful insect problems by using this ion channel inhibitor as a biological pesticide.</p>
+
  
−
    <p class="content-1">Target insect:</p>
+
<div>
−
    <li class="list">House cricket (<i>Acheta domesticus</i>) </li>
+
      <img src="https://static.igem.org/mediawiki/2016/a/ad/NCTU_FORMOSA_S1.png" class="picture">
−
    <li class="list"><i>Musca domestica</i></li>
+
</div>
−
    <li class="list"><i>Amblyomma americanum</i></li>
+
−
    <li class="list"><i>Heliothis virescens</i></li>
+
  
−
    <p class="content-1">Experiment</p>
+
<p class="content">
−
    <p class="content-2">1. Cloning </p>
+
T7 promoter+RBS+Sf1a+linker+His-tag+terminator
−
    <p class="content">After assembling the DNA sequences from the basic parts, we recombined each T7 Promoter+B0034+toxin +linker+6xHistag  gene to pSB1C3 backbones and conducted a PCR experiment to check the size of each part. The DNA sequence length of these parts is around 250-500 bp. In this PCR experiment, the toxin product's size should be near at 450-700 bp. </p>
+
<br>PANTIDE that contains the gene coding for Sf1a linked with His-tag.
−
        <div>
+
</p>
−
            <img src="e">
+
−
        </div>
+
−
    <p class="content">proved that we successfully ligated the toxin sequence onto an ideal backbone.</p>
+
−
    <p class="content-2">2. Expressing</p>
+
−
    <p class="content"><i>E.coli</i>(DE3) express the protein and form the disulfide in the cytoplasm. We sonicated the bacteria and purified the protein by 6xHis-tag behind the toxin using Nickel resin column. </p>
+
−
    <p class="content-2">3. Analysis</p>
+
−
    <p class="content">We do the Bradford analysis to get the protein concentration. </p>
+
−
        <div>
+
−
            <img src="f">
+
−
        </div>
+
−
    <p class="content">Also, we do the UV test and model the degradation rate.</p>
+
−
        <div>
+
−
            <img src="g">
+
−
        </div>
+
−
    <p class="content">We did the feeding assay on the Tabacco cutworm (Spodoptera litura) to calculate the LC<sub>50/sub>.</p>
+
−
    <p class="content">放蟲抽搐、死亡之影片</p>
+
−
    <p class="content-2">4.Modeling</p>
+
−
    <p class="content">According to reference, the energy of Ultraviolet will break the disulfide bonds and the toxicity is also decreased. To take the parameter into consideration for our automatic system, we modeled the degradation rate of the protein and modified the program in our device.</p>
+
−
        <div>
+
−
            <img src="h">放預測降解速率的圖
+
−
        </div>
+
−
    <p class="content-2">5. Device</p>
+
−
    <p class="content">We designed a device that contains a detector, a sprinkler, and an integrated hardware with users by APP through IoT talk. We use an infrared detector to detect the number of the pest and predict what time to spray the farmland. Furthermore, other detectors like temperature, humidity, lamination, pressure of carbon dioxide are also installed in our device. At the same time, the APP that displays all the information about the farmland would contact the users and spray biological pesticides automatically. This device can make farmers control the farmland remotely.</p>
+
−
        <div>
+
−
            <img src="i">放device的真實圖
+
−
        </div>
+
  
−
    </div>
+
<!-------OAIP------------->
 +
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974013" class="biobrick"><p class="content-1">BBa_K1974013</p></a>
  
−
  <div>
+
<div>
−
    <p class="title">U2-segestritoxin-Sf1a</p>
+
      <img src="https://static.igem.org/mediawiki/2016/2/21/NCTU_FORMOSA_O.png" class="picture">
−
    <p class="content">T7 Promoter+RBS+Sf1a+linker+6xhistag</p>
+
</div>
−
    <p class="content-1">Introduction:</p>
+
−
        <div>
+
−
            <img src="a">
+
−
        </div>
+
−
    <p class="content">By ligating the IPTG induced promoter T7 (BBa_ I712074), strong ribosome binding site (BBa_B0034), sf1a, linker, and the 6xHistag (BBa_ K1223006), we can express Sf1a, the toxin by IPTG induction.</p>
+
−
    <p class="content">This year we create a revolutionary system that integrates biological pesticides, an automatic detector, a sprinkler, and IoT. We made a database that contains most of the spider toxins and selected the target toxins by programming. U2-segestritoxin-Sf1a is coded for the venom of a spider, <i>Segestria florentina</i>. It is under the control of the strong T7 promoter. A 6xHistag is added for further protein purification.</p>
+
−
    <p class="content-1">Mechanism of Sf1a</p>
+
−
    <p class="content">U2-segestritoxin-Sf1a has a structure called ICK(inhibitor cysteine knot). This kind of structure contains four disulfide bonds. With this structure, Sf1a can resist the high temperature, acid-base solution and the digest juice of insect gut. Sf1a can bind on insect voltage-gated sodium channel Site-1, making it paralyze and die eventually.</p>
+
−
        <div>
+
−
            <img src="b">
+
−
        </div>
+
−
    <p class="content-1">Features of Sf1a</p>
+
−
    <p class="content">1.Non-toxic: U2-segestritoxin-Sf1a is non-toxic to mammals and bees. Since the structure of the target ion channel is different, U2-segestritoxin-Sf1a does not harm mammals and bees. So it is safe to use it as a biological pesticide.</p>
+
−
    <p class="content">2.Biodegradable: The toxin is a peptide, so it must degrade over time. After degradation, the toxin will become nutrition in the soil.</p>
+
−
    <p class="content">3.Species-specific: According to reference, U2-segestritoxin-Sf1a has specificity to Lepidopteran (moths) and Dipteran (flies). So another insect such as bees will not be killed.</p>
+
−
    <p class="content">4.Eco-friendly: Compare with a chemical pesticide, U2-segestritoxin-Sf1a will not remain in soil and water so that it will not pollute the environment and won’t harm the ecosystem. </p>
+
−
    <p class="content">Together, using Sf1a is totally an environmentally friendly way for solving harmful insect problems by using this ion channel inhibitor as a biological pesticide.</p>
+
−
    <p class="content-1">Target insects</p>
+
−
    <li class="list"><i>Lucilia cuprina</i></li>
+
−
    <li class="list"><i>Heliothis virescens</i></li>
+
−
    <p class="content-1">Experiment</p>
+
−
    <p class="content-2">1. Cloning</p>
+
−
    <p class="content">After assembling the DNA sequences from the basic parts, we recombined each T7 Promoter+B0034+toxin +linker+6xHistag  gene to pSB1C3 backbones and conducted a PCR experiment to check the size of each part. The DNA sequence length of these parts is around 250-500 bp. In this PCR experiment, the toxin product's size should be near at 450-700 bp. </p>
+
−
        <div>
+
−
            <img src="c">放PCR電泳跑的圖(證明大小正確)
+
−
        </div>
+
−
    <p class="content">proved that we successfully ligated the toxin sequence onto an ideal backbone.</p>
+
−
    <p class="content-2">2. Expressing</p>
+
−
    <p class="content"><i>E.coli</i>(DE3) expressed the protein and form the disulfide in the cytoplasm. We sonicated the bacteria and purify the protein by 6xHis-tag behind the toxin using Nickel resin column. </p>
+
−
    <p class="content-2">3. Analysis</p>
+
−
    <p class="content">We do the Bradford analysis to get the protein concentration. </p>
+
−
        <div>
+
−
            <img src="d">放SDS-PAGE圖，證明我們有表現出來
+
−
        </div>
+
−
    <p class="content">Also, we do the UV test and model the degradation rate.</p>
+
−
        <div>
+
−
            <img src="d">放濃度對時間作圖
+
−
        </div>
+
−
    <p class="content">We did the feed assay on the Tabacco cutworm (<i>Spodoptera litura</i>) to calculate the LC<sub>50</sub>.</p>
+
−
    <p class="content">放蟲抽搐、死亡之影片</p>
+
−
    <p class="content-2">4.Modeling</p>
+
−
    <p class="content">According to reference, the energy of Ultraviolet will break the disulfide bonds and the toxicity is also decreased. To take the parameter into consideration for our automatic system, we modeled the degradation rate of the protein and modified the program in our device.</p>
+
−
        <div>
+
−
            <img src="f">放預測降解速率的圖
+
−
        </div>
+
−
    <p class="content-2">5. Device</p>
+
−
    <p class="content">We designed a device that contains detector, sprinkler, and integrated hardware with users by APP through IoT talk. We use an infrared detector to detect the number of the pest and predict what time to spray the farmland. Furthermore, other detectors like temperature, humidity, lamination, pressure of carbon dioxide and on also install in our device. At the same time, the APP would contact the users that all the information about the farmland and spray biological pesticides automatically. This device can make farmers control the farmland remotely.</p>
+
−
        <div>
+
−
            <img src="f">放device的真實圖
+
−
        </div>
+
  
 +
<p class="content">
 +
T7 promoter+RBS+OAIP+linker+His-tag+terminator
 +
<br>PANTIDE that contains the gene coding for OAIP linked with His-tag.
 +
</p>
  
−
  </div>
 
  
−
  <div>
 
−
    <p class="title">Orally Active Insecticidal Peptide</p>
 
−
    <p class="content">T7 Promoter+RBS+Sf1a+linker+6xhistag</p>
 
−
    <p class="content-1">Introduction:</p>
 
−
        <div>
 
−
            <img src="a">
 
−
        </div>
 
−
    <p class="content">By ligating the IPTG induced promoter T7 (BBa_ I712074), strong ribosome binding site (BBa_B0034), OAIP, linker, and the 6xHistag (BBa_ K1223006), we are able to express OAIP, the toxin by IPTG induction.</p>
 
−
    <p class="content">This year we create a revolutionary system that integrates biological pesticides, automatic detector, sprinkler, and IoT. We made a database that contains most of the spider toxins and selected the target toxins by programming. Orally Active Insecticidal Peptide is coded for the venom of a spider, <i>Selenotypus plumipes</i>. It is under the control of the strong T7 promoter. A 6xHistag is added for further protein purification.</p>
 
−
    <p class="content-1">Mechanism of OAIP</p>
 
−
    <p class="content">Orally Active Insecticidal Peptide has a structure called ICK (inhibitor cysteine knot). This kind of structure contains three disulfide bonds. With this structure OAIP can resist the high temperature, acid base solution and the digest juice of insect gut. OAIP can bind on the voltage-gated sodium channel in the insect’s nervous system, making it paralyze and die eventually.</p>
 
−
        <div>
 
−
            <img src="b">
 
−
        </div>
 
−
    <p class="content-1">Features of OAIP</p>
 
−
    <p class="content">1.Non-toxic: Orally Active Insecticidal Peptide is non-toxic to mammals and bees. Since the structure of the target ion channel is different, Orally Active Insecticidal Peptide does not harm mammals and bees. So it is safe to use it as a biological pesticide.</p>
 
−
    <p class="content">2.Biodegradable: The toxin is a peptide, so it must degrade over time. After degradation, the toxin will become nutrition inside the soil.</p>
 
−
    <p class="content">3.Species-specific: According to reference, Orally Active Insecticidal Peptide has specificity to Lepidopteran (moths), Coleopteran (beetles) and Isopteran (termite). So another insect such as bees will not be killed.</p>
 
−
    <p class="content">4.Eco-friendly: Compare with a chemical pesticide, Orally Active Insecticidal Peptide will not remain in soil and water so that it will not pollute the environment and won’t harm the ecosystem. </p>
 
−
    <p class="content">Together, using OAIP is totally an environmentally friendly way for solving harmful insect problems by using this ion channel inhibitor as a biological pesticide.</p>
 
  
−
    <p class="content-1">Experiment</p>
+
<!---------------lectin---------------->
−
    <p class="content-2">1. Cloning</p>
+
−
    <p class="content">After assembling the DNA sequences from the basic parts, we recombined each T7 Promoter+B0034+toxin +linker+6xHistag  gene to pSB1C3 backbones and conducted a PCR experiment to check the size of each part. The DNA sequence length of these parts is around 250-500 bp. In this PCR experiment, the toxin product's size should be near at 450-700 bp. </p>
+
−
        <div>
+
−
            <img src="c">放PCR電泳跑的圖(證明大小正確)
+
−
        </div>
+
−
    <p class="content">proved that we successfully ligated the toxin sequence onto an ideal backbone.</p>
+
−
    <p class="content-2">2. Expressing</p>
+
−
    <p class="content"><i>E.coli</i>(DE3) express the protein and form the disulfide in the cytoplasm. We sonicated the bacteria and purified the protein by 6xHis-tag behind the toxin using Nickel resin column. </p>
+
−
    <p class="content-2">3. Analysis</p>
+
−
    <p class="content">We do the Bradford analysis to get the protein concentration. </p>
+
−
        <div>
+
−
            <img src="d">放SDS-PAGE圖，證明我們有表現出來
+
−
        </div>
+
−
    <p class="content">Also, we do the UV test and model the degradation rate.</p>
+
−
        <div>
+
−
            <img src="d">放濃度對時間作圖
+
−
        </div>
+
−
    <p class="content">We did the feed assay on the Tabacco cutworm (<i>Spodoptera litura</i>) to calculate the LC<sub>50</sub>.</p>
+
−
    <p class="content">放蟲抽搐、死亡之影片</p>
+
−
    <p class="content-2">4.Modeling</p>
+
−
    <p class="content">According to reference, the energy of Ultraviolet will break the disulfide bonds and the toxicity is also decreased. To take the parameter into consideration for our automatic system, we modeled the degradation rate of the protein and modified the program in our device.</p>
+
−
        <div>
+
−
            <img src="f">放預測降解速率的圖
+
−
        </div>
+
−
    <p class="content-2">5. Device</p>
+
−
    <p class="content">We designed a device that contains detector, sprinkler, and integrated hardware with users by APP through IoT talk. We use an infrared detector to detect the number of the pest and predict what time to spray the farmland. Furthermore, other detectors like temperature, humidity, lamination, pressure of carbon dioxide and on also install in our device. At the same time, the APP would contact the users that all the information about the farmland and spray biological pesticides automatically. This device can make farmers control the farmland remotely.</p>
+
−
        <div>
+
−
            <img src="f">放device的真實圖
+
−
        </div>
+
−
  </div>
+
  
−
  <div>
 
−
    <p class="title">Omega-hexatoxin-Hv1a</p>
 
−
    <p class="content">T7promoter+RBS+Hv1a+linker+snoedrop-lectin+linker+6xhistag></p>
 
−
    <p class="content-1">Introduction:</p>
 
−
        <div>
 
−
            <img src="a">
 
−
        </div>
 
−
    <p class="content">By ligating the IPTG induced promoter T7 (BBa_ I712074), strong ribosome binding site (BBa_B0034), Hv1a, snowdrop-lectin, and the 6xHistag (BBa_ K1223006). Moreover, we added two kinds of the linker to connect Hv1a, snowdrop-lectin. Two linkers are different size of amino acids. We can express the toxin fused with snowdrop lectin by IPTG induction.</p>
 
−
    <p class="content">This year we create a revolutionary system that integrates biological pesticides, automatic detector, sprinkler, and IoT. We made a database that contains most of the spider toxins and selected the target toxins by programming. Omega-hexatoxin-Hv1a is coded for the venom of a spider, <i>Hadronyche versuta</i>.</p>
 
−
    <p class="content">It is under the control of the strong T7 promoter. Snowdrop-lectin acts as a carrier that could transport the toxin to insect’s nervous system, hemolymph and can improve the oral activity. A 6xHistag is added for further protein purification.</p>
 
−
    <p class="content-1">Mechanism of Hv1a</p>
 
−
    <p class="content">According to reference, snowdrop-lectin is resistant to high temperature and would not be degraded by digestive juice. The species-specificity is based on the toxin, and the snowdrop lectin is the role of the carrier.</p>
 
−
    <p class="content">According to reference, Omega-hexatoxin-Hv1a has a structure called ICK(inhibitor cysteine knot).<sub>[1]</sub> This kind of structure contains three disulfide bonds and beta-sheet. With this structure, Hv1a can resist the high temperature, acid-base solution and the digest juice of insect gut. Hv1a can bind on insect voltage-gated Calcium channels (CaV1) in the central nervous system, making it paralyze and die eventually. </p>
 
  
−
    <p class="content-1">Features of Hv1a</p>
 
−
    <p class="content">1.Non-toxic: Omega-hexatoxin-Hv1a is non-toxic to mammals and Hymenoptera (bees). Since the structure of the target ion channel is different, omega-hexatoxin-Hv1a does not harm mammals and bees. So it is safe to use it as a biological pesticide.</p>
 
−
    <p class="content">2.Biodegradable: Omega-hexatoxin-Hv1a is a polypeptide so it must degrade over time. After degradation, the toxin will become nutrition in the soil.</p>
 
−
    <p class="content">3.Species-specific: According to reference, Omega-hexatoxin-Hv1a has specificity to Lepidopteran (moths), Dipteran (flies) and Orthopteran (grasshoppers).</p>
 
−
    <p class="content">4.Eco-friendly: Compare with chemical pesticides, Omega-hexatoxin-Hv1a will not remain in soil and water so that it will not pollute the environment and won’t harm the ecosystem. </p>
 
−
    <p class="content">Together, using Hv1a is totally an environmentally friendly way for solving harmful insect problems by using this ion channel inhibitor as a biological pesticide.</p>
 
−
    <p class="content-1">Target insect:</p>
 
−
    <li class="list">House cricket (<i>Acheta domesticus</i>) </li>
 
−
    <li class="list"><i>Musca domestica</i></li>
 
−
    <li class="list"><i>Amblyomma americanum</i></li>
 
−
    <li class="list"><i>Heliothis virescens</i></li>
 
  
−
    <p class="content-1">Experiment</p>
+
<p class="title">PANTIDE with Lectin</p>
−
    <p class="content-2">1. Cloning</p>
+
<p class="content"> PANTIDE can be a single toxic peptide of the three toxins we chose, Hv1a, Sf1a, and OAIP respectively with the addition of Galanthus nivalis agglutinin (GNA), for GNA has the function of acting as a carrier of PANTIDE from insect gut to the haemolymph and finally PANTIDE can transmit to the action site. </p>
−
    <p class="content">After assembling the DNA sequences from the basic parts, we recombined each T7 Promoter+B0034+toxin +linker+6xHistag  gene to pSB1C3 backbones and conducted a PCR experiment to check the size of each part. The DNA sequence length of these parts is around 250-500 bp. In this PCR experiment, the toxin product's size should be near at 450-700 bp. </p>
+
−
        <div>
+
−
            <img src="c">放PCR電泳跑的圖(證明大小正確)
+
−
        </div>
+
−
    <p class="content">proved that we successfully ligated the toxin sequence onto an ideal backbone.</p>
+
−
    <p class="content-2">2. Expressing</p>
+
−
    <p class="content"><i>E.coli</i>(DE3) express the protein and form the disulfide in the cytoplasm. We sonicated the bacteria and purified the protein by 6xHis-tag behind the toxin using Nickel resin column. </p>
+
−
    <p class="content-2">3. Analysis</p>
+
−
    <p class="content">We do the Bradford analysis to get the protein concentration. </p>
+
−
        <div>
+
−
            <img src="d">放SDS-PAGE圖，證明我們有表現出來
+
−
        </div>
+
−
    <p class="content">Also, we do the UV test and model the degradation rate.</p>
+
−
        <div>
+
−
            <img src="d">放濃度對時間作圖
+
−
        </div>
+
−
    <p class="content">We did the feed assay on the Tabacco cutworm (<i>Spodoptera litura</i>) to calculate the LC<sub>50</sub>.</p>
+
−
    <p class="content">放蟲抽搐、死亡之影片</p>
+
−
    <p class="content-2">4.Modeling</p>
+
−
    <p class="content">According to reference, the energy of Ultraviolet will break the disulfide bonds and the toxicity is also decreased. To take the parameter into consideration for our automatic system, we modeled the degradation rate of the protein and modified the program in our device.</p>
+
−
        <div>
+
−
            <img src="f">放預測降解速率的圖
+
−
        </div>
+
−
    <p class="content-2">5. Device</p>
+
−
    <p class="content">We designed a device that contains detector, sprinkler, and integrated hardware with users by APP through IoT talk. We use an infrared detector to detect the number of the pest and predict what time to spray the farmland. Furthermore, other detectors like temperature, humidity, lamination, pressure of carbon dioxide and on also install in our device. At the same time, the APP would contact the users that all the information about the farmland and spray biological pesticides automatically. This device can make farmers control the farmland remotely.</p>
+
−
        <div>
+
−
            <img src="f">放device的真實圖
+
−
        </div> 
+
−
  </div>
+
  
 +
<!----------Hv1a lectin----------->
 +
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974021" class="biobrick"><p class="content-1">BBa_K1974021</p></a>
  
 +
<div>
 +
      <img src="https://static.igem.org/mediawiki/2016/b/b6/NCTU_FORMOSA_HL.png" class="picture">
 +
</div>
 +
 +
<p class="content">
 +
T7 promoter+RBS+Hv1a+linker+lectin+linker+His-tag+terminator
 +
<br>PANTIDE that contains the gene coding for Hv1a and the snowdrop lectin linked with His-tag.
 +
 +
</p>
 +
 +
<!---------Sf1a lectin---------->
 +
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974022" class="biobrick"><p class="content-1">BBa_K1974022</p></a>
 +
 +
<div>
 +
      <img src="https://static.igem.org/mediawiki/2016/c/c3/NCTU_FORMOSA_SL.png" class="picture">
 +
</div>
 +
 +
<p class="content">
 +
T7 promoter+RBS+Sf1a+linker+lectin+linker+His-tag+terminator
 +
<br>PANTIDE that contains the gene coding for Sf1a and the snowdrop lectin linked with His-tag.
 +
 +
</p>
 +
 +
<!--------OAIP lectin------>
 +
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974023" class="biobrick"><p class="content-1">BBa_K1974023</p></a>
 +
 +
<div>
 +
      <img src="https://static.igem.org/mediawiki/2016/2/27/NCTU_FORMOSA_OL.png" class="picture">
 +
</div>
 +
 +
<p class="content">
 +
T7 promoter+RBS+OAIP+linker+lectin+linker+His-tag+terminator
 +
<br>PANTIDE that contains the gene coding for OAIP and the snowdrop lectin linked with His-tag.
 +
 +
</p>
 +
 +
<!---------------GS linker----------------->
 +
 +
<p class="title">PANTIDE with GS Linker</p>
 +
<p class="content"> In this part, to facilitate the expression of PANTIDE, we extended the linker with the substitution of GS linker that is even better than the original linker for the more correcter folding of the proteins structure.</p>
 +
 +
<!-----------OAIP GS linker--------------->
 +
<a href="http://parts.igem.org/wiki/index.php?title=Part:BBa_K1974033" class="biobrick"><p class="content-1">BBa_K1974033</p></a>
 +
 +
<div>
 +
      <img src="https://static.igem.org/mediawiki/2016/d/d6/NCTU_FORMOSA_GS1.png" class="picture">
 +
</div>
 +
 +
<p class="content">
 +
T7 promoter+RBS+Hv1a+GS linker+lectin+linker+His-tag+terminator
 +
<br>PANTIDE that contains the gene coding for Hv1a and the snowdrop lectin. And improve the correct folding of PANTIDE by substituting with GS linker linked with His-tag.
 +
 +
 +
</p>
 +
 +
 +
 +
</div>
 
</section>
 
</section>
−
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+
 
 +
 
 +
 
 +
 
 +
 
  
 
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var stepDistance = 100;
+
var stepDistance = 101;
 
var documentHeight = window.innerHeight;
 
var documentHeight = window.innerHeight;
 
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var positions = [];
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{{Team:NCTU_Formosa/footer}}
 
{{Team:NCTU_Formosa/footer}}

Latest revision as of 03:13, 4 November 2016

Composite Parts

We have known the function of the three toxic peptides of PANTIDE, Omega-hexatoxin-Hv1a, µ-segestritoxin-Sf1a, and Orally Active Insecticidal Peptide(OAIP) respectively and they are easy to express with the help of the bacteria E. coli BL21 Rosetta Gami strain. So, we then start to create BioBricks of these three toxins which we have obtained the nucleotide sequences. We use a strong T7 promoter that is suitable with the strain of E. coli we chose and also powerful at expressing proteins with multiple disulfide bonds, and we add His-tag for protein purification.

PANTIDE without Lectin

PANTIDE can be a single toxic peptide of the three toxins we chose, Hv1a, Sf1a, and OAIP respectively, which derive from the natural predators spiders.

BBa_K1974011

T7 promoter+RBS+Hv1a+linker+His-tag+terminator
PANTIDE that contains the gene coding for Hv1a linked with His-tag.

BBa_K1974012

T7 promoter+RBS+Sf1a+linker+His-tag+terminator
PANTIDE that contains the gene coding for Sf1a linked with His-tag.

BBa_K1974013

T7 promoter+RBS+OAIP+linker+His-tag+terminator
PANTIDE that contains the gene coding for OAIP linked with His-tag.

PANTIDE with Lectin

PANTIDE can be a single toxic peptide of the three toxins we chose, Hv1a, Sf1a, and OAIP respectively with the addition of Galanthus nivalis agglutinin (GNA), for GNA has the function of acting as a carrier of PANTIDE from insect gut to the haemolymph and finally PANTIDE can transmit to the action site.

BBa_K1974021

T7 promoter+RBS+Hv1a+linker+lectin+linker+His-tag+terminator
PANTIDE that contains the gene coding for Hv1a and the snowdrop lectin linked with His-tag.

BBa_K1974022

T7 promoter+RBS+Sf1a+linker+lectin+linker+His-tag+terminator
PANTIDE that contains the gene coding for Sf1a and the snowdrop lectin linked with His-tag.

BBa_K1974023

T7 promoter+RBS+OAIP+linker+lectin+linker+His-tag+terminator
PANTIDE that contains the gene coding for OAIP and the snowdrop lectin linked with His-tag.

PANTIDE with GS Linker

In this part, to facilitate the expression of PANTIDE, we extended the linker with the substitution of GS linker that is even better than the original linker for the more correcter folding of the proteins structure.

BBa_K1974033

T7 promoter+RBS+Hv1a+GS linker+lectin+linker+His-tag+terminator
PANTIDE that contains the gene coding for Hv1a and the snowdrop lectin. And improve the correct folding of PANTIDE by substituting with GS linker linked with His-tag.