Team:NYMU-Taipei/ymis4.html

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   <p>Sulfite reductase stable expressing cyanobacteria cost longer time and a  lot of money to set up. To check the function of constructed gene in advance,  we perform a chemical nephelometry assay to analyze the  function. We follow a paper done by Prof. I. Lavilla (Microvolume  turbidimetry for rapid and sensitive determination of the acid labile sulfide  fraction in waters after headspace single-drop microextraction with <em>in situ </em>generation of volatile hydrogen  sulfide, Analytica Chimica Acta 647 (2009) 112–116) to detect H2S. Due to our pTrc-kan-Cys I and pTrc-kan-Dsr  gene can turn on in E.coli, we transform these gene into DH5 alpha E.coli and  incubate in HSO3- contained LB medium. After 24hr and 48hr, we check  the medium and detect H2S concentration.    <br />
   <p>Sulfite reductase stable expressing cyanobacteria cost longer time and a  lot of money to set up. To check the function of constructed gene in advance,  we perform a chemical nephelometry assay to analyze the  function. We follow a paper done by Prof. I. Lavilla (Microvolume  turbidimetry for rapid and sensitive determination of the acid labile sulfide  fraction in waters after headspace single-drop microextraction with <em>in situ </em>generation of volatile hydrogen  sulfide, Analytica Chimica Acta 647 (2009) 112–116) to detect H2S. Due to our pTrc-kan-Cys I and pTrc-kan-Dsr  gene can turn on in E.coli, we transform these gene into DH5 alpha E.coli and  incubate in HSO3- contained LB medium. After 24hr and 48hr, we check  the medium and detect H2S concentration.    <br />
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   <div class=out style='text-align:center'><img src="images/s12.gif" alt="" width="554" height="220" /><br />
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   <div class=out style='text-align:center'><img src="http://igem.org/wiki/images/5/5c/Ymis12.gif" alt="" width="554" height="220" /><br />
     <p align="center">Using Microvolume turbidimetry method can detect micro amount H2S</p>
     <p align="center">Using Microvolume turbidimetry method can detect micro amount H2S</p>
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     (The same method used  in Environmental Protection Administration  Executive Yuan, Taiwan    <a href="http://www.niea.gov.tw/niea/AIR/A40671A.htm">http://www.niea.gov.tw/niea/AIR/A40671A.htm</a> )</p>
     (The same method used  in Environmental Protection Administration  Executive Yuan, Taiwan    <a href="http://www.niea.gov.tw/niea/AIR/A40671A.htm">http://www.niea.gov.tw/niea/AIR/A40671A.htm</a> )</p>
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   <div class=out style='text-align:center'><img src="images/s13.gif" alt="" width="548" height="246" /><br />
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   <div class=out style='text-align:center'><img src="http://igem.org/wiki/images/2/21/Ymis13.gif" alt="" width="548" height="246" /><br />
     <p>DPDA structure and DPDA solution, when H2S exist, DPDA solution turn  turbid </p></div>
     <p>DPDA structure and DPDA solution, when H2S exist, DPDA solution turn  turbid </p></div>
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         <div class=out style='text-align:center'><img src="images/s14.gif" alt="" width="553" height="263" /><br />
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         <div class=out style='text-align:center'><img src="http://igem.org/wiki/images/2/29/Ymis14.gif" alt="" width="553" height="263" /><br />
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         H2S serial dilution and standard curve detection        <br />
         H2S serial dilution and standard curve detection        <br />
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       <div class=out style='text-align:center'><span class="out" style="text-align:center"><img src="images/s15.gif" alt="" width="553" height="263" /></span><br />
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       <div class=out style='text-align:center'><span class="out" style="text-align:center"><img src="http://igem.org/wiki/images/5/57/Ymis15.gif" alt="" width="553" height="263" /></span><br />
         <p>Fortunately, we find out our Cys I and Dsr  enzyme are working!! </p>
         <p>Fortunately, we find out our Cys I and Dsr  enzyme are working!! </p>
       </div></div></div>
       </div></div></div>

Revision as of 03:42, 27 September 2012

NYMU iGEM

Result

Function of Sulfite in E.coli

Sulfite reductase stable expressing cyanobacteria cost longer time and a lot of money to set up. To check the function of constructed gene in advance, we perform a chemical nephelometry assay to analyze the function. We follow a paper done by Prof. I. Lavilla (Microvolume turbidimetry for rapid and sensitive determination of the acid labile sulfide fraction in waters after headspace single-drop microextraction with in situ generation of volatile hydrogen sulfide, Analytica Chimica Acta 647 (2009) 112–116) to detect H2S. Due to our pTrc-kan-Cys I and pTrc-kan-Dsr gene can turn on in E.coli, we transform these gene into DH5 alpha E.coli and incubate in HSO3- contained LB medium. After 24hr and 48hr, we check the medium and detect H2S concentration.


Using Microvolume turbidimetry method can detect micro amount H2S


Instead of using complex machine and expensive HPLC, we use Zn(NH3)4·(OH)2 to absorp H2S, wich can form complex salt,〔Zn(OH3)6〕S. And then, add N,N–dimethy1-1,4-phenylenediammoniumdichloride(DPDA solution) and ferrite chloride to form methyl blue. Finally, by analyzing O.D.670, we can easily get H2S concentration. The reaction fomula are as followed:

H2S+Zn(NH3)4·(OH)2→〔Zn(OH3)6〕S+2H2O

(The same method used in Environmental Protection Administration Executive Yuan, Taiwan    http://www.niea.gov.tw/niea/AIR/A40671A.htm )


DPDA structure and DPDA solution, when H2S exist, DPDA solution turn turbid



H2S serial dilution and standard curve detection


Fortunately, we find out our Cys I and Dsr enzyme are working!!