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Team:Sumbawagen/future direction
2015-02-19T07:13:45Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2><b>Future Directions</b></h2><br />
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<p> With limited facilities such as in availability of high speed microcentrifuge, UV and fluorescence spectrophotometers, incubators with shakers, we were able to adapt several standard protocols – written in Notebook section – to achieve certains goals in synthetic biology. We have been able to construct 2 new parts namely, crr gene encoding IIA(Glc) from E. coli alone (BBa_K1508002), and crr gene with double terminator (BBa_K1508003), which we have been deposited in iGEM Registry. Both plasmids have been sequenced for confirmation.</p><br />
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<p> In term of our projects goal on measuring honey glucose concentration, we already succeeded in developing protocols of measurements using (1) glucose meter, and (2) mRFP reporter gene expression based on the phenomenon of catabolite repression. The second protocol – called ECONEY – was a combination of innovative color intensity measurement with mobile phone, programming has advantage of easy use, and attractive appearance. Both protocols are very affordable for honey farmers in Sumbawa Island who still out of access for conventional method such as glucose refractometer. Therefore, our human practice has big success in introducing the power of synthetic biology to our people who were never exposed to knowledge in biotechnology although already used several products such as glucose meter and recombinant insulin which are available in drug stores.</p><br />
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<p> In the future, we will use these experiences to continue constructing BBa_J04450 + our original constructs (containing either crr or cyaA) and see whether they will really influence the catabolite repression sensitivity, resulted in shifted linear range of measureable region of glucose concentration. If this has been achieved, we can replace mRFP with amilCP (blue color), then we will able to get different color of medium when different honey added to the medium. This system can be used to easily and interestingly shown whether the honey is original Sumbawa or from other sources because they have different concentration of glucose</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/future_direction
Team:Sumbawagen/future direction
2015-02-19T07:12:19Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2><b>Future Directions</b></h2><br />
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<p>With limited facilities such as in availability of high speed microcentrifuge, UV and fluorescence spectrophotometers, incubators with shakers, we were able to adapt several standard protocols – written in Notebook section – to achieve certains goals in synthetic biology. We have been able to construct 2 new parts namely, crr gene encoding IIA(Glc) from E. coli alone (BBa_K1508002), and crr gene with double terminator (BBa_K1508003), which we have been deposited in iGEM Registry. Both plasmids have been sequenced for confirmation.</p><br />
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<p> In term of our projects goal on measuring honey glucose concentration, we already succeeded in developing protocols of measurements using (1) glucose meter, and (2) mRFP reporter gene expression based on the phenomenon of catabolite repression. The second protocol – called ECONEY – was a combination of innovative color intensity measurement with mobile phone, programming has advantage of easy use, and attractive appearance. Both protocols are very affordable for honey farmers in Sumbawa Island who still out of access for conventional method such as glucose refractometer. Therefore, our human practice has big success in introducing the power of synthetic biology to our people who were never exposed to knowledge in biotechnology although already used several products such as glucose meter and recombinant insulin which are available in drug stores.</p><br />
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<p>In the future, we will use these experiences to continue constructing BBa_J04450 + our original constructs (containing either crr or cyaA) and see whether they will really influence the catabolite repression sensitivity, resulted in shifted linear range of measureable region of glucose concentration. If this has been achieved, we can replace mRFP with amilCP (blue color), then we will able to get different color of medium when different honey added to the medium. This system can be used to easily and interestingly shown whether the honey is original Sumbawa or from other sources because they have different concentration of glucose</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/project/software
Team:Sumbawagen/project/software
2015-02-19T04:40:10Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2><b>Software</b></h2><br />
<br />
<p>Not everybody knows how to measure glucose concentration in honey. Our team found the best way to apply our project, by creating a device that capable to measure the glucose concentration in honey. To ease people use the device, we create a simple software, which is cheap and easy to use </p><br />
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<p> ECONEY Project is a colaboration between biotechnology and informatics technology. We createECONEYsoftwarebased on android smartphone technology, which can be used to measure glucose concentration using catabolite repression phenomenon. ECONEY software will detect RGB value and color code from single picture captured from smartphone camera.Then the RGB Luminance value will be calculated using the RGB Value detected earlier and the result showed on screen directly.Our software has graphics submenu which provide you with the standard curve. This curve is used to calculate the glucose concentration of honey.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/project/software
Team:Sumbawagen/project/software
2015-02-19T04:38:37Z
<p>Adel: </p>
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<p>Not everybody knows how to measure glucose concentration in honey. Our team found the best way to apply our project, by creating a device that capable to measure the glucose concentration in honey. To ease people use the device, we create a simple software, which is cheap and easy to use </p><br />
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<p> ECONEY Project is a colaboration between biotechnology and informatics technology. We createECONEYsoftwarebased on android smartphone technology, which can be used to measure glucose concentration using catabolite repression phenomenon. ECONEY software will detect RGB value and color code from single picture captured from smartphone camera.Then the RGB Luminance value will be calculated using the RGB Value detected earlier and the result showed on screen directly.Our software has graphics submenu which provide you with the standard curve. This curve is used to calculate the glucose concentration of honey.</p><br />
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http://2014.igem.org/Team:Sumbawagen/Notebook3
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<p>Not everybody knows how to measure glucose concentration in honey. Our team found the best way to apply our project, by creating a device that capable to measure the glucose concentration in honey. To ease people use the device, we create a simple software, which is cheap and easy to use. <br><br></p><br />
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<p>ECONEY Project is acolaboration between biotechnology and informatics technology. We createECONEYsoftwarebased on android smartphone technology, which can be used to measure glucose concentration using catabolite repression phenomenon. ECONEY software will detect RGB value and color code from single picture captured from smartphone camera.Then the RGB Luminance value will be calculated using the RGB Value detected earlier and the result showed on screen directly.Our software has graphics submenu which provide you with the standard curve. This curve is used to calculate the glucose concentration of honey. <br><br></p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Attributions
Team:Sumbawagen/Attributions
2015-02-19T04:13:47Z
<p>Adel: </p>
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>Attributions</h2><br />
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<p>Our team consists of 15 students, 5 instructors, and 2 advisors. We wouldn't have been able to do much without the sponsors, Sumbawa society, and also blessing from God.So, we would like to show our gratitude to the people who were very important for us:</p><br><br />
<br />
<ul style="padding-left:15px"><br />
<li><strong>Sumbawa University of Technology</strong>, which has facilitated us in term of administration, registration and raising funds. So, Sumbawagen team can join this competition.</li><br />
<li><strong>BLKM (Public Health Laboratory) of Sumbawa island</strong>, which has facilitated a laboratory and tools for the wetlab activity.</li><br />
<li><strong>Alzyme</strong>, which has facilitated us in providing tools and materials for wetlab activity.</li><br />
<li><strong>KPHP (Provincial Forest Management Unit) of Batu Lanteh</strong>, Sumbawa island, which has facilitated us in collecting honey sample.</li><br />
<li><strong>The Goverment of Sumbawa Regency</strong>, which has supported our project.</li><br />
<li><a href ="https://2014.igem.org/Team:Dundee"><strong>Dundee iGEM Team</strong> </a>, thanks for wiki Template.</li> <br />
<li><strong>And also for Sumbawa society</strong>, expecially for Sumbawa honey farmers that always give us support to finish this project.</li></ul><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Attributions
Team:Sumbawagen/Attributions
2015-02-19T04:12:52Z
<p>Adel: </p>
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>Attributions</h2><br />
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<br />
<p>Our team consists of 15 students, 5 instructors, and 2 advisors. We wouldn't have been able to do much without the sponsors, Sumbawa society, and also blessing from God.So, we would like to show our gratitude to the people who were very important for us:</p><br><br />
<br />
<ul style="padding-left:15px"><br />
<li><strong>Sumbawa University of Technology</strong>, which has facilitated us in term of administration, registration and raising funds. So, Sumbawagen team can join this competition.</li><br />
<li><strong>BLKM (Public Health Laboratory) of Sumbawa island</strong>, which has facilitated a laboratory and tools for the wetlab activity.</li><br />
<li><strong>Alzyme</strong>, which has facilitated us in providing tools and materials for wetlab activity.</li><br />
<li><strong>KPHP (Provincial Forest Management Unit) of Batu Lanteh</strong>, Sumbawa island, which has facilitated us in collecting honey sample.</li><br />
<li><strong>The Goverment of Sumbawa Regency</strong>, which has supported our project.</li><br />
<li><a href ="https://2014.igem.org/Team:Dundee/"><strong>Dundee iGEM Team</strong> </a>, thanks for wiki Template.</li> <br />
<li><strong>And also for Sumbawa society</strong>, expecially for Sumbawa honey farmers that always give us support to finish this project.</li></ul><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Attributions
Team:Sumbawagen/Attributions
2015-02-19T04:01:43Z
<p>Adel: </p>
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>Attributions</h2><br />
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<p>Our team consists of 15 students, 5 instructors, and 2 advisors. We wouldn't have been able to do much without the sponsors, Sumbawa society, and also blessing from God.So, we would like to show our gratitude to the people who were very important for us:</p><br><br />
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<ul style="padding-left:15px"><br />
<li><strong>Sumbawa University of Technology</strong>, which has facilitated us in term of administration, registration and raising funds. So, Sumbawagen team can join this competition.</li><br />
<li><strong>BLKM (Public Health Laboratory) of Sumbawa island</strong>, which has facilitated a laboratory and tools for the wetlab activity.</li><br />
<li><strong>Alzyme</strong>, which has facilitated us in providing tools and materials for wetlab activity.</li><br />
<li><strong>KPHP (Provincial Forest Management Unit) of Batu Lanteh</strong>, Sumbawa island, which has facilitated us in collecting honey sample.</li><br />
<li><strong>The Goverment of Sumbawa Regency</strong>, which has supported our project.</li><br />
<li><strong>Dundee iGEM Team</strong>, thanks for wiki Template.</li><br />
<li><strong>And also for Sumbawa society</strong>, expecially for Sumbawa honey farmers that always give us support to finish this project.</li></ul><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol8
Team:Sumbawagen/Notebook/protocol8
2015-02-19T03:52:07Z
<p>Adel: </p>
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<h2>Notebook – Protocol – 8. Transformation (Heat Shock)</h2<br />
<br />
<p>Protocols from the kit were followed by adaptation because we have no high speed micro centrifuge, but only a 6,000 rpm small micro centrifuge </p><br />
<br />
<p>1. Add plasmid into tubes containing E. coli competent cell.</p><br />
<p>2. Mix by pipetting using micropipette (5-10 times pipetting). </p><br />
<p>3. Incubate at 4 °C for 40 minutes (inside a refrigerator).</p><br />
<p>4. Prepare the hot water with the temperature of 42 °C, measure the temperature using thermometer. </p><br />
<p>5. Heat shock the cells by put the bottom of the tube on hot water for 45 seconds.</p><br />
<p>6. Incubate at 4 °C for 5 minutes.</p><br />
<p>7. Add 1 ml LB liquid medium sterile (without antibiotic) into each tubes.</p><br />
<p>8. Incubate using shaker for 2 hours</p><br />
<p>9. Plating the transformation by add 100 µl transformation reaction to LB agar plates containing chloramphenicol (final concentration 50 µg/ml).</p><br />
<p>10. Give label for the plates.</p><br />
<p>11. Centrifuge for 6,000 rpm for 3 minutes.</p><br />
<p>12. Discard the flow-through as much as 700 µl.</p><br />
<p>13. Suspend the remaining flow-through and pellets by pipetting.</p><br />
<p>14. Add 100 µl to agar plate containing appropriate antibiotic.</p><br />
<p>15. Give label for the plates.</p><br />
<p>16. Incubate at 37 °C overnight.</p><br />
<p>17. Check the result later. </p><br />
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<h2> Notebook – Protocol – 10. DNA Sequencing</h2><br />
<p> DNA sequencing was done by ordering to commercial company.</a></p><br />
<p>1. Premix sample was prepared with the following composition:</p><br />
<ul style="padding-left:25px;"><br />
<li> Plasmid sample, 10 to 20 ul*</li><br />
<li>10 uM primer, 1 ul </li><br />
<li> Sterilized distilled water, 9 to 19 ul</li><br />
Total volume, 21 ul </li><br><br />
Concentration < 900 ng </li><br />
</ul><br />
<br />
<p>2. Samples were sent to the company<br><br />
<p> 3. According to the company, DNA sequencing was done using the following conditions:<br><br />
<ul style="padding-left:25px;"><br />
<li> Sequencing kit, BigDye® Terminator v3.1 Cycle Sequencing Kit (Applied Biosystems)</li><br />
<li> Purification kit, BigDye XTerminator® Purification Kit (Applied Biosystems) </u></li><br />
<li> Sequencer, Applied Biosystems 3730xl DNA Analyzer</li></p></ul><br />
<br />
<al><p>4. DNA sequencing data in .ab1 files were received by email, then analyzed using Bioedit program. For alignment, online CLUSTAL program was used.</p></al><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/Results1
Team:Sumbawagen/Notebook/Results1
2015-01-20T22:01:11Z
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<h2><b>Notebook – Results – 1. Construction of BBa_J04450 + constitutive promoter/crr or cyaA</b></h2><br />
<br />
<br />
<p>We planned to construct BBa_J04450 as circuit #1 (original circuit) followed downstream by our original circuit #2. Circuit #2 is composed by BBa_J2319 (constitutive promoter) / BBa_B0034 (RBS) / crr (IIA(Glc) or cyaA (Adenylate cyclase) / BBa_B0015 (double terminator).</p><br />
<br />
<p>So the first step was to clone either cyaA or crr from E. coli to Biobrick construct. PCR fragments with correct size were able to be obtained. After direct restriction with EcoRI and SpeI, the fragments were cloned into pSB1C3 backbone. </p><br />
<br />
<p>Next step was to construct, BBa_J2319 (chloramphenicol) / BBa_B0034 (ampicillin) and crr or cyaA (chloramphenicol) / BBa_B0015 (chloramphenicol). For BBa_J2319 / BBa_B0034 construction, prefix insertion was used. Thus ligation product was screened on ampicillin LB agar plate. While for crr or cyaA / BBa_B0015 construction, suffix insertion was used because the large size of crr or cyaA could be isolated by agarose gel excision. After BBa_J2319 / BBa_0034 (ampicillin) and crr or cyaA / BBa_0015 (chloramphenicol) constructs were obtained, prefix insertion was done to get BBa_J2319 / BBa_B0034 / crr or cyaA / BBa_B0015 in pSB1C3 backbone. Final step was prefix insertion of BBa_J00450 the original constructs.</p><br />
<br />
<p>In every steps of works, we only confirmed the insertion by colony selection in appropriate antibiotics containing LB agar plate, and purification of backbone plasmid for insertion by gel electrophoresis to completely separated small insert fragments. Thus in theory there will be no self-ligation. However for either prefix or suffix inserts, the digested samples were not purified, instead used directly for ligation reaction. </p><br />
<br />
<p>Below some representative gel electrophoresis data were shown.</p><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/24/Project_-_1_fig_1.jpg"></center><br><br />
<center>Figure 1. Cloning of adenylate cyclease full sequence (ACFL), adenylate cyclase catalytic domain (ACCD) and IIA(Glc) (2AG) by PCR</center><br><br />
<br />
<br><br />
<center><img src="https://static.igem.org/mediawiki/2014/4/45/Sumbawagen.result1.pict2.jpg"></center><br><br />
<center>Figure 2. Confirmation of insertion of ACFL, ACCD and IIA(Glc) into pSB1C3 by colony PCR. Only IIA(Glc) was confirmed to be inserted by increasing band size in this figure.</center><br><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/Results1
Team:Sumbawagen/Notebook/Results1
2015-01-20T22:00:10Z
<p>Adel: </p>
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<h2>Notebook – Results – 1. Construction of BBa_J04450 + constitutive promoter/crr or cyaA</h2><br />
<br />
<br />
<p>We planned to construct BBa_J04450 as circuit #1 (original circuit) followed downstream by our original circuit #2. Circuit #2 is composed by BBa_J2319 (constitutive promoter) / BBa_B0034 (RBS) / crr (IIA(Glc) or cyaA (Adenylate cyclase) / BBa_B0015 (double terminator).</p><br />
<br />
<p>So the first step was to clone either cyaA or crr from E. coli to Biobrick construct. PCR fragments with correct size were able to be obtained. After direct restriction with EcoRI and SpeI, the fragments were cloned into pSB1C3 backbone. </p><br />
<br />
<p>Next step was to construct, BBa_J2319 (chloramphenicol) / BBa_B0034 (ampicillin) and crr or cyaA (chloramphenicol) / BBa_B0015 (chloramphenicol). For BBa_J2319 / BBa_B0034 construction, prefix insertion was used. Thus ligation product was screened on ampicillin LB agar plate. While for crr or cyaA / BBa_B0015 construction, suffix insertion was used because the large size of crr or cyaA could be isolated by agarose gel excision. After BBa_J2319 / BBa_0034 (ampicillin) and crr or cyaA / BBa_0015 (chloramphenicol) constructs were obtained, prefix insertion was done to get BBa_J2319 / BBa_B0034 / crr or cyaA / BBa_B0015 in pSB1C3 backbone. Final step was prefix insertion of BBa_J00450 the original constructs.</p><br />
<br />
<p>In every steps of works, we only confirmed the insertion by colony selection in appropriate antibiotics containing LB agar plate, and purification of backbone plasmid for insertion by gel electrophoresis to completely separated small insert fragments. Thus in theory there will be no self-ligation. However for either prefix or suffix inserts, the digested samples were not purified, instead used directly for ligation reaction. </p><br />
<br />
<p>Below some representative gel electrophoresis data were shown.</p><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/24/Project_-_1_fig_1.jpg"></center><br><br />
<center>Figure 1. Cloning of adenylate cyclease full sequence (ACFL), adenylate cyclase catalytic domain (ACCD) and IIA(Glc) (2AG) by PCR</center><br><br />
<br />
<br><br />
<center><img src="https://static.igem.org/mediawiki/2014/4/45/Sumbawagen.result1.pict2.jpg"></center><br><br />
<center>Figure 2. Confirmation of insertion of ACFL, ACCD and IIA(Glc) into pSB1C3 by colony PCR. Only IIA(Glc) was confirmed to be inserted by increasing band size in this figure.</center><br><br />
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<h2>A. Team Discussion</h2><br />
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<p>Policy and practices is a way for us to introduce our project to society. We discussed our policy and practices project first on June 23, 2014. The points of our discussion were determining our survey targets and making questionnaire design. After we shared our best ideas for the project, we decided the purpose of the project which is to introduce ECONEY to Sumbawa society and to discover more information about everything related to Sumbawa honey. Because of our questionnaire design was still not complete, we had another disscussions then we report to our instructors to get the revisions. The problems were about the question complexity and its relation to the project. And finally, our questionnaire finished on July 6th.<br></p><br />
<center><img src="https://static.igem.org/mediawiki/2014/4/40/Team_Disscustion.jpg"></center><br><br />
<br />
<p>The next discussion was about the respondents. We determined the region target and the number of the respondents. We devided the respondents in honey production regions. We divided the respondents into producers, distributors, and consumers. We targetted to have 630 respondents which consist of 210 producers, 210 distributors, and 210 consumers using random sampling design.<br></p><br />
<br />
<h2>B. Idea</h2><br />
<br />
We surveyed 16 regions consist of 7 producer regions and 9 consumer regions, while the distributors were exist in both of the areas. The producer regions located in villages. On the other hand, we choose Sumbawa Besar (the capital city of Sumbawa regency) as consumer regions. Sumbawagen team then divided into 7 teams to do the survey. Each team did the survey at 1 producer regions and 1 consumer regions. The survey was held on July 14-17 2014.<br><br />
<br />
<br />
<h2>C. Data Analysis</h2><br />
We would like to give our gratitude to all Sumbawa society that had been participated in our policy and practices project. There are 672 respondents which consist of 130 producers, 71 distributors, and 471 consumers participated in the project. The data analysis were given below:<br><br><br />
<br />
<strong><i>1. Have you ever consume Sumbawa honey?</i></strong><br><br />
Most of our respondents have consumed Sumbawa honey. It shows that Sumbawa honey is a very important commodity for Sumbawa society.<br><br />
<br />
<center><img src="https://static.igem.org/mediawiki/2014/9/94/Diagram1.jpg"></center><br><br />
<br />
<strong><i>2. Is there any tools which can be used to measure glucose level in Sumbawa honey?</i></strong><br />
<p>Many of our respondents said they don’t know the tools to measure the glucose level in Sumbawa honey.</P><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/0c/Diagram_2.jpg"></center><br><br />
<br />
<strong><i>3. If the answer for no. 2 is YES, please mention the tools</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/24/Diagram_3.jpg"></center><br><br />
<p>None of our respondents could mention the tool used to measure glucose level in Sumbawa honey.<br> <br />
Although some of our respondents said that there is a tool exist to measure glucose in Sumbawa honey, they couldn’t mention the name. After interviewed them little bit deeper we figured out that society has perception that the tool is the same with the tool used to measure water level, but the fact it's different. If they want to know glucose level in Sumbawa honey (especially the distributor) they have to send honey sample to Mataram (the capital city of West Nusa Tenggara province).<br></p><br />
<br />
<strong><i>4. If there is a tool created to measure the glucose level in honey, how would you respond?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/d/d9/Sumbawagen.4.jpg"></center><br><br />
<p>The society gave the positive response for this question. Most of producers want to use (58%), and just 7% didn’t care about it. The distributor respondents who want to learn and use the tool are almost same (47% and 46%). Meanwhile, most of the consumers want to learn (53%).<br></p><br />
<br />
<br />
<strong><i>5. Are you android smartphone user?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/b/ba/Diagram_5.jpg"></center><br><br />
<p>In Sumbawa society, android smartphone is familiar among teens. Hence in the survey, only a small number of respondents using android smartphone: producer is only 29%, distributor is only 15%, and the consumer is only 30%. <br></p><br />
<br />
<strong><i>6. If the answer of question number 5 is “YES”, are you using android smartphone application?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/03/Diagram_6.jpg"></center><br><br />
<p>Most of android user use android aplication beacuse there are various and cheap applications in android market, even some are distributed for free.<br></p><br />
<br />
<strong><i>7. If the answer is “YES” for question number 6, is android smartphone application easy to use?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/7/7e/Diagram_7.jpg"></center><br><br />
<p>All of our producer respondents agree if android smartphone applications are easy to use. Whereas 40% from our distributor respondents and 5% from consumer respondents said that the applications are difficult to use.<br></p><br />
<br />
<strong><i>8. If the application can meassure glucose level in honey, what is your response?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/5/57/8_diagram.jpg"></center><br><br />
<p>Our respondents were very interest with our project especially for producer and distributor category as show from the survey result.<br> </p><br />
<br />
<strong><i>9. As sumbawa people, are you proud of Sumbawa honey?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/6/6b/Diagram_9.jpg"></center><br><br />
<p>We can conclude that Sumbawa people are absolutely proud of Sumbawa honey. Sumbawa honey is the icon of Sumbawa.</p><br />
<br />
<h2>Conclusion</h2><br />
<p>Sumbawa honey as the icon of Sumbawa contains high glucose level. However, there is no tools for measuring glucose concentration exist in Sumbawa. That's why people of Sumbawa they really appreciate our ECONEY project. We hope our ECONEY project could be applied to help people in Sumbawa measure glucose level in honey.</p><br />
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Adel
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2014-11-22T11:29:17Z
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Adel
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2014-11-20T07:40:35Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>A. Team Discussion</h2><br />
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<p>Policy and practices is a way for us to introduce our project to society. We discussed our policy and practices project first on June 23, 2014. The points of our discussion were determining our survey targets and making questionnaire design. After we shared our best ideas for the project, we decided the purpose of the project which is to introduce ECONEY to Sumbawa society and to discover more information about everything related to Sumbawa honey. Because of our questionnaire design was still not complete, we had another disscussions then we report to our instructors to get the revisions. The problems were about the question complexity and its relation to the project. And finally, our questionnaire finished on July 6th.<br></p><br />
<center><img src="https://static.igem.org/mediawiki/2014/4/40/Team_Disscustion.jpg"></center><br><br />
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<p>The next discussion was about the respondents. We determined the region target and the number of the respondents. We devided the respondents in honey production regions. We divided the respondents into producers, distributors, and consumers. We targetted to have 630 respondents which consist of 210 producers, 210 distributors, and 210 consumers using random sampling design.<br></p><br />
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<h2>B. Idea</h2><br />
<br />
We surveyed 16 regions consist of 7 producer regions and 9 consumer regions, while the distributors were exist in both of the areas. The producer regions located in villages. On the other hand, we choose Sumbawa Besar (the capital city of Sumbawa regency) as consumer regions. Sumbawagen team then divided into 7 teams to do the survey. Each team did the survey at 1 producer regions and 1 consumer regions. The survey was held on July 14-17 2014.<br><br />
<br />
<br />
<h2>C. Data Analysis</h2><br />
We would like to give our gratitude to all Sumbawa society that had been participated in our policy and practices project. There are 672 respondents which consist of 130 producers, 71 distributors, and 471 consumers participated in the project. The data analysis were given below:<br><br><br />
<br />
<strong><i>1. Have you ever consume Sumbawa honey?</i></strong><br><br />
Most of our respondents have consumed Sumbawa honey. It shows that Sumbawa honey is a very important commodity for Sumbawa society.<br><br />
<br />
<center><img src="https://static.igem.org/mediawiki/2014/9/94/Diagram1.jpg"></center><br><br />
<br />
<strong><i>2. Is there any tools which can be used to measure glucose level in Sumbawa honey?</i></strong><br />
<p>Many of our respondents said they don’t know the tools to measure the glucose level in Sumbawa honey.</P><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/0c/Diagram_2.jpg"></center><br><br />
<br />
<strong><i>3. If the answer for no. 2 is YES, please mention the tools</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/24/Diagram_3.jpg"></center><br><br />
<p>None of our respondents could mention the tool used to measure glucose level in Sumbawa honey.<br> <br />
Although some of our respondents said that there is a tool exist to measure glucose in Sumbawa honey, they couldn’t mention the name. After interviewed them little bit deeper we figured out that society has perception that the tool is the same with the tool used to measure water level, but the fact it's different. If they want to know glucose level in Sumbawa honey (especially the distributor) they have to send honey sample to Mataram (the capital city of West Nusa Tenggara province).<br></p><br />
<br />
<strong><i>4. If there is a tool created to measure the glucose level in honey, how would you respond?</i></strong><br><br />
<p>The society gave the positive response for this question. Most of producers want to use (58%), and just 7% didn’t care about it. The distributor respondents who want to learn and use the tool are almost same (47% and 46%). Meanwhile, most of the consumers want to learn (53%).<br></p><br />
<br />
<br />
<strong><i>5. Are you android smartphone user?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/b/ba/Diagram_5.jpg"></center><br><br />
<p>In Sumbawa society, android smartphone is familiar among teens. Hence in the survey, only a small number of respondents using android smartphone: producer is only 29%, distributor is only 15%, and the consumer is only 30%. <br></p><br />
<br />
<strong><i>6. If the answer of question number 5 is “YES”, are you using android smartphone application?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/03/Diagram_6.jpg"></center><br><br />
<p>Most of android user use android aplication beacuse there are various and cheap applications in android market, even some are distributed for free.<br></p><br />
<br />
<strong><i>7. If the answer is “YES” for question number 6, is android smartphone application easy to use?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/7/7e/Diagram_7.jpg"></center><br><br />
<p>All of our producer respondents agree if android smartphone applications are easy to use. Whereas 40% from our distributor respondents and 5% from consumer respondents said that the applications are difficult to use.<br></p><br />
<br />
<strong><i>8. If the application can meassure glucose level in honey, what is your response?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/5/57/8_diagram.jpg"></center><br><br />
<p>Our respondents were very interest with our project especially for producer and distributor category as show from the survey result.<br> </p><br />
<br />
<strong><i>9. As sumbawa people, are you proud of Sumbawa honey?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/6/6b/Diagram_9.jpg"></center><br><br />
<p>We can conclude that Sumbawa people are absolutely proud of Sumbawa honey. Sumbawa honey is the icon of Sumbawa.</p><br />
<br />
<h2>Conclusion</h2><br />
<p>Sumbawa honey as the icon of Sumbawa contains high glucose level. However, there is no tools for measuring glucose concentration exist in Sumbawa. That's why people of Sumbawa they really appreciate our ECONEY project. We hope our ECONEY project could be applied to help people in Sumbawa measure glucose level in honey.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook
Team:Sumbawagen/Notebook
2014-10-18T03:50:41Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2><b>POLICY AND PRACTICE</b> </h2><br />
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<h3><b>Part 1 Survey to Society</b> </h3><br><br />
<br />
<ul style="padding-left:25px;"><br />
<p>1. 1 - 4th July, 2014</p><br />
<p>Discussion of Human Practice (now Policy & Practice) project plan </p><br />
<p>2. 8 - 11th July, 2014</p><br />
<p>Permission confirmation to the district and village office to do the survey for the society</p><br />
<p>3. 14 - 17th July, 2014</p><br />
<p>Data collection (survey)</p><br />
<p>4. 21th July, 2014</p><br />
<p>Data analysis </p><br />
<p>5. 22 - 26th July, 2014</p><br />
<p>Result discussion</p><br />
</p></ul><br />
<br />
<br />
<h3><b>Part 2 Demo of Our Laboratory Activity</b> </h3><br><br />
<br />
<ul style="padding-left:25px;"><br />
<p>6. 7-8th September, 2014</p><br />
<p>#Permission to village office for Policy and Practice activity in Batu Dulang village, Batu Lanteh District and Gapit Village, Sumbawa Regency<br />
#Analysis the last laboratory result<br />
</p><br />
<p>7. 9-10th September, 2014</p><br />
<p>Policy and Practice activity in Batu Dulang village</p><br />
<p>8. 11-12th September, 2014</p><br />
<p>Policy and Practice activity in Gapit village</p><br />
<p>9. 13-15th September, 2014</p><br />
<p>Edit video Policy and Practice activity in Batu Dulang and Gapit village </p><br />
</p></ul><br />
<br />
<br />
<h3><b>Part 3 Taking Video about Societies Opinion about Biotechnology</b> </h3><br><br />
<br />
<ul style="padding-left:25px;"><br />
<p>10. 16th September, 2014</p><br />
<p>Permission to some government office for take video about the opinion of public policy officer and head of religion community in Sumbawa Regency</p><br />
<p>11. 17-19th September, 2014</p><br />
<p>Take video about the opinion of high public policy officer and head of religion community in Sumbawa Regency</p><br />
<p>12. 20-21th September, 2014</p><br />
<p>Edit video about the opinion of public policy officer and head of religion community in Sumbawa Regency</p><br />
</p></ul><br />
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http://2014.igem.org/File:Lab.jpg
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2014-10-18T03:34:51Z
<p>Adel: uploaded a new version of &quot;File:Lab.jpg&quot;</p>
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Adel
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2014-10-18T03:33:12Z
<p>Adel: </p>
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Adel
http://2014.igem.org/Team:Sumbawagen/overviews/Econey
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2014-10-18T03:30:45Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>SUMBAWAGEN PROJECT OVERVIEW</h2><br />
<h3>Background</h3><br />
<br />
<p>Biotechnology Faculty, Sumbawa University of Technology (SUT) was a new university, establish at May 2013 in Sumbawa Regency of West Nusa Tenggara Province of Indonesia. An Island located in east part of Indonesia. <br></p><br />
<br />
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<br />
<p>As the impact of the geographical position according to the capital city of Indonesia which located Java island, Sumbawa has limited accessibility to the economic and education sector. There are only two university in Sumbawa, and the SUT in the one and the only technology university in Sumbawa which has Biotechnology Faculty as their prime major, especially in eastern part of Indonesia.<br></p><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/2b/UTS_Campus.jpg campus 1"></center><br><br />
<center><strong>Picture 1. Campus view of Sumbawa University of Tecnology, the “Orange Building”</strong><br><br></center><br />
<br />
<center><img src="https://static.igem.org/mediawiki/2014/2/20/UTS_Campus_2.jpg"></center><br><br />
<center><strong>Picture 2. Campus view of Sumbawa University of Tecnology, the “Blue Building”</strong><br><br></center><br />
<br />
<p>Which these such condition Sumbawagen team started their project from the very fresh beginning. The basic reason that motivated the Sumbawagen team is back to their first survey on introducing synthetic biology to Sumbawa society at 2013. Based on the survey results we had conclusion that most of Sumbawa people are already in touch with various of biotechnology product in their daily life. On the other side this fact does not goes in the same direction with the understanding of Sumbawa society to the biotechnology products. This condition lead our team to go deeper into the topic which involves synthetic biology and the development prospect among Sumbawa society.<br></p><br />
<br />
<p>Eventhough Sumbawagen team consist of sophomore and in shortage condition to the facility and accessibility such as, inspite of using the refrigerated microsentrifuge for the transformation process we using minicentrifuge inside the refrigerator. We also created home made shaker for our cultivation project. Those condition did not made our team lost the spirit to participated in the prestigious competition of IGEM (International Genetically Enginereed Machine) 2014. We encourage ourself to overcome the obstacle in front of us, like the quote from our Rector “ When there is a will, there is a way”. </p><br />
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<br />
<br />
<h3>Project Idea</h3><br />
<p>Sumbawagen 2014 project is ECONEY (E. coli for Honey). ECONEY is engineered E.coli that capable to measure different level of glucose consentration in honey. BBa_J04450 has a lac promoter and mRFP gene, thus E. coli harboring this plasmid will give red color when the bacteria were grown with lactose or IPTG. Oppositely, the expression of mRFP will be decreased when glucose was present in medium through a mechanism called catabolite repression. We use this phenomenon to measure the concentration of glucose in honey by calibrating the color of E. coli medium using android-based mobile phone. Creation of novel circuit which consists of constitutive promoter followed by either Adenylate Cyclase or IIAGlc genes, which placed downward lac-mRFP circuit, may affect the sensitivity of catabolite repression. We expect an engineered E. coli, which shows red color expression in different range of glucose concentration. By changing mRFP gene with amilCP, a blue fluorescent protein, we may obtain E. coli expressing either red or blue color when different honey added to the medium due to different glucose concentration.<br></p><br />
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Adel
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>A. Team Discussion</h2><br />
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<p>Policy and practices is a way for us to introduce our project to society. We discussed our policy and practices project first on June 23, 2014. The points of our discussion were determining our survey targets and making questionnaire design. After we shared our best ideas for the project, we decided the purpose of the project which is to introduce ECONEY to Sumbawa society and to discover more information about everything related to Sumbawa honey. Because of our questionnaire design was still not complete, we had another disscussions then we report to our instructors to get the revisions. The problems were about the question complexity and its relation to the project. And finally, our questionnaire finished on July 6th.<br></p><br />
<center><img src="https://static.igem.org/mediawiki/2014/4/40/Team_Disscustion.jpg"></center><br><br />
<br />
<p>The next discussion was about the respondents. We determined the region target and the number of the respondents. We devided the respondents in honey production regions. We divided the respondents into producers, distributors, and consumers. We targetted to have 630 respondents which consist of 210 producers, 210 distributors, and 210 consumers using random sampling design.<br></p><br />
<br />
<h2>B. Idea</h2><br />
<br />
We surveyed 16 regions consist of 7 producer regions and 9 consumer regions, while the distributors were exist in both of the areas. The producer regions located in villages. On the other hand, we choose Sumbawa Besar (the capital city of Sumbawa regency) as consumer regions. Sumbawagen team then divided into 7 teams to do the survey. Each team did the survey at 1 producer regions and 1 consumer regions. The survey was held on July 14-17 2014.<br><br />
<br />
<br />
<h2>C. Data Analysis</h2><br />
We would like to give our gratitude to all Sumbawa society that had been participated in our policy and practices project. There are 672 respondents which consist of 130 producers, 71 distributors, and 471 consumers participated in the project. The data analysis were given below:<br><br><br />
<br />
<strong><i>1. Have you ever consume Sumbawa honey?</i></strong><br><br />
Most of our respondents have consumed Sumbawa honey. It shows that Sumbawa honey is a very important commodity for Sumbawa society.<br><br />
<br />
<center><img src="https://static.igem.org/mediawiki/2014/9/94/Diagram1.jpg"></center><br><br />
<br />
<strong><i>2. Is there any tools which can be used to measure glucose level in Sumbawa honey?</i></strong><br />
<p>Many of our respondents said they don’t know the tools to measure the glucose level in Sumbawa honey.</P><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/0c/Diagram_2.jpg"></center><br><br />
<br />
<strong><i>3. If the answer for no. 2 is YES, please mention the tools</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/24/Diagram_3.jpg"></center><br><br />
<p>None of our respondents could mention the tool used to measure glucose level in Sumbawa honey.<br> <br />
Although some of our respondents said that there is a tool exist to measure glucose in Sumbawa honey, they couldn’t mention the name. After interviewed them little bit deeper we figured out that society has perception that the tool is the same with the tool used to measure water level, but the fact it's different. If they want to know glucose level in Sumbawa honey (especially the distributor) they have to send honey sample to Mataram (the capital city of West Nusa Tenggara province).<br></p><br />
<br />
<strong><i>4. If there is a tool created to measure the glucose level in honey, how would you respond?</i></strong><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/2b/Diagram4.jpg"></center><br><br />
<p>The society gave the positive response for this question. Most of producers want to use (58%), and just 7% didn’t care about it. The distributor respondents who want to learn and use the tool are almost same (47% and 46%). Meanwhile, most of the consumers want to learn (53%).<br></p><br />
<br />
<br />
<strong><i>5. Are you android smartphone user?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/b/ba/Diagram_5.jpg"></center><br><br />
<p>In Sumbawa society, android smartphone is familiar among teens. Hence in the survey, only a small number of respondents using android smartphone: producer is only 29%, distributor is only 15%, and the consumer is only 30%. <br></p><br />
<br />
<strong><i>6. If the answer of question number 5 is “YES”, are you using android smartphone application?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/03/Diagram_6.jpg"></center><br><br />
<p>Most of android user use android aplication beacuse there are various and cheap applications in android market, even some are distributed for free.<br></p><br />
<br />
<strong><i>7. If the answer is “YES” for question number 6, is android smartphone application easy to use?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/7/7e/Diagram_7.jpg"></center><br><br />
<p>All of our producer respondents agree if android smartphone applications are easy to use. Whereas 40% from our distributor respondents and 5% from consumer respondents said that the applications are difficult to use.<br></p><br />
<br />
<strong><i>8. If the application can meassure glucose level in honey, what is your response?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/5/57/8_diagram.jpg"></center><br><br />
<p>Our respondents were very interest with our project especially for producer and distributor category as show from the survey result.<br> </p><br />
<br />
<strong><i>9. As sumbawa people, are you proud of Sumbawa honey?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/6/6b/Diagram_9.jpg"></center><br><br />
<p>We can conclude that Sumbawa people are absolutely proud of Sumbawa honey. Sumbawa honey is the icon of Sumbawa.</p><br />
<br />
<h2>Conclusion</h2><br />
<p>Sumbawa honey as the icon of Sumbawa contains high glucose level. However, there is no tools for measuring glucose concentration exist in Sumbawa. That's why people of Sumbawa they really appreciate our ECONEY project. We hope our ECONEY project could be applied to help people in Sumbawa measure glucose level in honey.</p><br />
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<h2>SUMBAWAGEN PROJECT OVERVIEW</h2><br />
<h3>Background</h3><br />
<br />
<p>Biotechnology Faculty, Sumbawa University of Technology (SUT) was a new university, establish at May 2013 in Sumbawa Regency of West Nusa Tenggara Province of Indonesia. An Island located in east part of Indonesia. <br></p><br />
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<center><img src="https://static.igem.org/mediawiki/2014/4/4d/Biotek_camp_1.png"></center><br><br />
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<p>As the impact of the geographical position according to the capital city of Indonesia which located Java island, Sumbawa has limited accessibility to the economic and education sector. There are only two university in Sumbawa, and the SUT in the one and the only technology university in Sumbawa which has Biotechnology Faculty as their prime major, especially in eastern part of Indonesia.<br></p><br />
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<p>Which these such condition Sumbawagen team started their project from the very fresh beginning. The basic reason that motivated the Sumbawagen team is back to their first survey on introducing synthetic biology to Sumbawa society at 2013. Based on the survey results we had conclusion that most of Sumbawa people are already in touch with various of biotechnology product in their daily life. On the other side this fact does not goes in the same direction with the understanding of Sumbawa society to the biotechnology products. This condition lead our team to go deeper into the topic which involves synthetic biology and the development prospect among Sumbawa society.<br></p><br />
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<p>Eventhough Sumbawagen team consist of sophomore and in shortage condition to the facility and accessibility such as, inspite of using the refrigerated microsentrifuge for the transformation process we using minicentrifuge inside the refrigerator. We also created home made shaker for our cultivation project. Those condition did not made our team lost the spirit to participated in the prestigious competition of IGEM (International Genetically Enginereed Machine) 2014. We encourage ourself to overcome the obstacle in front of us, like the quote from our Rector “ When there is a will, there is a way”. </p><br />
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<h3>Project Idea</h3><br />
<p>Sumbawagen 2014 project is ECONEY (E. coli for Honey). ECONEY is engineered E.coli that capable to measure different level of glucose consentration in honey. BBa_J04450 has a lac promoter and mRFP gene, thus E. coli harboring this plasmid will give red color when the bacteria were grown with lactose or IPTG. Oppositely, the expression of mRFP will be decreased when glucose was present in medium through a mechanism called catabolite repression. We use this phenomenon to measure the concentration of glucose in honey by calibrating the color of E. coli medium using android-based mobile phone. Creation of novel circuit which consists of constitutive promoter followed by either Adenylate Cyclase or IIAGlc genes, which placed downward lac-mRFP circuit, may affect the sensitivity of catabolite repression. We expect an engineered E. coli, which shows red color expression in different range of glucose concentration. By changing mRFP gene with amilCP, a blue fluorescent protein, we may obtain E. coli expressing either red or blue color when different honey added to the medium due to different glucose concentration.<br></p><br />
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<h2>SUMBAWAGEN PROJECT OVERVIEW</h2><br />
<h3>Background</h3><br />
<br />
<p>Biotechnology Faculty, Sumbawa University of Technology (SUT) was a new university, establish at May 2013 in Sumbawa Regency of West Nusa Tenggara Province of Indonesia. An Island located in east part of Indonesia. <br></p><br />
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<center><img src="https://static.igem.org/mediawiki/2014/4/4d/Biotek_camp_1.png"></center><br><br />
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<p>As the impact of the geographical position according to the capital city of Indonesia which located Java island, Sumbawa has limited accessibility to the economic and education sector. There are only two university in Sumbawa, and the SUT in the one and the only technology university in Sumbawa which has Biotechnology Faculty as their prime major, especially in eastern part of Indonesia.<br></p><br />
<center><img src="https://static.igem.org/mediawiki/2014/6/66/Biotech_camp-2.png"></center><br><br />
<br />
<p>Which these such condition Sumbawagen team started their project from the very fresh beginning. The basic reason that motivated the Sumbawagen team is back to their first survey on introducing synthetic biology to Sumbawa society at 2013. Based on the survey results we had conclusion that most of Sumbawa people are already in touch with various of biotechnology product in their daily life. On the other side this fact does not goes in the same direction with the understanding of Sumbawa society to the biotechnology products. This condition lead our team to go deeper into the topic which involves synthetic biology and the development prospect among Sumbawa society.<br></p><br />
<br />
<p>Eventhough Sumbawagen team consist of sophomore and in shortage condition to the facility and accessibility such as, inspite of using the refrigerated microsentrifuge for the transformation process we using minicentrifuge inside the refrigerator. We also created home made shaker for our cultivation project. Those condition did not made our team lost the spirit to participated in the prestigious competition of IGEM (International Genetically Enginereed Machine) 2014. We encourage ourself to overcome the obstacle in front of us, like the quote from our Rector “ When there is a will, there is a way”. </p><br />
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<h1>Project Idea</h1><br />
<p>Sumbawagen 2014 project is ECONEY (E. coli for Honey). ECONEY is engineered E.coli that capable to measure different level of glucose consentration in honey. BBa_J04450 has a lac promoter and mRFP gene, thus E. coli harboring this plasmid will give red color when the bacteria were grown with lactose or IPTG. Oppositely, the expression of mRFP will be decreased when glucose was present in medium through a mechanism called catabolite repression. We use this phenomenon to measure the concentration of glucose in honey by calibrating the color of E. coli medium using android-based mobile phone. Creation of novel circuit which consists of constitutive promoter followed by either Adenylate Cyclase or IIAGlc genes, which placed downward lac-mRFP circuit, may affect the sensitivity of catabolite repression. We expect an engineered E. coli, which shows red color expression in different range of glucose concentration. By changing mRFP gene with amilCP, a blue fluorescent protein, we may obtain E. coli expressing either red or blue color when different honey added to the medium due to different glucose concentration.<br></p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol12
Team:Sumbawagen/Notebook/protocol12
2014-10-18T02:09:09Z
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<h2>Notebook – Protocol – 12. Honey glucose meter measurement</h2><br />
<ul style="padding-left:25px;"><br />
<p>1. Glucose dissolved in distilled water to a concentrations of 100 mM was used as stock. From that stock, various concentration of glucose were prepared by dilution. The concentrations were 5 / 10 / 15 / 20 / 25 mM.</p><br />
<p>2. Those glucose stocks were measured its mg/dl value by a glucose meter. The values then plotted as y axis, against the concentration as x axis. Linear standard curve was obtained and the formula was used to calculate the concentration of unknown sample.</p><br />
<p>3. Honey in 1.5 ml polypropylene tube was warmed at 60 oC for 10 minutes, then transferred 100 ul to a new tube which contained 900 ul distilled water to obtain 1/10 dilution.</p><br />
<p>4. This sample was used for measurement of glucose concentration by glucose meter, and its glucose concentration was calculated using standard curve obtained before.</p><br />
<p>5. When the mg/dl value still exceeded the linear curve, dilution rate was increased.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol13
Team:Sumbawagen/Notebook/protocol13
2014-10-18T02:07:36Z
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>Notebook – Protocol – 13. SDS-PAGE</h2><br />
<ul style="padding-left:25px;"><br />
<p>1. 1 ml of E. coli BL21(DE3) overnight culture was added to 1.5 ml propylene tube. Then centrifuge at 6,000 rpm, 5 minutes at room temperature. Then supernatant was discarded.</p><br />
<p>2. Pellet was added with 300 ul of 1x Bug Buster (Novagen). Tube was vortexed.</p><br />
<p>3. Tube was put on boiled water for 5 minutes.</p><br />
<p>4. 50 ul of samples was aliquoted and used as “total protein” sample.</p><br />
<p>5. The rest of the sample in the tube was centrifuged at 6,000 rpm, 10 min, room temperature.</p><br />
<p>6. 50 ul of supernatant was aliquoted and used as “soluble protein’ sample.</p><br />
<p>7. Protein concentration of “total protein” and “soluble protein” were estimated using Lowry method based on the kit from Biorad. We have no UV spectrophotometer for measurement of real concentration at 750 nm. But according to our chief instructor Dr. Arief previous experiment, the concentration of “soluble protein” will be about 1,000 – 2,000 ug/ml, while the concentration of “total protein” will be about 10,000 – 20,000 ug/ml.</p><br />
<p>8. “Soluble protein” sample was used directly, while “total protein” sample was diluted 1/10th to obtain samples with roughly the same concentration for SDS-PAGE.</p><br />
<p>9. 15 ul sample was added with 15 ul dye containing SDS and Coomassie Brilliant Blue (CBB) in 500 ul polypropylene tube, then the tube was put in boiled water for 15 min</p><br />
<p>10. 15 ul of sample – protein quantity about 10 ug – was electrophoresed. Marker used was Blue Star Prestained Protein Marker (Nippon Genetics). The marker has proteins bands of 170 / 125 / 93 / 68 / 53 / 41 / 32 / 23 / 14 / 10 kD.</p><br />
<p>11. SDSPAGE was done with commercial 10-20% SDS gel, and 1x SDS buffer. 10x SDS buffer was prepared using 250 mM Tris, 1,920 mM Glycine, and 1% w/v SDS.</p><br />
<p>12. SDSPAGE was done for 45 minutes with equipment setting of 40 mA.</p><br />
<p>13. Staining of SDSPAGE gel was done using Rapid CBB kit (Kanto Chemicals). Staining was done 1-3 hours.</p><br />
<p>14. Destaining of SDSPAGE gel was done using solution containing 75 ml distilled water, 10 ml acetic acid, 15 ml ethanol. Destaining was done overnight.</p><br />
<p>15. Gel was documented with pocket camera, and file was evaluated with Photoshop program in a computer.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol13
Team:Sumbawagen/Notebook/protocol13
2014-10-18T02:05:09Z
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<h2>Notebook – Protocol – 13. SDS-PAGE</h2><br />
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<p>1. 1 ml of E. coli BL21(DE3) overnight culture was added to 1.5 ml propylene tube. Then centrifuge at 6,000 rpm, 5 minutes at room temperature. Then supernatant was discarded.</p><br />
<p>2. Pellet was added with 300 ul of 1x Bug Buster (Novagen). Tube was vortexed.</p><br />
<p>3. Tube was put on boiled water for 5 minutes.</p><br />
<p>4. 50 ul of samples was aliquoted and used as “total protein” sample.</p><br />
<p>5. The rest of the sample in the tube was centrifuged at 6,000 rpm, 10 min, room temperature.</p><br />
<p>6. 50 ul of supernatant was aliquoted and used as “soluble protein’ sample.</p><br />
<p>7. Protein concentrations of “total protein” and “soluble protein” were estimated using Lowry method based on the kit from Biorad. We have no UV spectrophotometer for measurement of real concentration at 750 nm. But according to our chief instructor Dr. Arief previous experiment, the concentration of “soluble protein” will be about 1,000 – 2,000 ug/ml, while the concentration of “total protein” will be about 10,000 – 20,000 ug/ml.</p><br />
<p>8. “Soluble protein” sample was used directly, while “total protein” sample was diluted 1/10th to obtain samples with roughly the same concentration for SDS-PAGE.</p><br />
<p>9. 15 ul sample was added with 15 ul dye containing SDS and Coomassie Brilliant Blue (CBB) in 500 ul polypropylene tube, then the tube was put in boiled water for 15 min</p><br />
<p>10. 15 ul of sample – protein quantity about 10 ug – was electrophoresed. Marker used was Blue Star Prestained Protein Marker (Nippon Genetics). The marker has proteins bands of 170 / 125 / 93 / 68 / 53 / 41 / 32 / 23 / 14 / 10 kD.</p><br />
<p>11. SDSPAGE was done with commercial 10-20% SDS gel, and 1x SDS buffer. 10x SDS buffer was prepared using 250 mM Tris, 1,920 mM Glycine, and 1% w/v SDS.</p><br />
<p>12. SDSPAGE was done for 45 minutes with equipment setting of 40 mA.</p><br />
<p>13. Staining of SDSPAGE gel was done using Rapid CBB kit (Kanto Chemicals). Staining was done 1-3 hours.</p><br />
<p>14. Destaining of SDSPAGE gel was done using solution containing 75 ml distilled water, 10 ml acetic acid, 15 ml ethanol. Destaining was done overnight.</p><br />
<p>15. Gel was documented with pocket camera, and file was evaluated with Photoshop program in a computer.</p><br />
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http://2014.igem.org/Team:Sumbawagen/Notebook/protocol14
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2014-10-18T02:04:29Z
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<h2>Notebook – Protocol – 14. Measurement</h2><br />
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<p>1. Overnight culture of “main culture” was taken from the shaker, then put in rack for measurement.</p><br />
<p>2. First, picture was taken with pocket camera.</p><br />
<p>3. Then color of the medium was measured its RGB value by ColorMeter program from an android-base mobile phone. For each side, 3 pictures were shot.</p><br />
<p>4. For validation, another mobile phone with different resolution of camera was used. Also besides, ColorMeter, we also used different program for RGB measurement namely Color Detector.</p><br />
<p>5. Values of RGB were converted to luminance value to obtain only the brightness of color (http://www.scantips.com/lumin.html; accessed on Sep. 24th, 2014). The formula used is:<br />
Luminance value = (R x 0.30) + (G x 0.59) + (B x 0.11)<br />
</p><br />
<p>6. Luminance values from 3 RGB data were averaged and calculated its standard deviation.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol14
Team:Sumbawagen/Notebook/protocol14
2014-10-18T02:03:20Z
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<h2>Notebook – Protocol – 14. Measurement</h2><br />
<ul style="padding-left:25px;"><br />
<p>1. Overnight culture of “main culture” was taken from the shaker, then put in rack for measurement.</p><br />
<p>2. First, picture was taken with pocket camera.</p><br />
<p>3. Then color of the medium was measured its RGB value by ColorMeter program from an android-base mobile phone. For each side, 3 pictures were shot.</p><br />
<p>4. For validation, an other mobile phone with different resolution of camera was used. Also besides, ColorMeter, we also used different program for RGB measurement namely Color Detector.</p><br />
<p>5. Values of RGB were converted to luminance value to obtain only the brightness of color (http://www.scantips.com/lumin.html; accessed on Sep. 24th, 2014). The formula used is:<br />
Luminance value = (R x 0.30) + (G x 0.59) + (B x 0.11)<br />
</p><br />
<p>6. Luminance values from 3 RGB data were averaged and calculated its standard deviation.</p><br />
</p></ul><br />
<br />
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<h2>Notebook – Protocol – 14. Measurement</h2><br />
<p><u>DNA Purification from Liquid Samples</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Overnight culture of “main culture” was taken from the shaker, then put in rack for measurement.</p><br />
<p>2. First, picture was taken with pocket camera.</p><br />
<p>3. Then color of the medium was measured its RGB value by ColorMeter program from an android-base mobile phone. For each side, 3 pictures were shot.</p><br />
<p>4. For validation, an other mobile phone with different resolution of camera was used. Also besides, ColorMeter, we also used different program for RGB measurement namely Color Detector.</p><br />
<p>5. Values of RGB were converted to luminance value to obtain only the brightness of color (http://www.scantips.com/lumin.html; accessed on Sep. 24th, 2014). The formula used is:<br />
Luminance value = (R x 0.30) + (G x 0.59) + (B x 0.11)<br />
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<p>6. Luminance values from 3 RGB data were averaged and calculated its standard deviation.</p><br />
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<h2>Notebook – Protocol – 13. SDS-PAGE</h2><br />
<ul style="padding-left:25px;"><br />
<p>1. 1 ml of E. coli BL21(DE3) overnight culture was added to 1.5 ml propylene tube. Then centrifuge at 6,000 rpm, 5 minutes at room temperature. Then supernatant was discarded.</p><br />
<p>2. Pellet was added with 300 ul of 1x Bug Buster (Novagen). Tube was vortexed.</p><br />
<p>3. Tube was put on boiled water for 5 minutes.</p><br />
<p>4. 50 ul of samples was aliquoted and used as “total protein” sample.</p><br />
<p>5. The rest of the sample in the tube was centrifuged at 6,000 rpm, 10 min, room temperature.</p><br />
<p>6. 50 ul of supernatant was aliquoted and used as “soluble protein’ sample.</p><br />
<p>7. Protein concentration of “total protein” and “soluble protein” were estimated using Lowry method based on the kit from Biorad. We have no UV spectrophotometer for measurement of real concentration at 750 nm. But according to our chief instructor Dr. Arief previous experiment, the concentration of “soluble protein” will be about 1,000 – 2,000 ug/ml, while the concentration of “total protein” will be about 10,000 – 20,000 ug/ml.</p><br />
<p>8. “Soluble protein” sample was used directly, while “total protein” sample was diluted 1/10th to obtain samples with roughly the same concentration for SDS-PAGE.</p><br />
<p>9. 15 ul sample was added with 15 ul dye containing SDS and Coomassie Brilliant Blue (CBB) in 500 ul polypropylene tube, then the tube was put in boiled water for 15 min</p><br />
<p>10. 15 ul of sample – protein quantity about 10 ug – was electrophoresed. Marker used was Blue Star Prestained Protein Marker (Nippon Genetics). The marker has proteins bands of 170 / 125 / 93 / 68 / 53 / 41 / 32 / 23 / 14 / 10 kD.</p><br />
<p>11. SDSPAGE was done with commercial 10-20% SDS gel, and 1x SDS buffer. 10x SDS buffer was prepared using 250 mM Tris, 1,920 mM Glycine, and 1% w/v SDS.</p><br />
<p>12. SDSPAGE was done for 45 minutes with equipment setting of 40 mA.</p><br />
<p>13. Staining of SDSPAGE gel was done using Rapid CBB kit (Kanto Chemicals). Staining was done 1-3 hours.</p><br />
<p>14. Destaining of SDSPAGE gel was done using solution containing 75 ml distilled water, 10 ml acetic acid, 15 ml ethanol. Destaining was done overnight.</p><br />
<p>15. Gel was documented with pocket camera, and file was evaluated with Photoshop program in a computer.</p><br />
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<h2>Notebook – Protocol – 12. Honey glucose meter measurement</h2><br />
<ul style="padding-left:25px;"><br />
<p>1. Glucose dissolved in distilled water to a concentration of 100 mM was used as stock. From that stock, various concentration of glucose were prepared by dilution. The concentrations were 5 / 10 / 15 / 20 / 25 mM.</p><br />
<p>2. Those glucose stocks were measured its mg/dl value by a glucose meter. The values then plotted as y axis, against the concentration as x axis. Linear standard curve was obtained and the formula was used to calculate the concentration of unknown sample.</p><br />
<p>3. Honey in 1.5 ml polypropylene tube was warmed at 60 oC for 10 minutes, then transferred 100 ul to a new tube which contained 900 ul distilled water to obtain 1/10 dilution.</p><br />
<p>4. This sample was used for measurement of glucose concentration by glucose meter, and its glucose concentration was calculated using standard curve obtained before.</p><br />
<p>5. When the mg/dl value still exceeded the linear curve, dilution rate was increased.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol4
Team:Sumbawagen/Notebook/protocol4
2014-10-18T01:51:51Z
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<h2>Notebook – Protocol – 4. DNA Purification</h2><br />
<p>Spin column based purification was used. The kit is Gel/PCR extraction kit (Nippon Genetics). Protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge. All steps were done at room temperature.</p><br />
<p><u>DNA Purification from Liquid Samples</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Into a clean, sterilized 0.5 ul polypropylene tube, add PCR products : buffer GP1 = 1 : 5 (e.g. 40 ul : 200 ul). Vortexing.</p><br />
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.</p><br />
<p>5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube.</p><br />
<p>6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>7. Obtain the DNA as the flowthrough.</p><br />
</p></ul><br />
<br />
<p><u>DNA Purification from Agarose Gel</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Into a clean, sterlized 1.5 ul polypropylene tube, add cutted gel containing DNA of interest : buffer GP1 = < 300 mg : 500 ul. Vortexing. Incubate at 55 oC; 10-15 minutes. Invert the tube.</p><br />
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.</p><br />
<p>5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube</p><br />
<p>6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>7. Obtain the DNA as the flowthrough.</p><br />
</p></ul><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol4
Team:Sumbawagen/Notebook/protocol4
2014-10-18T01:50:50Z
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<h2>Notebook – Protocol – 4. DNA Purification</h2><br />
<p>Spin column based purification was used. The kit is Gel/PCR extraction kit (Nippon Genetics). Protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge. All steps were done at room temperature.</p><br />
<p><u>DNA Purification from Liquid Samples</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Into a clean, sterillized 0.5 ul polypropylene tube, add PCR products : buffer GP1 = 1 : 5 (e.g. 40 ul : 200 ul). Vortexing.</p><br />
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.</p><br />
<p>5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube.</p><br />
<p>6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>7. Obtain the DNA as the flowthrough.</p><br />
</p></ul><br />
<br />
<p><u>DNA Purification from Agarose Gel</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Into a clean, sterlized 1.5 ul polypropylene tube, add cutted gel containing DNA of interest : buffer GP1 = < 300 mg : 500 ul. Vortexing. Incubate at 55 oC; 10-15 minutes. Invert the tube.</p><br />
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.</p><br />
<p>5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube</p><br />
<p>6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>7. Obtain the DNA as the flowthrough.</p><br />
</p></ul><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol3
Team:Sumbawagen/Notebook/protocol3
2014-10-18T01:49:47Z
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<h2>Notebook – Protocol – 3. Electrophoresis and DNA visualization</h2><br />
<br />
<br />
<p>1.50x TAE (Thermoscientific) was diluted to 1x TAE (1 ml 50x TAE + 49 ml distilled water).</p><br />
<p>2. 3 g of Agarose S (Nippongene) was added into 200 ml of 1x TAE to get 1.5 % agarose gel in a glass flask.</p> <br />
<p>3. Agarose was melted by putting the glass flask in boiled water – we have no microwave – and sometimes stirred slowly to avoid bubble formation with stirrer bar on a stirrer.</p><br />
<p>4. Completely melted agarose solution was poured into the casting tray with comb.</p><br />
<p>5. Let the gel became solid by incubating at room temperature for 30 minutes.</p><br />
<p>6. Electrophoresis was done in 1x TAE buffer in an electrophoresis tank (Mupid)</p> <br />
<p>7. The volume of samples used for electrophoresis was 7 ul for 1 kB DNA ladder and 10 ul (PCR product), or 15 ul (Restriction enzyme digestion product) for DNA samples for each well.</p><br />
<p>8. Electrophoresis was done for 20 minutes using 135V setting.</p><br />
<p>9. After electrophoresis, gel was put into 1x TAE solution containing Sybrsafe to stain the DNA for about 20 minutes.</p><br />
<p>10. Then gel was placed on trans illuminator for viewing and documentation using pocket digital camera.</p> <br />
<p>11. The file was viewed with computer using Photoshop.</p><br />
<p>12. For disposal, gel and buffer were dried then burnt together with any tissues or papers in contact.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Human_practice/survey
Team:Sumbawagen/Human practice/survey
2014-10-18T01:47:31Z
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<h2>A. Team Discussion</h2><br />
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<p>Policy and practices is a way for us to introduce our project to society. We discussed our policy and practices project first on June 23 2014. The points of our discussion were determined our survey targets and making questionnaire design. After we shared our best ideas for the project, we decided the purpose of the project is to introduce our ECONEY project to society in Sumbawa and discover more information about everything related to Sumbawa honey. Because of our questionnaire design was still not complete, we had another disscussions then we report to our instructors to get the revisions. The problems were about the question complexity and the relation to the project. And finally, our questionnaire finished on July 6th.<br></p><br />
<center><img src="https://static.igem.org/mediawiki/2014/4/40/Team_Disscustion.jpg"></center><br><br />
<br />
<p>The next was discussion about the respondents. We discussed about the region target and the amount of the people. Since our project was related to honey, we focused to the respondents in honey production regions. We divided them into producers, distributors, and consumers. We targetted to have 630 respondents which consist of 210 producers, 210 distributors, and 210 consumers.<br></p><br />
<br />
<h2>B. Idea</h2><br />
<br />
We did the survey by visiting respondent’s house and interview them randomly. Our region targets were divided into 16 regions consist of 7 producer regions and 9 consumer regions, while the distributors were exist in both of the areas. The producer regions located in villages. On the other hand, we choose Sumbawa Besar (the capital city of Sumbawa regency) as consumer regions. Sumbawagen team then divided into 7 teams to do the survey. Each team did the survey at 1 producer regions and 1 consumer regions. The survey was held on July 14-17 2014.<br><br />
<br />
<br />
<h2>C. Data Analysis</h2><br />
We would like to give our gratitude to all Sumbawa society that had been participated in our policy and practices project. There are 672 respondents which consist of 130 producers, 71 distributors, and 471 consumers participated in the project. The data analysis were given below:<br><br><br />
<br />
<strong><i>1. Have you ever consume sumbawa honey?</i></strong><br><br />
Most of our respondents have consumed Sumbawa honey. It shows that Sumbawa honey is a very important commodity for Sumbawa society.<br><br />
<br />
<center><img src="https://static.igem.org/mediawiki/2014/9/94/Diagram1.jpg"></center><br><br />
<br />
<strong><i>2. Is there any tools which can be used to measure glucose level in Sumbawa honey?</i></strong><br />
<p>Many of our respondents said they don’t know the tools to measure the glucose level in Sumbawa honey.</P><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/0c/Diagram_2.jpg"></center><br><br />
<br />
<strong><i>3. If the answerfor no. 2 is A, please mention the tools</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/24/Diagram_3.jpg"></center><br><br />
<p>From our survey activity no one from our respondents could mention the name of the tool to measure glucose level in Sumbawa honey.<br> <br />
Although most of our respondents said if they know the tools to measure glucose in Sumbawa honey, they didn’t mention the tool’s name. Then we conclude that nobody knows and ever use the tools to measure glucose level in honey. The society thought if the tool to measure glucose level is same with the tool to measure water level, but in fact it is different. Sumbawa society are familiar with tools for measuring water level. If they want to know glucose level in Sumbawa honey (especially for distributor) they should deliver honey sample to Mataram (capital city of West Nusa Tenggara province).<br></p><br />
<br />
<strong><i>4. If there is a tool created to measure the glucose level in honey, how would you respond?</i></strong><br />
<center><img src="https://static.igem.org/mediawiki/2014/2/2b/Diagram4.jpg"></center><br><br />
<p>The society gave the positive response for this question. Most of producers want to use (58%), and just 7% didn’t care about it. The distributor respondents who want to learn and use the tool are almost same (47% and 46%). Meanwhile, most of the consumers want to learn (53%).<br></p><br />
<br />
<br />
<strong><i>5. Are you android smartphone user?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/b/ba/Diagram_5.jpg"></center><br><br />
<p>In Sumbawa society, android smartphone is familiar in among teens. Hence in the survey, only a small of our respondents using android smartphone: producer is only 29%, distributor is only 15%, and the consumer is only 30%. <br></p><br />
<br />
<strong><i>6. If the answer of question number is “a”, are you using android smartphone application?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/0/03/Diagram_6.jpg"></center><br><br />
<p>Almost all android user using in android smartphone. No wonder if they use it, because there are various and cheap applications in android smartphone, even there are many applications for free.<br></p><br />
<br />
<strong><i>7. If the answer is “a” for number 6, what’s your opinion when using android smartphone application?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/7/7e/Diagram_7.jpg"></center><br><br />
<p>All of our producer respondents agree if android smartphone applications are easy to use. Whereas 45% from our distributor respondents and 5% from consumer respondents said that the applications are difficult to use.<br></p><br />
<br />
<strong><i>8. If the application can meassure glucose level in honey, what is your response?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/5/57/8_diagram.jpg"></center><br><br />
<p>Our respondents were very interest with our project especially for producer and distributor category. It is proved in the survey result that 100% answer if they want to use the application.<br> </p><br />
<br />
<strong><i>9. As sumbawa people, are you proud of Sumbawa honey?</i></strong><br><br />
<center><img src="https://static.igem.org/mediawiki/2014/6/6b/Diagram_9.jpg"></center><br><br />
<p>We can conclude that Sumbawa people are absolutely proud of Sumbawa honey. Sumbawa honey is an icon of Sumbawa. Only 1 % of respondent (consumer category) doesn’t feel proud with Sumbawa honey.</p><br />
<br />
<h2>Conclusion</h2><br />
<p>Sumbawa honey has been the icon of Sumbawa. Sumbawa honey contains high glucose level. However most of them have not known the tools for measuring glucose concentration level yet. Thus, they really appreciate our ECONEY project. We hope our ECONEY project could be applied to help people in Sumbawa measure glucose level in honey.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol5
Team:Sumbawagen/Notebook/protocol5
2014-10-18T01:35:03Z
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<h2>Notebook – Protocol – 5. Restriction enzymes digestion</h2><br />
<p>1. Restriction enzymes digestion was done using Fast Digest product (Thermoscientific) using the following composition:</p><br />
<ul style="padding-left:25px;"><br />
<li>DNA sample (1-3 ug), 15 ul</li><br />
<li>Fast Digest restriction enzyme #1, 2 ul</li><br />
<li>Fast Digest restriction enzyme #2, 2 ul</li><br />
<li>10x buffer, 3 ul</li><br />
<li>Sterilized distilled water, 6 ul</li><br />
Total volume, 30 ul</li><br />
</p></ul><br />
<br />
<p>2. Samples were incubated at 37 °C for more than 1 hour (typically 3 hours).</p><br />
<p>3. When necessary, samples were electrophoresed using protocol “3. Electrophoresis”(typically 3 hours).</p><br />
<p>4. Samples were purified using protocol “4. DNA purification”.</p><br />
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<h2>Notebook – Protocol – 4. DNA Purification</h2><br />
<p>Spin column based purification was used. The kit is Gel/PCR extraction kit (Nippon Genetics). Protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge. All steps were done at room temperature.</p><br />
<p><u>DNA Purification from Liquid Samples</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Into a clean, sterlized 0.5 ul polypropylene tube, add PCR products : buffer GP1 = 1 : 5 (e.g. 40 ul : 200 ul). Vortexing.</p><br />
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.</p><br />
<p>5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube.</p><br />
<p>6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>7. Obtain the DNA as the flowthrough.</p><br />
</p></ul><br />
<br />
<p><u>DNA Purification from Agarose Gel</u></p><br />
<ul style="padding-left:25px;"><br />
<p>1. Into a clean, sterlized 1.5 ul polypropylene tube, add cutted gel containing DNA of interest : buffer GP1 = < 300 mg : 500 ul. Vortexing. Incubate at 55 oC; 10-15 minutes. Invert the tube.</p><br />
<p>2. Load the samples onto the spin column equipped with collection tube. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>3. Add 600 ul of GP2. Centrifuge for 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>4. Centrifuge for 6,000 rpm; 5 minutes to dry the column.</p><br />
<p>5. Replace collecting tube with clean, sterilized 1.5 ml size polypropylene tube</p><br />
<p>6. Add 30 ul of GP3. Incubate 2 minutes at room temperature. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>7. Obtain the DNA as the flowthrough.</p><br />
</p></ul><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol7
Team:Sumbawagen/Notebook/protocol7
2014-10-18T01:07:54Z
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<h2>Notebook – Protocol – 7. Ligation</h2><br />
<p>1. Ligation reaction was as follow:</p><br />
<ul style="padding-left:25px;"><br />
<li>Insert DNA, 2,5 ul*</li><br />
<li>Plasmid DNA, 1.5 ul*</li><br />
<li>Ligation kit (Takara), 5 ul</li><br />
Total volume, 10 ul</li><br><br />
* final quantity insert DNA > plasmid DNA </li><br />
</ul><br />
<br />
<p>2. Samples were incubated at 4 oC for overnight (inside a refrigerator).</p><br />
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<h2>Notebook – Protocol – 7. Ligation</h2><br />
<p>1. Ligation reaction was as follow:</p><br />
<ul style="padding-left:25px;"><br />
<li>Insert DNA, 2,5 ul*</li><br />
<li>Plasmid DNA, 1.5 ul*</li><br />
<li>Ligation kit (Takara), 5 ul</li><br />
Total volume, 10 ul</li><br />
* final quantity insert DNA > plasmid DNA <br />
</p></ul><br />
<br />
<p>2. Samples were incubated at 4 oC for overnight (inside a refrigerator).</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol6
Team:Sumbawagen/Notebook/protocol6
2014-10-18T01:01:17Z
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<h2>Notebook – Protocol – 6. Plasmid purification</h2><br />
<p>Spin column based purification was used. The kit is Plasmid mini kit (Nippon Genetics). Standard protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge.</p><br />
<br />
<ul><br />
<p>1. Add 2 ml of E. coli overnight culture into 2 ml polypropylene tube. Centrifuge for 6,000 rpm; 5 minutes. Remove the supernatant.</><br />
<p>2. Add 200 ul of mP1; vortexing. Add 200 ul of mP2; invert the tube 10 times. Incubate at 2 minutes at room temperature. Add 300 ul of mP3; invert the tube. </p><br />
<p>3. Immediately centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>4. Load the supernatant onto spin column. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>5. Add 400 ul of mP4. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>6. Add 600 ul of mP5. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.</p><br />
<p>7. Centrifuge 6,000 rpm; 5 minute to dry the column.</p><br />
<p>8. Replace collecting tube with clean, sterlized 1,5 ml size polypropylene tube.</p><br />
<p>9. Add 50 ul of mP6. Incubate at room temperature for 2 minutes. Centrifuge for 6,000 rpm; 5 minutes.</p><br />
<p>10. Obtain the plasmid as the flowthrough.</p><br />
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Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol6
Team:Sumbawagen/Notebook/protocol6
2014-10-18T00:55:48Z
<p>Adel: </p>
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>Notebook – Protocol – 6. Plasmid purification</h2><br />
<p>Spin column based purification was used. The kit is Plasmid mini kit (Nippon Genetics). Standard protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge.</p><br />
<br />
<ul><br />
1. Add 2 ml of E. coli overnight culture into 2 ml polypropylene tube. Centrifuge for 6,000 rpm; 5 minutes. Remove the supernatant.<br />
2. Add 200 ul of mP1; vortexing. Add 200 ul of mP2; invert the tube 10 times. Incubate at 2 minutes at room temperature. Add 300 ul of mP3; invert the tube.<br />
3. Immediately centrifuge for 6,000 rpm; 5 minutes.<br />
4. Load the supernatant onto spin column. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.<br />
5. Add 400 ul of mP4. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.<br />
6. Add 600 ul of mP5. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.<br />
7. Centrifuge 6,000 rpm; 5 minute to dry the column.<br />
8. Replace collecting tube with clean, sterlized 1,5 ml size polypropylene tube.<br />
9. Add 50 ul of mP6. Incubate at room temperature for 2 minutes. Centrifuge for 6,000 rpm; 5 minutes.<br />
10. Obtain the plasmid as the flowthrough.<br />
<br />
</ul><br />
<br />
<p>2. Samples were incubated at 37 oC for more than 1 hour (typically 3 hours).</p><br />
<p>3. When necessary, samples were electrophoresed using protocol “3. Electrophoresis”(typically 3 hours).</p><br />
<p>4. Samples were purified using protocol “4. DNA purification”.</p></div>
Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol6
Team:Sumbawagen/Notebook/protocol6
2014-10-18T00:55:20Z
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<h2>Notebook – Protocol – 6. Plasmid purification</h2><br />
<p>Spin column based purification was used. The kit is Plasmid mini kit (Nippon Genetics). Standard protocols from the kit were followed by adaptation because we have no high speed microcentrifuge, but only a 6,000 rpm small microcentrifuge.</p><br />
<br />
<p><ul><br />
1. Add 2 ml of E. coli overnight culture into 2 ml polypropylene tube. Centrifuge for 6,000 rpm; 5 minutes. Remove the supernatant.<br />
2. Add 200 ul of mP1; vortexing. Add 200 ul of mP2; invert the tube 10 times. Incubate at 2 minutes at room temperature. Add 300 ul of mP3; invert the tube.<br />
3. Immediately centrifuge for 6,000 rpm; 5 minutes.<br />
4. Load the supernatant onto spin column. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.<br />
5. Add 400 ul of mP4. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.<br />
6. Add 600 ul of mP5. Centrifuge 6,000 rpm; 1 minute. Discard the flowthrough.<br />
7. Centrifuge 6,000 rpm; 5 minute to dry the column.<br />
8. Replace collecting tube with clean, sterlized 1,5 ml size polypropylene tube.<br />
9. Add 50 ul of mP6. Incubate at room temperature for 2 minutes. Centrifuge for 6,000 rpm; 5 minutes.<br />
10. Obtain the plasmid as the flowthrough.<br />
<br />
</p></ul><br />
<br />
<p>2. Samples were incubated at 37 oC for more than 1 hour (typically 3 hours).</p><br />
<p>3. When necessary, samples were electrophoresed using protocol “3. Electrophoresis”(typically 3 hours).</p><br />
<p>4. Samples were purified using protocol “4. DNA purification”.</p></div>
Adel
http://2014.igem.org/Team:Sumbawagen/Notebook/protocol5
Team:Sumbawagen/Notebook/protocol5
2014-10-18T00:44:45Z
<p>Adel: </p>
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<h2>Notebook – Protocol – 5. Restriction enzymes digestion</h2><br />
<p>1. Restriction enzymes digestion was done using Fast Digest product (Thermoscientific) using the following composition:</p><br />
<ul style="padding-left:25px;"><br />
<li>DNA sample (1-3 ug), 15 ul</li><br />
<li>Fast Digest restriction enzyme #1, 2 ul</li><br />
<li>Fast Digest restriction enzyme #2, 2 ul</li><br />
<li>10x buffer, 3 ul</li><br />
<li>Sterilized distilled water, 6 ul</li><br />
Total volume, 30 ul</li><br />
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<p>2. Samples were incubated at 37 oC for more than 1 hour (typically 3 hours).</p><br />
<p>3. When necessary, samples were electrophoresed using protocol “3. Electrophoresis”(typically 3 hours).</p><br />
<p>4. Samples were purified using protocol “4. DNA purification”.</p><br />
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<h1 class="firstHeading">Team:Sumbawagen/Team</h1><br />
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<h2>Notebook – Protocol – 5. Restriction enzymes digestion</h2><br />
<p>1. Restriction enzymes digestion was done using Fast Digest product (Thermoscientific) using the following composition:</p><br />
<ul style="padding-left:25px;"><br />
<li>DNA sample (1-3 ug), 15 ul</li><br />
<li>Fast Digest restriction enzyme #1, 2 ul</li><br />
<li>Fast Digest restriction enzyme #2, 2 ul</li><br />
<li>10x buffer, 3 ul</li><br />
<li>Sterilized distilled water, 6 ul</li><br />
Total volume, 30 ul</li><br />
</p></ul><br />
<br />
<p>2. Samples were incubated at 37 oC for more than 1 hour (typically 3 hours).</p><br />
<p>3. When necessary, samples were electrophoresed using protocol “3. Electrophoresis”(typically 3 hours).</p><br />
<p>4. Samples were purified using protocol “4. DNA purification”.</p></div>
Adel
http://2014.igem.org/File:8_diagram.jpg
File:8 diagram.jpg
2014-10-18T00:31:24Z
<p>Adel: </p>
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Adel