Difference between revisions of "Team:Warwick/Results"

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$("#Content1").html("<h2>Did we get our plasmids in?</h2>GELS: To assess whether the transformations we carried were correct, we amplified the identified and ran gels to compare the lengths of the DNA present in the plasmids present in the cells. Though we did not do this for every transformation, some of the most exciting one which we ran gels of were: * The first of our transformations was inserting JDE131 with GvpA into DH5A (E. coli). As can be seen from the gel, the bands amplified were at just under 500bp which is close to the expected length of the amplicon which would be 468bp. <img src='https://~~l.messenger.com/l.php?u=http%3A%2F%2F2018~~.igem.org~~%2FFile%3AT~~--Warwick--2018-FMX1.png~~&h=AT0L2PEQieLC2g1emnsiShdv2dP-E5lpl4A-F36SnIuGP72NTj-JyXzhR69Du3VpycZ06AVjP3nE7444rrahtLczJdDyFTRgw0hEX3FOCzUoMRibXiLC2ZZekQm0lFVr52UCKg~~'> * The next of our transformations was inserting ECE174 with GvpC into DH5A (E. coli). As can be seen from the gel, the bands amplified were at around 500bp which is close to the expected length of the amplicon, which only amplified the GvpC gene, which is 500bp. <img src=''> Another one our transformations was inserting JDE131 with CFP into DH5A (E. coli). As can be seen from the gel, the bands amplified were at … which is close to the expected length of the amplicon which would be 961bp. <img src='https://static.igem.org/mediawiki/2018/9/9d/T--Warwick--2018-FMX3.png'>");

+

$("#Content1").html("<h2>Did we get our plasmids in?</h2>GELS: To assess whether the transformations we carried were correct, we amplified the identified and ran gels to compare the lengths of the DNA present in the plasmids present in the cells. Though we did not do this for every transformation, some of the most exciting one which we ran gels of were: * The first of our transformations was inserting JDE131 with GvpA into DH5A (E. coli). As can be seen from the gel, the bands amplified were at just under 500bp which is close to the expected length of the amplicon which would be 468bp. <img src='https://static.igem.org/mediawiki/2018/e/eb/T--Warwick--2018-FMX1.png'> * The next of our transformations was inserting ECE174 with GvpC into DH5A (E. coli). As can be seen from the gel, the bands amplified were at around 500bp which is close to the expected length of the amplicon, which only amplified the GvpC gene, which is 500bp. <img src='https://static.igem.org/mediawiki/parts/d/d0/T--Warwick--2018-GelGvpC.png'> Another one our transformations was inserting JDE131 with CFP into DH5A (E. coli). As can be seen from the gel, the bands amplified were at … which is close to the expected length of the amplicon which would be 961bp. <img src='https://static.igem.org/mediawiki/2018/9/9d/T--Warwick--2018-FMX3.png'>");

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$("#Content2").html("<h2>Plate growth shows that the plasmids did get in as they survived in antiobiotics</h2> * Our plasmids have antibiotic resistant genes to ensure that the cell retains the associated genes, and thus can be grown on antibiotic plates. * We observed LBA Ampicillin plates with GvpA E. coli and CFP E. coli grown on them. * We also observed LBA Spectinomycin with GvpA B. subtilis grown on them and LBA Chloramphenicol plates with GvpC B. subtilis grown on them. <h2>Sequencing the data of plasmids</h2>We were also able to miniprep or PCR purify the following inserts to sequence verify them: GvpA E. coli, GvpC E. coli, GvpA B. sub, CFP E. coli. The sequencing results can be seen below, and show that they were correctly inserted without significant mutations. GvpA E. coli <img src=''> GvpC E. coli <img src=''> CFP E. coli <img src=''> GvpA B. subtilis <img src=''>");

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$("#Content2").html("<h2>Plate growth shows that the plasmids did get in as they survived in antiobiotics</h2> * Our plasmids have antibiotic resistant genes to ensure that the cell retains the associated genes, and thus can be grown on antibiotic plates. * We observed LBA Ampicillin plates with GvpA E. coli and CFP E. coli grown on them. * We also observed LBA Spectinomycin with GvpA B. subtilis grown on them and LBA Chloramphenicol plates with GvpC B. subtilis grown on them. <h2>Sequencing the data of plasmids</h2>We were also able to miniprep or PCR purify the following inserts to sequence verify them: GvpA E. coli, GvpC E. coli, GvpA B. sub, CFP E. coli. The sequencing results can be seen below, and show that they were correctly inserted without significant mutations. GvpA E. coli <img src='https://static.igem.org/mediawiki/2018/3/33/T--Warwick--2018-FMX5.gif'> GvpC E. coli <img src='https://static.igem.org/mediawiki/2018/c/ca/T--Warwick--2018-FMX7.png'> CFP E. coli <img src='https://static.igem.org/mediawiki/2018/1/1c/T--Warwick--2018-FMX9.png'> GvpA B. subtilis <img src='https://static.igem.org/mediawiki/2018/d/dc/T--Warwick--leadthing.png'>");

$("#Content3").html("<h2>Did we get our plasmids to work?</h2>In order to characterise our promoters, we inserted a yellow fluorescent protein after both of the bacillus promoters (Hyperspank and Popp). We collected results that suggested that these promoters were successful in transcribing the fluorescent proteins and were thus functional in Bacillus subtilis. The data showed that the higher level of IPTG present, the higher level of fluorescence in the inducible promoter (Hyperspank). The data also compares the difference between the inducible (Hyperspank) and the constitutive (Popp) promoters fluorescence, showing that Popp has almost three-fold expression. The full data for our characterisation is available on our registry, as well as extensively detailed on our models page. We also got addgene mega cluster into E. coli and conducted several floating experiments to see whether or not the cells floated. We saw qualitative preliminary data, that suggested gas vesicles may have been formed. This is shown in example, by the image below which shows the formation of bubbles at the top of the culture based on the level of IPTG concentration: at 0ul IPTG and 200ul IPTG there were no bubble present; this is likely because the promoters were not induced and over-worked respectively. 2ul shows the highest amount of bubbles, this is expected number at which the T7 promoter would be most efficient and, 20ul has a few bubbles, as, again, it is likely overworked. As mentioned, this is extremely preliminary data, but was replicated and showed similar qualitative results.<img src=''>");

Difference between revisions of "Team:Warwick/Results"

Latest revision as of 03:54, 18 October 2018

Results

Contact Us

igem@warwick.ac.uk

Sponsors

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          $("#inorgIMG").css("background-color", "grey");
-         $("#Content1").html("<h2>Did we get our plasmids in?</h2><p>GELS:<br>To assess whether the transformations we carried were correct, we amplified the identified and ran gels to compare the lengths of the DNA present in the plasmids present in the cells. Though we did not do this for every transformation, some of the most exciting one which we ran gels of were:<br><br> * The first of our transformations was inserting JDE131 with GvpA into DH5A (E. coli). As can be seen from the gel, the bands amplified were at just under 500bp which is close to the expected length of the amplicon which would be 468bp.</p><br><img src='https://l.messenger.com/l.php?u=http%3A%2F%2F2018.igem.org%2FFile%3AT--Warwick--2018-FMX1.png&h=AT0L2PEQieLC2g1emnsiShdv2dP-E5lpl4A-F36SnIuGP72NTj-JyXzhR69Du3VpycZ06AVjP3nE7444rrahtLczJdDyFTRgw0hEX3FOCzUoMRibXiLC2ZZekQm0lFVr52UCKg'><br><p><br> * The next of our transformations was inserting ECE174 with GvpC into DH5A (E. coli). As can be seen from the gel, the bands amplified were at around 500bp which is close to the expected length of the amplicon, which only amplified the GvpC gene, which is 500bp.</p><br><img src=''><br><p>Another one our transformations was inserting JDE131 with CFP into DH5A (E. coli). As can be seen from the gel, the bands amplified were at … which is close to the expected length of the amplicon which would be 961bp.</p><br><img src='https://static.igem.org/mediawiki/2018/9/9d/T--Warwick--2018-FMX3.png'>");
+         $("#Content1").html("<h2>Did we get our plasmids in?</h2><p>GELS:<br>To assess whether the transformations we carried were correct, we amplified the identified and ran gels to compare the lengths of the DNA present in the plasmids present in the cells. Though we did not do this for every transformation, some of the most exciting one which we ran gels of were:<br><br> * The first of our transformations was inserting JDE131 with GvpA into DH5A (E. coli). As can be seen from the gel, the bands amplified were at just under 500bp which is close to the expected length of the amplicon which would be 468bp.</p><br><img src='https://static.igem.org/mediawiki/2018/e/eb/T--Warwick--2018-FMX1.png'><br><p><br> * The next of our transformations was inserting ECE174 with GvpC into DH5A (E. coli). As can be seen from the gel, the bands amplified were at around 500bp which is close to the expected length of the amplicon, which only amplified the GvpC gene, which is 500bp.</p><br><img src='https://static.igem.org/mediawiki/parts/d/d0/T--Warwick--2018-GelGvpC.png'><br><p>Another one our transformations was inserting JDE131 with CFP into DH5A (E. coli). As can be seen from the gel, the bands amplified were at … which is close to the expected length of the amplicon which would be 961bp.</p><br><img src='https://static.igem.org/mediawiki/2018/9/9d/T--Warwick--2018-FMX3.png'>");
-         $("#Content2").html("<h2>Plate growth shows that the plasmids did get in as they survived in antiobiotics</h2><p> * Our plasmids have antibiotic resistant genes to ensure that the cell retains the associated genes, and thus can be grown on antibiotic plates.<br> * We observed LBA Ampicillin plates with GvpA E. coli and CFP E. coli grown on them.<br> * We also observed LBA Spectinomycin with GvpA B. subtilis grown on them and LBA Chloramphenicol plates with GvpC B. subtilis grown on them.</p><br><br><h2>Sequencing the data of plasmids</h2><p>We were also able to miniprep or PCR purify the following inserts to sequence verify them: GvpA E. coli, GvpC E. coli, GvpA B. sub, CFP E. coli. The sequencing results can be seen below, and show that they were correctly inserted without significant mutations.<br><br>GvpA E. coli</p><br><img src=''><br><br><p>GvpC E. coli</p><br><img src=''><br><br><p>CFP E. coli</p><br><img src=''><br><br><p>GvpA B. subtilis</p><br><img src=''>");
+         $("#Content2").html("<h2>Plate growth shows that the plasmids did get in as they survived in antiobiotics</h2><p> * Our plasmids have antibiotic resistant genes to ensure that the cell retains the associated genes, and thus can be grown on antibiotic plates.<br> * We observed LBA Ampicillin plates with GvpA E. coli and CFP E. coli grown on them.<br> * We also observed LBA Spectinomycin with GvpA B. subtilis grown on them and LBA Chloramphenicol plates with GvpC B. subtilis grown on them.</p><br><br><h2>Sequencing the data of plasmids</h2><p>We were also able to miniprep or PCR purify the following inserts to sequence verify them: GvpA E. coli, GvpC E. coli, GvpA B. sub, CFP E. coli. The sequencing results can be seen below, and show that they were correctly inserted without significant mutations.<br><br>GvpA E. coli</p><br><img src='https://static.igem.org/mediawiki/2018/3/33/T--Warwick--2018-FMX5.gif'><br><br><p>GvpC E. coli</p><br><img src='https://static.igem.org/mediawiki/2018/c/ca/T--Warwick--2018-FMX7.png'><br><br><p>CFP E. coli</p><br><img src='https://static.igem.org/mediawiki/2018/1/1c/T--Warwick--2018-FMX9.png'><br><br><p>GvpA B. subtilis</p><br><img src='https://static.igem.org/mediawiki/2018/d/dc/T--Warwick--leadthing.png'>");
          $("#Content3").html("<h2>Did we get our plasmids to work?</h2><p>In order to characterise our promoters, we inserted a yellow fluorescent protein after both of the bacillus promoters (Hyperspank and Popp). We collected results that suggested that these promoters were successful in transcribing the fluorescent proteins and were thus functional in Bacillus subtilis. The data showed that the higher level of IPTG present, the higher level of fluorescence in the inducible promoter (Hyperspank). The data also compares the difference between the inducible (Hyperspank) and the constitutive (Popp) promoters fluorescence, showing that Popp has almost three-fold expression. The full data for our characterisation is available on our registry, as well as extensively detailed on our models page.<br><br><br>We also got addgene mega cluster into E. coli and conducted several floating experiments to see whether or not the cells floated. We saw qualitative preliminary data, that suggested gas vesicles may have been formed. This is shown in example, by the image below which shows the formation of bubbles at the top of the culture based on the level of IPTG concentration: at 0ul IPTG and 200ul IPTG there were no bubble present; this is likely because the promoters were not induced and over-worked respectively. 2ul shows the highest amount of bubbles, this is expected number at which the T7 promoter would be most efficient and, 20ul has a few bubbles, as, again, it is likely overworked.<br><br>As mentioned, this is extremely preliminary data, but was replicated and showed similar qualitative results.</p><img src=''>");