"
Team:SYSU-China/file/Project/Result/Integration.html
From 2014.igem.org
MolidantaTW (Talk | contribs) |
|||
| Line 4: | Line 4: | ||
<p>We construct plac-Or2-FourU-GFP on the pSB4A5 for the test.So that we can see whether FourU can control GFP translation in a temperature dependent manner.</p> | <p>We construct plac-Or2-FourU-GFP on the pSB4A5 for the test.So that we can see whether FourU can control GFP translation in a temperature dependent manner.</p> | ||
<a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/c/ce/Sysu-OFG.png"><img src="https://static.igem.org/mediawiki/2014/c/ce/Sysu-OFG.png" style="width:450px" alt="" /></a> <br /> | <a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/c/ce/Sysu-OFG.png"><img src="https://static.igem.org/mediawiki/2014/c/ce/Sysu-OFG.png" style="width:450px" alt="" /></a> <br /> | ||
| - | Figure 1 | + | <p1 style="text-align: center">Figure 1</p1> |
<p>We co-transformed the plasmid as the following groups:</p> | <p>We co-transformed the plasmid as the following groups:</p> | ||
<a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png"><img src="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png" alt="" /></a> <br /> | <a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png"><img src="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png" alt="" /></a> <br /> | ||
| - | Figure 2 | + | <p1 style="text-align: center">Figure 2</p1> |
<p>These four groups of bacteria were cultured at 30℃ and 37℃ respectively. All the OD was modified to be the same before the sample was tested by 510nm as excitation wavelength and 550nm as emission wavelength.</p> | <p>These four groups of bacteria were cultured at 30℃ and 37℃ respectively. All the OD was modified to be the same before the sample was tested by 510nm as excitation wavelength and 550nm as emission wavelength.</p> | ||
<a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/9/93/SysuchinaRNAT1.png"><img src="https://static.igem.org/mediawiki/2014/9/93/SysuchinaRNAT1.png" alt="" /></a> <br /> | <a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/9/93/SysuchinaRNAT1.png"><img src="https://static.igem.org/mediawiki/2014/9/93/SysuchinaRNAT1.png" alt="" /></a> <br /> | ||
| - | Figure 3 | + | <p1 style="text-align: center">Figure 3</p1> |
<h2><b>2.Bacterial Two-Hybrid System integrated with M13</b></h2> | <h2><b>2.Bacterial Two-Hybrid System integrated with M13</b></h2> | ||
<p>The reporter gene on pRPT is gene-Ⅷ from M13 bacteriophage. The corresponding plasmid is named pRPT-gⅧ.</p> | <p>The reporter gene on pRPT is gene-Ⅷ from M13 bacteriophage. The corresponding plasmid is named pRPT-gⅧ.</p> | ||
| Line 16: | Line 16: | ||
"><img src="https://static.igem.org/mediawiki/2014/c/c9/Sysu-Plasmid_O8.png | "><img src="https://static.igem.org/mediawiki/2014/c/c9/Sysu-Plasmid_O8.png | ||
" style="width:450px" alt="" /></a> <br /> | " style="width:450px" alt="" /></a> <br /> | ||
| - | Figure 4 | + | <p1 style="text-align: center">Figure 4</p1> |
<p>We co-transformed the plasmid as the following groups:</p> | <p>We co-transformed the plasmid as the following groups:</p> | ||
<a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png"><img src="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png" alt="" /></a> <br /> | <a class="fancybox" rel="group" href="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png"><img src="https://static.igem.org/mediawiki/2014/c/cd/Sysu-Biao_FG.png" alt="" /></a> <br /> | ||
| - | Figure 5 | + | <p1 style="text-align: center">Figure 5</p1> |
<p> | <p> | ||
After the co-transformations as follows, we tested if gene Ⅷ can be expressed in Bacterial Two-Hybrid System. | After the co-transformations as follows, we tested if gene Ⅷ can be expressed in Bacterial Two-Hybrid System. | ||
| Line 26: | Line 26: | ||
"><img src="https://static.igem.org/mediawiki/2014/6/6a/SysuchinaRNAT2.png | "><img src="https://static.igem.org/mediawiki/2014/6/6a/SysuchinaRNAT2.png | ||
" alt="" /></a> | " alt="" /></a> | ||
| - | Figure 6 | + | <p1 style="text-align: center">Figure 6</p1> |
<p> | <p> | ||
LB with 1/4 of each basic antibiotic concentration was used to screen for the bacteria containing all three plasmids in each group. Then liquid LB with 1/3 of each basic antibiotic concentration was used to culture the bacteria for 24 hours before the OD was modified to be consistent. After that, the bacteria were broken down and centrifuged. The intact bacteria, the supernatant and sediment from the centrifugation were all prepared for samples of Western Blot. gene Ⅷ was fused to FLAG for the following Western Blotting analysis. | LB with 1/4 of each basic antibiotic concentration was used to screen for the bacteria containing all three plasmids in each group. Then liquid LB with 1/3 of each basic antibiotic concentration was used to culture the bacteria for 24 hours before the OD was modified to be consistent. After that, the bacteria were broken down and centrifuged. The intact bacteria, the supernatant and sediment from the centrifugation were all prepared for samples of Western Blot. gene Ⅷ was fused to FLAG for the following Western Blotting analysis. | ||
</p> | </p> | ||
<!--SYSUCHINA!--> | <!--SYSUCHINA!--> | ||
Revision as of 02:22, 18 October 2014
Integration
Bacterial Two-Hybrid System integrate with RNAT
We construct plac-Or2-FourU-GFP on the pSB4A5 for the test.So that we can see whether FourU can control GFP translation in a temperature dependent manner.
<a class="fancybox" rel="group" href="
"><img src="" style="width:450px" alt="" /></a>
<p1 style="text-align: center">Figure 1</p1>
We co-transformed the plasmid as the following groups:
<a class="fancybox" rel="group" href="
"><img src="" alt="" /></a>
<p1 style="text-align: center">Figure 2</p1>
These four groups of bacteria were cultured at 30℃ and 37℃ respectively. All the OD was modified to be the same before the sample was tested by 510nm as excitation wavelength and 550nm as emission wavelength.
<a class="fancybox" rel="group" href="
"><img src="" alt="" /></a>
<p1 style="text-align: center">Figure 3</p1>
2.Bacterial Two-Hybrid System integrated with M13
The reporter gene on pRPT is gene-Ⅷ from M13 bacteriophage. The corresponding plasmid is named pRPT-gⅧ.
<a class="fancybox" rel="group" href="
"><img src="
" style="width:450px" alt="" /></a>
<p1 style="text-align: center">Figure 4</p1>
We co-transformed the plasmid as the following groups:
<a class="fancybox" rel="group" href=""><img src="" alt="" /></a>
<p1 style="text-align: center">Figure 5</p1>
After the co-transformations as follows, we tested if gene Ⅷ can be expressed in Bacterial Two-Hybrid System.
<a class="fancybox" rel="group" href="
"><img src="
" alt="" /></a>
<p1 style="text-align: center">Figure 6</p1>
LB with 1/4 of each basic antibiotic concentration was used to screen for the bacteria containing all three plasmids in each group. Then liquid LB with 1/3 of each basic antibiotic concentration was used to culture the bacteria for 24 hours before the OD was modified to be consistent. After that, the bacteria were broken down and centrifuged. The intact bacteria, the supernatant and sediment from the centrifugation were all prepared for samples of Western Blot. gene Ⅷ was fused to FLAG for the following Western Blotting analysis.
