Difference between revisions of "Part:BBa K1363400"
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<p>This part is our integral blue light pathway. It consists of a blue light inducing amilCP expression system, a blue light sensor where YF1 and FixJ sense the light and lambda CI acts as the not gate to control blue protein expression.</p> | <p>This part is our integral blue light pathway. It consists of a blue light inducing amilCP expression system, a blue light sensor where YF1 and FixJ sense the light and lambda CI acts as the not gate to control blue protein expression.</p> | ||
| − | <p>The circuit is as following: We combine the light sensor and amiICP expression system on the same plasmid and it can let the bacteria complete blue photography independently. | + | <p>The circuit is as following: We combine the light sensor and amiICP expression system on the same plasmid and it can let the bacteria complete blue photography independently.[fig 1]</p> |
<figure><img src="https://static.igem.org/mediawiki/parts/5/50/USTC-5.jpg" width="300"/><figcaption>Fig 1</figcaption></figure> | <figure><img src="https://static.igem.org/mediawiki/parts/5/50/USTC-5.jpg" width="300"/><figcaption>Fig 1</figcaption></figure> | ||
<p>In the blue light-inducible system, FixK2 promoter is the downstream signaling promoter of YF1/FixJ blue light-sensing system. The YF1/FixJ system becomes inactive when illuminated by blue otherwise active. An inverter is placed behind FixK2 promoter.
In the absence of blue light, YF1 phosphorylates FixJ that activates FixK2 promoter, allowing the transcription of lambda CI, thus repressing the amilCP output (Blue Chromophore Protein).</p> | <p>In the blue light-inducible system, FixK2 promoter is the downstream signaling promoter of YF1/FixJ blue light-sensing system. The YF1/FixJ system becomes inactive when illuminated by blue otherwise active. An inverter is placed behind FixK2 promoter.
In the absence of blue light, YF1 phosphorylates FixJ that activates FixK2 promoter, allowing the transcription of lambda CI, thus repressing the amilCP output (Blue Chromophore Protein).</p> | ||
<h3>Experiments:</h3> | <h3>Experiments:</h3> | ||
<h4>1. Test in solid media</h4> | <h4>1. Test in solid media</h4> | ||
| − | <p>Without induced expression, bacteria containing blue light sensing-imaging system grew up and developed in the LB media. Then we divided the media into two regions, one of them received blue light simulation while the other one kept darkness by a base plate to block blue light. The testing conditions are showed in the following</p> | + | <p>Without induced expression, bacteria containing blue light sensing-imaging system grew up and developed in the LB media. Then we divided the media into two regions, one of them received blue light simulation while the other one kept darkness by a base plate to block blue light. The testing conditions are showed in the following[fig 2]</p> |
<figure><img src="https://static.igem.org/mediawiki/parts/3/3d/USTC-2.png" width="300"/><figcaption>Fig 2</figcaption></figure> | <figure><img src="https://static.igem.org/mediawiki/parts/3/3d/USTC-2.png" width="300"/><figcaption>Fig 2</figcaption></figure> | ||
| − | <p>After 12-hours' development, the result showed the bacteria could produce blue chromo-protein when stimulated by blue light and kept its color in the darkness:</p> | + | <p>After 12-hours' development, the result showed the bacteria could produce blue chromo-protein when stimulated by blue light and kept its color in the darkness:[fig 3]</p> |
<figure><img src="https://static.igem.org/mediawiki/parts/9/95/USTC-3.png" width="300"/><figcaption>Fig 3</figcaption></figure> | <figure><img src="https://static.igem.org/mediawiki/parts/9/95/USTC-3.png" width="300"/><figcaption>Fig 3</figcaption></figure> | ||
<h4>2. Test in liquid media</h4> | <h4>2. Test in liquid media</h4> | ||
<p>The idea of control experiment is the same as that ain solid media. We prepared two test tubes containing equivalent bacterium solution. The test was conducted at 18:00, August 4th when one of them was put under blue light and the other was developed in the darkness. </p> | <p>The idea of control experiment is the same as that ain solid media. We prepared two test tubes containing equivalent bacterium solution. The test was conducted at 18:00, August 4th when one of them was put under blue light and the other was developed in the darkness. </p> | ||
<p>At 11:00, August 5th, the tube stimulated under blue light turned light blue while the other one kept the original color. </p> | <p>At 11:00, August 5th, the tube stimulated under blue light turned light blue while the other one kept the original color. </p> | ||
| − | <p>After 24 hours, the blue bacteria showed evidently. Using centrifuge to get the bacteria showed extremely distinct blue chromo-protein expression based on blue light activation was come true.</p> | + | <p>After 24 hours, the blue bacteria showed evidently. Using centrifuge to get the bacteria showed extremely distinct blue chromo-protein expression based on blue light activation was come true.[fig 4]</p> |
<figure><img src="https://static.igem.org/mediawiki/parts/9/94/USTC-1.png" width="300"/><figcaption>Fig 4</figcaption></figure> | <figure><img src="https://static.igem.org/mediawiki/parts/9/94/USTC-1.png" width="300"/><figcaption>Fig 4</figcaption></figure> | ||
<h4>3. Pattern Test</h4> | <h4>3. Pattern Test</h4> | ||
| − | <p>Test of a blue sun pattern focused on sharp shape of a picture is in the following. The result showed bacteria could produce protein at the edge of the picture accurately.</p> | + | <p>Test of a blue sun pattern focused on sharp shape of a picture is in the following. The result showed bacteria could produce protein at the edge of the picture accurately.[fig 5]</p> |
<figure><img src="https://static.igem.org/mediawiki/parts/c/c1/USTC-4.jpg" width="300"/><figcaption>Fig 5</figcaption></figure> | <figure><img src="https://static.igem.org/mediawiki/parts/c/c1/USTC-4.jpg" width="300"/><figcaption>Fig 5</figcaption></figure> | ||
<h4>4.The interference of other color </h4> | <h4>4.The interference of other color </h4> | ||
<p>In order to see if other colors will influence the blue inducible bacteria, we observed cell populations containing the blue light inducible system grown in four tubes under same conditions.However are differently illuminated with red light, green light and blue light and in dark. Their growing curves are approximately the same with acceptable deviation. The growth of dark exposed bacteria slightly surpassed others since they eat a lot and produce less, which should has nothing to do with repress from the light.</p> | <p>In order to see if other colors will influence the blue inducible bacteria, we observed cell populations containing the blue light inducible system grown in four tubes under same conditions.However are differently illuminated with red light, green light and blue light and in dark. Their growing curves are approximately the same with acceptable deviation. The growth of dark exposed bacteria slightly surpassed others since they eat a lot and produce less, which should has nothing to do with repress from the light.</p> | ||
| − | <p>We may conclude that neither red nor green light has repressive effects on blue inducible bacteria</p> | + | <p>We may conclude that neither red nor green light has repressive effects on blue inducible bacteria[fig 6]</p> |
<figure><img src="https://static.igem.org/mediawiki/parts/d/de/USTC-Grow.png" width="300"/><figcaption>Fig 6</figcaption></figure> | <figure><img src="https://static.igem.org/mediawiki/parts/d/de/USTC-Grow.png" width="300"/><figcaption>Fig 6</figcaption></figure> | ||
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<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
<partinfo>BBa_K1363400 SequenceAndFeatures</partinfo> | <partinfo>BBa_K1363400 SequenceAndFeatures</partinfo> | ||
Revision as of 05:25, 1 November 2014
R0010-K592016-B0015-K502020
This part is our integral blue light pathway. It consists of a blue light inducing amilCP expression system, a blue light sensor where YF1 and FixJ sense the light and lambda CI acts as the not gate to control blue protein expression.
The circuit is as following: We combine the light sensor and amiICP expression system on the same plasmid and it can let the bacteria complete blue photography independently.[fig 1]
In the blue light-inducible system, FixK2 promoter is the downstream signaling promoter of YF1/FixJ blue light-sensing system. The YF1/FixJ system becomes inactive when illuminated by blue otherwise active. An inverter is placed behind FixK2 promoter. In the absence of blue light, YF1 phosphorylates FixJ that activates FixK2 promoter, allowing the transcription of lambda CI, thus repressing the amilCP output (Blue Chromophore Protein).
Experiments:
1. Test in solid media
Without induced expression, bacteria containing blue light sensing-imaging system grew up and developed in the LB media. Then we divided the media into two regions, one of them received blue light simulation while the other one kept darkness by a base plate to block blue light. The testing conditions are showed in the following[fig 2]
After 12-hours' development, the result showed the bacteria could produce blue chromo-protein when stimulated by blue light and kept its color in the darkness:[fig 3]
2. Test in liquid media
The idea of control experiment is the same as that ain solid media. We prepared two test tubes containing equivalent bacterium solution. The test was conducted at 18:00, August 4th when one of them was put under blue light and the other was developed in the darkness.
At 11:00, August 5th, the tube stimulated under blue light turned light blue while the other one kept the original color.
After 24 hours, the blue bacteria showed evidently. Using centrifuge to get the bacteria showed extremely distinct blue chromo-protein expression based on blue light activation was come true.[fig 4]
3. Pattern Test
Test of a blue sun pattern focused on sharp shape of a picture is in the following. The result showed bacteria could produce protein at the edge of the picture accurately.[fig 5]
4.The interference of other color
In order to see if other colors will influence the blue inducible bacteria, we observed cell populations containing the blue light inducible system grown in four tubes under same conditions.However are differently illuminated with red light, green light and blue light and in dark. Their growing curves are approximately the same with acceptable deviation. The growth of dark exposed bacteria slightly surpassed others since they eat a lot and produce less, which should has nothing to do with repress from the light.
We may conclude that neither red nor green light has repressive effects on blue inducible bacteria[fig 6]
Sequence and Features