Difference between revisions of "Part:BBa T9002"
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The luxR based receiver, F2620 (formerly I13270), controls the production of GFP. The GFP protein generator is the same as that found in I7101. | The luxR based receiver, F2620 (formerly I13270), controls the production of GFP. The GFP protein generator is the same as that found in I7101. | ||
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__NOTOC__ | __NOTOC__ | ||
| − | + | the characterization and improvement of BBa_T9002 | |
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(Characterized by Peking_2019)<br> | (Characterized by Peking_2019)<br> | ||
| − | We retest the characterization of this part.Using different concentration of AHL,we get the corresponding | + | We retest the characterization of this part.Using different concentration of AHL,we get the corresponding fluorescence intensity and the result is as follows. |
<center>https://2019.igem.org/wiki/images/c/cb/T--Peking--QS2.png</center> | <center>https://2019.igem.org/wiki/images/c/cb/T--Peking--QS2.png</center> | ||
| − | <center>Figure1 The corresponding | + | <center>Figure1 The corresponding fluorescence intensity of different concentration of AHL.</center><br> |
Experiment procedure:<br> | Experiment procedure:<br> | ||
1.E. coli DH5α transferred with PSB1C3-T9002 were incubated in LB liquid medium at 37℃, 220rpm overnight<br> | 1.E. coli DH5α transferred with PSB1C3-T9002 were incubated in LB liquid medium at 37℃, 220rpm overnight<br> | ||
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We used <partinfo>BBa_T9002</partinfo> to characterize quorum sensing system. But we found that this part has an obvious homologous recombination. So we changed the terminator of GFP to avoid it. | We used <partinfo>BBa_T9002</partinfo> to characterize quorum sensing system. But we found that this part has an obvious homologous recombination. So we changed the terminator of GFP to avoid it. | ||
<center>https://https://2019.igem.org/wiki/images/c/c6/T--Peking--QS-.png</center> | <center>https://https://2019.igem.org/wiki/images/c/c6/T--Peking--QS-.png</center> | ||
| − | <center>Figure2 The design of quorum sensing system</center> | + | <center>Figure2 The design of quorum sensing system.</center> |
<center>https://2019.igem.org/wiki/images/1/10/T--Peking--QS1.png</center> | <center>https://2019.igem.org/wiki/images/1/10/T--Peking--QS1.png</center> | ||
| − | <center>Figure3 | + | <center>Figure3 A time-scale quorum sensing system can be transformed to spatial-level through a donor/receiver system. The donor cells, which merely express and release AHL, would activate the GFP expression of receiver cells through AHL diffusion. This is validated on the agar plate, by dropping the donor cells in the center and receiver cells around them with different distances. We found a progressive decrease in fluorescence as the receiver cells locate farther from central AHL donor. </center> |
Experiment procedure:<br> | Experiment procedure:<br> | ||
1.Incubate donor and receiver cells in LB medium overnight.<br> | 1.Incubate donor and receiver cells in LB medium overnight.<br> | ||
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3.5 μL donor cell was dropped in the LB solid medium. <br> | 3.5 μL donor cell was dropped in the LB solid medium. <br> | ||
4.5 Μl receptor cell was dropped in different distances in the same LB solid medium.<br> | 4.5 Μl receptor cell was dropped in different distances in the same LB solid medium.<br> | ||
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| + | <!-- Add more about the biology of this part here | ||
| + | ===Usage and Biology=== | ||
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| + | <!-- --> | ||
| + | <span class='h3bb'>Sequence and Features</span> | ||
| + | <partinfo>BBa_T9002 SequenceAndFeatures</partinfo> | ||
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| + | <!-- Uncomment this to enable Functional Parameter display | ||
| + | ===Functional Parameters=== | ||
| + | <partinfo>BBa_T9002 parameters</partinfo> | ||
| + | <!-- --> | ||
Revision as of 23:23, 20 October 2019
GFP Producer Controlled by 3OC6HSL Receiver Device
The luxR based receiver, F2620 (formerly I13270), controls the production of GFP. The GFP protein generator is the same as that found in I7101.
the characterization and improvement of BBa_T9002
(Characterized by Peking_2019)
We retest the characterization of this part.Using different concentration of AHL,we get the corresponding fluorescence intensity and the result is as follows.
Experiment procedure:
1.E. coli DH5α transferred with PSB1C3-T9002 were incubated in LB liquid medium at 37℃, 220rpm overnight
2.Grown culture was 100-fold diluted in M9 medium for the same condition for 3 hour.
3.Culture was 40-fold diluted in M9 medium containing different concentration of AHL.
4.200 µl aliquots of each of the cultures were transferred into a flat-bottomed 96 well plate.
5.FI and OD600 was measured for 10 hours in microplate reader.
We used BBa_T9002 to characterize quorum sensing system. But we found that this part has an obvious homologous recombination. So we changed the terminator of GFP to avoid it.
Experiment procedure:
1.Incubate donor and receiver cells in LB medium overnight.
2.Grown culture was 100-fold diluted in M9 medium.
3.5 μL donor cell was dropped in the LB solid medium.
4.5 Μl receptor cell was dropped in different distances in the same LB solid medium.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 1004
Illegal BsaI.rc site found at 1732