Difference between revisions of "Part:BBa K2279001"
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<span class='h3bb'>Sequence and Features</span> | <span class='h3bb'>Sequence and Features</span> | ||
<partinfo>BBa_K2279001 SequenceAndFeatures</partinfo> | <partinfo>BBa_K2279001 SequenceAndFeatures</partinfo> | ||
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| + | ===Plasmid construction=== | ||
| + | We used PCR to produce AimP gene fragment. | ||
| + | |||
| + | [[file:T--TMMU-China--aimP.jpeg]] | ||
| + | |||
| + | Then we inserted this gene to plasmid pSB1C3. We transformed this plasmid plays (one contains gene AimR) into strain DH5α (E. coli). Then we picked some colonies for cultivation and confirmed the transformation result by enzyme digestion using PstI and EcoRI enzymes. From the result of electrophoresis, we confirmed the transformation of AimR was success. | ||
| + | |||
| + | [[file:T--TMMU-China--aimp1.jpeg]] | ||
| + | |||
| + | We then sequenced the positive bacteria and confirmed that the plasmid plays (one contains gene AimR) was indeed transferred to the bacteria. We expanded the bacteria and extracted plasmids from the bacteria. So far, we've successfully constructed AimR. | ||
| + | |||
===Biological Function=== | ===Biological Function=== | ||
The B.subtilis bacteriophage phi3T employs the AimR-AimP QS system to make the lysis-lysogeny decision. AimP is the pre-pro-peptide. It is 43aa long. The mature signal peptide of AimP is SAIRGA. Binding of the mature signal peptide to AimR will disrupt the dimer forms of AimR. After that, the AimR can no longer bind to the promoter of AimX, a potential non coding RNA involved in the process of lysis-lysogeny. | The B.subtilis bacteriophage phi3T employs the AimR-AimP QS system to make the lysis-lysogeny decision. AimP is the pre-pro-peptide. It is 43aa long. The mature signal peptide of AimP is SAIRGA. Binding of the mature signal peptide to AimR will disrupt the dimer forms of AimR. After that, the AimR can no longer bind to the promoter of AimX, a potential non coding RNA involved in the process of lysis-lysogeny. | ||
| − | [[file: | + | [[file:T--TMMU-China--aimfu.jpeg]] |
===Design=== | ===Design=== | ||
By combining the expression of AimR and AimP components, we want to develop a synthetic QS system in B. subtilis for target gene autoinhibition. | By combining the expression of AimR and AimP components, we want to develop a synthetic QS system in B. subtilis for target gene autoinhibition. | ||
Revision as of 13:51, 31 October 2017
AimP
AimP encodes a signal peptide.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 58
- 1000COMPATIBLE WITH RFC[1000]
Plasmid construction
We used PCR to produce AimP gene fragment.
Then we inserted this gene to plasmid pSB1C3. We transformed this plasmid plays (one contains gene AimR) into strain DH5α (E. coli). Then we picked some colonies for cultivation and confirmed the transformation result by enzyme digestion using PstI and EcoRI enzymes. From the result of electrophoresis, we confirmed the transformation of AimR was success.
We then sequenced the positive bacteria and confirmed that the plasmid plays (one contains gene AimR) was indeed transferred to the bacteria. We expanded the bacteria and extracted plasmids from the bacteria. So far, we've successfully constructed AimR.
Biological Function
The B.subtilis bacteriophage phi3T employs the AimR-AimP QS system to make the lysis-lysogeny decision. AimP is the pre-pro-peptide. It is 43aa long. The mature signal peptide of AimP is SAIRGA. Binding of the mature signal peptide to AimR will disrupt the dimer forms of AimR. After that, the AimR can no longer bind to the promoter of AimX, a potential non coding RNA involved in the process of lysis-lysogeny.
Design
By combining the expression of AimR and AimP components, we want to develop a synthetic QS system in B. subtilis for target gene autoinhibition.
A synthetic communication pathway between B. subtilis strains by co-culturing AimP-producing “sender” cells with AimR-sensing “receiver” cells to inhibit gene expression was also designed.
