Difference between revisions of "Part:BBa K1666006"
(→‎Result)
Line 13: Line 13:
 
[[File:NEFU_China_2015_Plsr_Blue.png|450px|thumb|center|'''Fig2. Part of the vector containing ''lsrA'' '''We use nisA promoter to initiate the transcription of ''lsrA'' gene]]
 
[[File:NEFU_China_2015_Plsr_Blue.png|450px|thumb|center|'''Fig2. Part of the vector containing ''lsrA'' '''We use nisA promoter to initiate the transcription of ''lsrA'' gene]]
  
−
===Result===
+
===Characterization===
  
−
[[File:NEFU_China_2015_pnisA_and_LsrA.png|500px|thumb|center|'''Fig3. Engineered E. coli Trans T1 contains pHY300PLK-Plsr-amilCP ''']]
+
[[File:NEFU_China_2015_Ecoli_Blue.png|500px|thumb|center|'''Fig3. Engineered ''E. coli'' Trans T1 contains pHY300PLK-P''lsr''-''amilCP'' ''']]
  
−
[[File:NEFU_China_2015_pnisA_and_LsrA.png|500px|thumb|center|'''Fig4. Yogurt guarder incubated with culture supernatant of different bacteria ''']]
+
[[File:NEFU_China_2015_Yogurt_Blue.png|500px|thumb|center|'''Fig4. Yogurt guarder incubated with culture supernatant of different bacteria ''']]
  
  

Revision as of 01:23, 19 September 2015

lsr promoter of LuxS/AI-2 signaling pathway in Salmonalla

Quorum sensing is a process of bacterial cell-to-cell communication involving the production and detection of extracellular signaling molecules called autoinducers. And autoinducer-2 (AI-2) has been proposed to serve as a 'universal signal' for interspecies communication. In the LuxS/AI-2 signaling system of Salmonella Typhimurium, AI-2 response involves ATP binding cassette transporter encoded by genes named Lsr (LuxS regulated). Plsr is the promoter of the lsr operon. In our project, we use it to trigger the expression of a reporter gene.

Usage and Biology

AI-2 is generated by many species of Gram-negative and Gram-positive bacteria. In a group of bacteria exemplified by Salmonella, AI-2 response involves lsr genes that encode ATP binding cassette-type transporter. Plsr is the promoter of the lsr operon.

Fig1. Schematic overview of the AI-2 response pathway in Salmonella TyphimuriumThe precursor of AI-2, 4,5-Dihydroxy-2,3-Pentanedione (DPD) , is a byproduct generated when LuxS converts S-Ribosylhomocysteine (SRH) to Homocysteine (HCY). DPD then undergoes spontaneously cyclization, forming AI-2, and exports to the culture supernatant. After that, extracellular AI-2 bounds to LsrB, following by passing the membrane channel and importing the cytoplasm. LsrK phosphorylates AI-2 afterwards. The lsr operon is repressed until phosphorylated AI-2 causes LsrR to relieve its repression on the promoter. And this allows further AI-2 import.

In our project, we set this protein-coding part under the regulation of a nisA promoter which can be activated by food-grade inducer, nisin. We linearized the related expression vectors and stably integrated them into the genome of the hosts. And together with other parts, we will construct a membrane channel for AI-2 generated by pathogens in the engineered bacteria.

Fig2. Part of the vector containing lsrA We use nisA promoter to initiate the transcription of lsrA gene

Characterization

Fig3. Engineered E. coli Trans T1 contains pHY300PLK-Plsr-amilCP
Fig4. Yogurt guarder incubated with culture supernatant of different bacteria


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]