Plasmid
pEryU

Part:BBa_K4383044

Designed by: Junxiao Zhang   Group: iGEM22_SCU-China   (2022-10-10)


pEryU

plasmid backbone is p15A, antibiotic resistance is kanamycin resistance. It functions by coding EryABCRHI, so that bacteria can utilize erythritol.

the heterogeneous expression of erythritol utilizing pathway could function successfully in E. coli.

Ery Metabolism.jpeg

figure1.Ery Metabolism pathway

Ery growth condition.png

figure2.Ery growth conditions of E. coliDH5a with pEryU in M9 erythritol solo carbon source

Ery growth curve.png

figure3.growth curve of E. coliDH5a in M9 erythritol solo carbon source

Scusu.png

figure4.Ery growth conditions of E. coliDH5a with EryABC, EryRHI, or pEryU in M9 erythritol solo carbon source (18 hours culture)

Scu6.png

figure5.growth curve of pEryU carrying probiotic E. coli Nissle 1917 under the solo carbon source erythritol

Scu7.png

figure6.growth curve of pEryU carrying probiotic E. coli Nissle 1917 under a gradient of erythritol, from 0.4g/L to 4g/L in M9 glucose culture

Scu8.png

figure7.growth curve of carrying probiotic E. coli Nissle 1917 under a gradient of different dietary intake proportions of erythritol and glucose

Scu9.png

figure8.growth curve of carrying probiotic E. coli Nissle 1917,10g/L glucose concentration as the intestinal environment under the normal diet for simulation. 2g/L erythritol to simulate taking 1-2 bottles of erythritol drinking with the normal diet and used 5g/L erythritol + 5g/L glucose, 10g/L erythritol simulating half of the sugar intake was replaced by erythritol, all of the sugar intake was replaced by erythritol, respectively

Scu39.png

figure9.RT-qpcr result of pEryU_pUC57-pJ23106-tmd_E. coli

In general, we successfully cloned and characterized the gene for erythritol utilization. The erythritol metabolic pathway was reconstructed in Escherichia coli to make it capable of using erythritol as an independent carbon source for growth. At the same time, we also investigated the effects of different carbon source ratios and different erythritol concentrations on probiotic E. coli Nissle 1917 (EcN) strains that could use erythritol (pEryU-EcN). Finally, we simulated the normal diet and found that a certain amount of erythritol in the normal diet increased the growth of pEryU-EcN, providing validation of our design.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
    Illegal NheI site found at 4355
    Illegal NheI site found at 4378
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 5998
    Illegal AgeI site found at 880
    Illegal AgeI site found at 1324
    Illegal AgeI site found at 2166
    Illegal AgeI site found at 3347
    Illegal AgeI site found at 5414
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 5501


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