Part:BBa_K1510002

PnlmC (S.mutans)

This part is derived from Streptococcus mutans UA159, a gram positive bacterium. According to study done by Liu et al.[http://link.springer.com/article/10.1007%2Fs13213-013-0629-6 Liu, T., et al., ComCED signal loop precisely regulates nlmC expression in Streptococcus mutans. Annals of Microbiology, 2014. 64(1): p. 31-38.], nlmC promoter can sense competence stimulating peptide (CSP).As a result, the main function of this promoter is to sense a quorum sensing chemical, competence-stimulating peptide (CSP), and trigger down string gene.

The picture is derived from the study done by Liu et al., indicating the CSP stimulation of nlmC promoter.

[http://link.springer.com/article/10.1007%2Fs13213-013-0629-6 Liu, T., et al., ComCED signal loop precisely regulates nlmC expression in Streptococcus mutans. Annals of Microbiology, 2014. 64(1): p. 31-38.]

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]

2020 SZPT-CHINA

In 2014, the NYMU-Taipei team cited the relevant data of the article ComCED signal loop precisely regulates nlmC expression in Streptococcus mutans published by [http://link.springer.com/article/10.1007%2Fs13213-013-0629-6 Liu, T., et al.]in Annals of Microbiology, but did not characterize it. This year, we tested the part in conjunction with our project.

Contribution

The recombinant strain E. coli BL21 (DE3) with pET28a-DEG vector was as the detector. Different concentrate CSP was added into the culture medium. When the concentration of CSP reached 0.5mg/ml, the comD-CSP complex would phosphorylation of comE, phosphorylated comE and nlmC promoter combined to start the expression of the downstream fluorescent gene GFP. The fluorescence intensity increased gradually as the culture time prolonged. After 24h, there was a significant difference in fluorescence intensity. This result showed that the quorum sensing signaling system in E. coli detector we constructed was functional.

Figure 2 shows the comparison with the control group, certain fluorescence intensity was measured in the engineering bacteria containing the nlmC promoter, indicating that the nlmC promoter has fluorescence leakage. In the future, we shall try to redesign or mutate a new low-leakage promoter based on the nlmC promoter and comE binding domain to achieve high sensitivity and low detection limit detection.