Part:BBa_K4849000

P_rbcL1C constitutive Synechocystis promoter

 

Description

P_rbcL is the promoter for the large subunit of Rubisco in Synechocystis sp. strain PCC 6803. Huang et al. (2010) constructed several variants of this promoter, some of which are shown in Figure 1, by PCR using genomic DNA from Synechocystis as template.

Figure 1. The upstream region (−277 to −1, relative to the ATG) of the rbcL gene in Synechocystis sp. strain PCC 6803. a: predicted NtcA-binding site, b: putative −10 element, c: putative −35 element, d: −10 element, e: putative ribosome-binding site (RBS), f: start codon ATG of the rbcL gene. The variants ‘1’ (PrbcL1A, PrbcL1B, PrbcL1C) lack the predicted NtcA-binding site and the AT-rich region upstream (−277 to −215, relative to the ATG of the rbcL gene). The variants ‘C’ (PrbcL1C, PrbcL2C) lack part of the 3′-end (−48 to −18, relative to the ATG of the rbcL gene), and the RBS (black box) is attached by standard assembly, resulting in an additional 8-bp scar (TACTAGAG, grey box). Taken from Huang et al. (2010).

 

Huang et al. (2010) also tested the strengths of these promoters by producing GFPmut3B reporter constructs and measuring the fluorescence of the reporter protein GFPmut3B (Figure 2 and Table 1). The specific promoter activities of the promoters P_rbcL2A, P_rbcL2C, P_rbcL1A, PrbcL1B, P_rbcL1C and P_rbcL2B in relation to P_rnpB, which as a weak constitutive Synechocystis promoter of the of the rnaseP gene, were 19, 10, 9, 5, 4 and 1, respectively (Figure 2 and Table 1).

 

Figure 2. Specific activities of the six rbcL promoter variants in comparison to P_rnpB and P_trc1O in Synechocystis. The activities were measured by means of GFPmut3B fluorescence and divided by the absorbance of the cultures at 750 nm. The data represent mean ± SD of triple measurements of three independent cultivations. Taken from Huang et al. (2010).

 

Table 1. Specific promoter activities in Synechocystis from Figure 2 in units relative to P_rnpB. Taken from Huang et al. (2010).

 

References

 

Huang, H.H., Camsund, D., Lindblad, P. and Heidorn, T., 2010. Design and characterization of molecular tools for a synthetic biology approach towards developing cyanobacterial biotechnology. Nucleic acids research, 38(8), pp.2577-2593.

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]