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Expression of the Proteus mirabilis recA gene in Bacillus subtilis is directed by its own promoter
Abstract:
The recA gene of Proteus mirabilis (recApm) has been cloned into the PstI site of the Bacillus promoter-probe plasmid pPL603. When present on this plasmid, the recApm1) gene is expressed in B. subtilis under the control of its own transcriptional and translational signals. It is concluded that the high AT-content of the DNA sequence upstream of the -35 region is of decisive importance for the usage of the recApm promoter by the B. subtilis RNA polymerase. The results are discussed in relation to the expression barriers found to exist for genes from gram-negative bacteria in the gram-positive B. subtilis.
Insights
The Proteus mirabilis recA gene (recApm) is expressed in Bacillus subtilis, showing that high AT-content DNA upstream of the -35 region is crucial for promoter recognition by B. subtilis RNA polymerase.
Area of Science:
- Molecular Biology
- Microbiology
- Genetics
Background:
- The recA gene is essential for DNA repair and recombination in bacteria.
- Understanding gene expression across different bacterial species, like Gram-negative Proteus mirabilis and Gram-positive Bacillus subtilis, is key to overcoming expression barriers.
Purpose of the Study:
- To investigate the expression of the recA gene from Proteus mirabilis (recApm) in Bacillus subtilis.
- To identify factors influencing the transcription of Gram-negative genes in a Gram-positive host.
Main Methods:
- Cloning the recApm gene into the Bacillus promoter-probe plasmid pPL603.
- Expressing the recApm gene in B. subtilis under its native regulatory elements.
- Analyzing the role of DNA sequence elements, specifically AT-content, in promoter function.
Main Results:
- The recApm gene was successfully expressed in B. subtilis.
- The high AT-content in the DNA sequence upstream of the -35 region was identified as critical for recApm promoter recognition by B. subtilis RNA polymerase.
Conclusions:
- The AT-rich upstream sequence is a decisive factor for B. subtilis RNA polymerase to utilize the recApm promoter.
- This finding sheds light on the expression barriers encountered when transferring genes from Gram-negative to Gram-positive bacteria.