Fluorosensor proteins to detect specific DNA sequences in living bacteria

Area of Science:

• Molecular Biology
• Biotechnology
• Genetics

Background:

• Existing DNA sensors often struggle to detect native double-stranded DNA (dsDNA) prevalent in physiological conditions.
• There is a need for sensitive and specific methods to monitor dsDNA within living cells.

Purpose of the Study:

• To engineer novel fluorosensor proteins capable of detecting specific dsDNA sequences in living bacteria.
• To create a biosensor for real-time monitoring of target DNA in bacterial hosts.

Main Methods:

• Constructed fusion proteins comprising zinc fingers, a dimerization motif, and a split yellow fluorescent protein (eYFP).
• Utilized Escherichia coli (E. coli) Top10 cells harboring plasmids encoding the fluorosensor proteins and target DNA.
• Induced protein expression and measured fluorescence intensity to assess sensor performance.

Main Results:

• A significant increase in fluorescence was observed in E. coli cells expressing the fluorosensor proteins in the presence of the target DNA sequence.
• Control experiments lacking the target DNA sequence showed minimal fluorescence, confirming sensor specificity.
• The developed fluorosensor proteins successfully detected specific dsDNA within living bacterial cells.

Conclusions:

• The novel fluorosensor proteins effectively detect specific dsDNA sequences in living bacteria.
• This engineered biosensor provides a promising tool for molecular biology and biotechnology applications requiring in vivo DNA detection.