DSF biosensor and the Xcc DSF overproduction mutant
ΔrpfC, methods for DSF extraction and purification were estab-
lished [5, 6].
Biosensor-independent methods for DSF extraction and purifi-
cation were also established [2, 7]. In order to improve detection
sensitivity, a quantitative detection of DSF family signals via LC-MS
was developed [8]. The condensed crude extracts fromXccsuper-
natants were separated by an ultra-performance liquid chro-
matographic (UPLC) system on a C 18 reverse-phase column,
followed by negative electrospray ionization (ESI) and detection by
a time-of-flight mass spectrometry (TOF-MS) analysis. As a result,
the minimal detectable level of DSF or BDSF was reduced to as low
as 1μM. This allowed fast and convenient determination of DSF and
BDSF levels in bacterial cultures and reaction mixtures [8, 9].2 Materials
Prepare all solutions using ultrapure water (prepared by purifying
deionized water, to attain a sensitivity of 18 MΩ-cm at 25C) and
analytical grade reagents. Prepare and store all reagents at room
temperature (unless indicated otherwise). Diligently follow all
waste disposal regulations when disposing waste materials.Fig. 1HPLC chromatogram of ethyl acetate extract of the culture supernatant of
the DSF hyperproduction mutantΔrpfCofXccstrain XC1. (a) Four members of
DSF family QS signals are detected in the supernatant ofΔrpfC, among which
DSF and BDSF are the major signal molecules. (b) The chemical structures of
DSF, BDSF, CDSF, and IDSF98 Lian Zhou et al.