Bink Control Background 8 __FULL__
LINK ===> https://urluso.com/2tcEad
Reversing the competitive disadvantage of the B. cenocepacia WT strain and conferring it to the BinK mutant was an essential step for the ability of the BinK mutant to colonize the squid. The BinK mutation that gives it this advantage is monomorphic for a B. cenocepacia-specific insertion in the array of Tn21-like transposons that occur between the V. fischeri fliK and kdpA flagellar operons. This insertion, after transposition, was found by performing PCR on the mutant background and determining which amplifiïers were present; the insertion caused a 55 bp deletion, with a unique 50 bp insertion, that caused a frame-shift and translated a truncated version of TnsD, the protein product of which is required for transposition (26). Because this insert is known to be fixed in population samples from geographically-diverse sources (25, 27), and the fliK operon was similarly found to be associated with a Tn21-like element in a study of tnphoA mutant isolates from the Baltic Sea (28, 29), the Tn21-like transposon was termed qseC, and the Tn21-like insertion termed qseC’. At all three character intervals (evenness, shoot formation, and bioluminescence), BinK mutant colonies were larger and more sustained, and thus, more competitive than those formed on the other treatments, suggesting that competition for space on the surface may be one of the ways that BinK promotes colonization (22). We likewise find that the mutant is more competitive during live squid bacterial outgrowth assays than the WT strain; we have also carried out tandem mass spectrometry experiments to show that there is a 40% reduction in expression of qseC’. A qseC’ mutant was generated by allelic replacement of the wild-type allele with a recA’ mutant to ensure a correct chromosomal copy of qseC’. In the absence of qseC’, the BinK mutant form more colonies on the squid surface but not as many colonies as the WT strain. d2c66b5586