A Novel Selection Marker for Efficient DNA Cloning and Recombineering in E. coli
Figure 6
Cells transformed with larger mfabI plasmids have a growth advantage.
(A) Larger colonies were formed by cells transformed with larger mfabI plasmids on plates. Shown are colonies of DH5α cells transformed by pF2 (i & iv), pF2-DTA-Rosa26 (ii & v), or pF2-DTA-Rosa26-Insert (iii & vi) and grown at 37°C (i, ii & iii) or 32°C (iv, v & vi) on 1 µM triclosan LB plates; insert in each panel is the low-magnification view of the whole plate. (B) Higher cell densities obtained in overnight cultures of larger mfabI plasmid transformants. Two colonies per group were picked from 32°C-grown plates and grew in 1 µM triclosan LB broth at 37°C for 17 hrs before O.D. measurements. *: p<0.05 **: p<0.01 in one-tailed, unpaired t-test (N = 2). Error bars show standard deviations. (C) Plasmid DNA yields from overnight cultures. Plasmid DNA was extracted from cultures used in (B). Error bars show standard deviations (N = 2). (D) Restriction enzyme (XhoI/XbaI)-digested plasmid DNA used in (C). Molar equalized amounts of DNA were digested. All clones showed expected correct digestion patterns (refer to maps in Fig. 5A). M is DNA molecular weight marker lane (Invitrogen 1 kb plus DNA ladder); numbers denote kb bands.