Dimension fastscan afm system

A 703 base pair (bp) region from the gltA promoter was gel-purified using the QIAquick Gel Extraction Kit (Qiagen) by using primer pairs gltA F and gltA R for S. Typhimurium (-689 bp to + 14 bp) and for E. coli (-694 bp to + 9 bp), respectively. A glutaraldehyde-modified mica surface was prepared as described in Chakraborty et al. (2017) (link). Ten nanograms of the gltA regulatory region was incubated with 30 nM OmpR for 15 min at RT. This mixture was then deposited on the mica for 15 min. Images were acquired on a Bruker Dimension FastScan AFM system using the tapping mode with a silicon nitride cantilever (FastScan C, Bruker). Raw AFM images were processed using Gwyddion software¹.

Glutaraldehyde-modified mica surface was prepared as decribed previously in (Liu et al., 2010 (link)) and references therein. A 755 bp sequence upstream to the +1 start site (Gerstel et al., 2003 (link)) of csgD was amplified and gel purified. This fragment also harbored a SsrB-specific site (Feng et al., 2004 (link); Walthers et al., 2007 (link)), TTATAAT sequence (Figure 6—figure supplement 4). A typical 50 μl reaction contained 10 ng of this DNA (755 bp of the csgD regulatory region) mixed with an appropriate amount of SsrB, SsrBc or D56A SsrB and incubated for 15 min at room temperature. This mixture was then deposited on glutaraldehyde-modified mica for 15 min. Images were acquired on a Bruker Dimension FastScan AFM system using the tapping mode with a silicon nitride cantilever (FastScan C, Bruker). Raw AFM images were processed using Gwyddion software (http://gwyddion.net/). The bending angle was analysed using a home-written Matlab code as described in the Supplementary methods.

4 μM ADP-Actin was polymerized first in F buffer (1 X KME in 1X G buffer) for 1 hour. Actin filaments were diluted 5 times in 30 μL F buffer with 100 nM proteins and incubated for 30 minutes.
For topography imaging of actin filaments, 0.8 μM of actin filaments in 30 μL F buffer containing indicated protein was deposited on 0.01% poly-L-lysine coated glass coverslips and incubated for extra 30 minutes before image. The coverslips were rinsed three times in F buffer to remove unattached actin filaments and 30 μl of F buffer was added to the sample immediately to prevent drying. A silicon nitride probe (Scanasyst Fluid+, Bruker) with nominal tip radius of 2 nm was used. All samples were imaged with a Bruker Dimension FastScan AFM system and Nanoscope V controller operating under ScanAsyst fluid mode. AFM images were acquired at a maximum scan rate of 2.5 Hz and 256 × 256 data points per image. Raw AFM data were analyzed using Bruker Nanoscope Analysis 1.90 software.

The SDS molecules were dissolved in water (0.6 wt%), and multilayer graphene flakes were transferred onto silicon substrates. We used this concentration because only the SDS aggregations were observed at higher concentration (2.0 wt%). An AFM probe tip and multilayer graphene on a silicon substrate were placed in a water droplet, into which the SDS solution could be slowly injected at a rate of 75 μL/h using a microsyringe pump. Consequently, the concentration of SDS molecules reached approximately 1.36 mM. All of the measurements were performed at room temperature.
The AFM measurements were carried out in “tapping in fluid” mode using a Dimension FastScan AFM system (Bruker, Santa Babara, CA, USA). In the examination of the effect of AFM tip scanning, the AFM images (Fig. 1b,c, and d) were obtained with a tip–sample force (f_ts) of 20 pN (see Supplementary Note 6 for the detailed estimation method) and a scan velocity (v_scan) of 4.3 μm/s. The intense tip scanning for generating ribbons was performed in 15 min with an f_ts of 30 pN and a v_scan of 55.7 μm/s. In independent 291 scanning experiments, the scanning time was set as about 20 min. In the demonstration of unidirectional ribbon formation, the AFM images (Fig. 4a,b) were collected with an f_ts of 27 pN and a v_scan of 1.3 μm/s. The unidirectional ribbon formation was achieved with an f_ts of 42 pN and a v_scan of 32.4 μm/s.