1,139 research outputs found
A master relation defines the nonlinear viscoelasticity of single fibroblasts
Cell mechanical functions like locomotion, contraction and division are
controlled by the cytoskeleton, a dynamic biopolymer network whose mechanical
properties remain poorly understood. We perform single-cell uniaxial stretching
experiments on 3T3 fibroblasts. By superimposing small amplitude oscillations
on a mechanically prestressed cell, we find a transition from linear
viscoelastic behavior to power-law stress stiffening. Data from different cells
over several stress decades can be uniquely scaled to obtain a master-relation
between the viscoelastic moduli and the average force. Remarkably, this
relation holds independently of deformation history, adhesion biochemistry, and
intensity of active contraction. In particular, it is irrelevant whether force
is actively generated by the cell or externally imposed by stretching. We
propose that the master-relation reflects the mechanical behavior of the force
bearing actin cytoskeleton, in agreement with stress stiffening known from
semiflexible filament networks.Comment: 12 pages, 11 figures. Accepted for publication in Biophysical
Journal, scheduled to appear in May 200
A versatile maskless microscope projection photolithography system and its application in light-directed fabrication of DNA microarrays
We present a maskless microscope projection lithography system (MPLS), in
which photomasks have been replaced by a Digital Micromirror Device type
spatial light modulator (DMD, Texas Instruments). Employing video projector
technology high resolution patterns, designed as bitmap images on the computer,
are displayed using a micromirror array consisting of about 786000 tiny
individually addressable tilting mirrors. The DMD, which is located in the
image plane of an infinity corrected microscope, is projected onto a substrate
placed in the focal plane of the microscope objective. With a 5x(0.25 NA) Fluar
microscope objective, a fivefold reduction of the image to a total size of 9
mm2 and a minimum feature size of 3.5 microns is achieved. Our system can be
used in the visible range as well as in the near UV (with a light intensity of
up to 76 mW/cm2 around the 365 nm Hg-line). We developed an inexpensive and
simple method to enable exact focusing and controlling of the image quality of
the projected patterns. Our MPLS has originally been designed for the
light-directed in situ synthesis of DNA microarrays. One requirement is a high
UV intensity to keep the fabrication process reasonably short. Another demand
is a sufficient contrast ratio over small distances (of about 5 microns). This
is necessary to achieve a high density of features (i.e. separated sites on the
substrate at which different DNA sequences are synthesized in parallel fashion)
while at the same time the number of stray light induced DNA sequence errors is
kept reasonably small. We demonstrate the performance of the apparatus in
light-directed DNA chip synthesis and discuss its advantages and limitations.Comment: 12 pages, 9 figures, journal articl
Bead-based assay for spatiotemporal gene expression control in cell-free transcription-translation systems
Cell-free gene expression has applications in synthetic biology, biotechnology and biomedicine. In this technique gene expression regulation plays an important role. Transcription factors do not completely suppress expression while other methods for expression control, for example CRISPR/Cas, often require important biochemical modifications. Here we use an all Escherichia coli-based cell-free expression system and present a bead-based method to instantly start and, at a later stage, completely stop gene expression. Magnetic beads coated with DNA of the gene of interest trigger gene expression. The expression stops if we remove the bead-bound DNA as well as transcribed mRNA by hybridization to bead-bound ssDNA. Our method is a simple way to control expression duration very accurately in time and space
DNA oligomer binding in competition exhibits cooperativity
Binding of two complementary DNA single strands to a double-helix, DNA hybridization, is a sequence specific molecular recognition process that plays important roles in biology and biotechnological applications. In the past much work has been devoted to understand double helix formation, however, DNA binding in complex situations often remains difficult to deal with. Here we use fluorescence anisotropy to assess the binding affinities of DNA oligonucleotide strands that compete for hybridization to the same probe molecule in thermal equilibrium. We find that the ratio of the binding constants in competition can change substantially compared to pairwise assessments. This is a signature of non-trivial interaction among the competitors: the binding microstates of each strand are affected by the presence of the other, but to a different degree. To our knowledge this type of phenomenon is not included in current equilibrium models of oligonucleotide binding. We suggest interactions beyond double helix conformations to cause the observed cooperative behavior. The cooperativity could produce more complex binding phenomena than previously thought
Vehicular Cooperative Maneuvers -- Quo Vaditis?
Vehicles will not only get more and more automated, but they will also
cooperate in new ways. Currently, human-driven vehicles begin to communicate
with each other using vehicle-to-everything technology. Future vehicles will
use communication to share sensor data and even negotiate cooperative
maneuvers. This lets them learn more about the environment and improves traffic
flow and passenger comfort as more predictable maneuvers are likely to lead to
a smoother ride. This paper introduces the most important concepts around
cooperative vehicular maneuvers. We also summarize currently open challenges
and questions to answer before a deployment can begin. Afterward, we give some
perspectives on the further evolution of cooperative maneuvers and beyond.Comment: 8 pages incl. references and author biographies, 4 figures incl.
multiple sub-figure
Hydra Molecular Network Reaches Criticality at the Symmetry-Breaking Axis-Defining Moment
We study biological, multicellular symmetry breaking on a hollow cell sphere as it occurs during hydra regeneration from a random cell aggregate. We show that even a weak temperature gradient directs the axis of the regenerating animal but only if it is applied during the symmetry-breaking moment. We observe that the spatial distribution of the early expressed, head-specific gene k s 1 has become scale-free and fractal at that point. We suggest the self-organized critical state to reflect long range signaling, which is required for axis definition and arises from cell next-neighbor communication
Full incorporation of the noncanonical amino acid hydroxylysine as a surrogate for lysine in green fluorescent protein
The canonical set of amino acids leads to an exceptionally wide range of protein functionality, nevertheless, this
set still exhibits limitations. The incorporation of noncanonical amino acids into proteins can enlarge its functional scope. Although proofreading will counteract the charging of tRNAs with other amino acids than the
canonical ones, the translation machinery may still accept noncanonical amino acids as surrogates and incorporate them at the canonically prescribed locations within the protein sequence. Here, we use a cell-free
expression system to demonstrate the full replacement of L-lysine by L-hydroxylysine at all lysine sites of
recombinantly produced GFP. In vivo, as a main component of collagen, post-translational L-hydroxylysine
generation enables the formation of cross-links. Our work represents a first step towards in vitro production of
(modified) collagens, more generally of proteins that can easily be crosslinke
- …