ABSTRACT
It has been hypothesized that the ribosome gains additional fidelity during protein translation by probing structural differences in tRNA species. We measure the translocation kinetics of different tRNA species through â¼3 nm diameter synthetic nanopores. Each tRNA species varies in the time scale with which it is deformed from equilibrium, as in the translocation step of protein translation. Using machine-learning algorithms, we can differentiate among five tRNA species, analyze the ratios of tRNA binary mixtures, and distinguish tRNA isoacceptors.
Subject(s)
Nanopores , Protein Biosynthesis , RNA, Transfer/chemistry , Binding Sites , Electrophoresis , Kinetics , Machine Learning , RNA, Transfer/genetics , Ribosomes/chemistry , Ribosomes/geneticsABSTRACT
Previous measurements of the electronic conductance of DNA nucleotides or amino acids have used tunnel junctions in which the gap is mechanically adjusted, such as scanning tunneling microscopes or mechanically controllable break junctions. Fixed-junction devices have, at best, detected the passage of whole DNA molecules without yielding chemical information. Here, we report on a layered tunnel junction in which the tunnel gap is defined by a dielectric layer, deposited by atomic layer deposition. Reactive ion etching is used to drill a hole through the layers so that the tunnel junction can be exposed to molecules in solution. When the metal electrodes are functionalized with recognition molecules that capture DNA nucleotides via hydrogen bonds, the identities of the individual nucleotides are revealed by characteristic features of the fluctuating tunnel current associated with single-molecule binding events.
Subject(s)
DNA , Microscopy, Scanning Tunneling/instrumentation , Nucleotides , DNA/chemistry , Electric Conductivity , Electrodes , Hydrogen Bonding , Microfluidic Analytical Techniques , Nucleotides/chemistry , Palladium/chemistry , Silicon/chemistryABSTRACT
We present a scripting toolkit for the acquisition and analysis of a wide variety of imaging data by integrating the ease of use of various programming environments such as LABVIEW, IGOR PRO, MATLAB, SCILAB, and others. This toolkit is designed to allow the user to quickly program a variety of standard microscopy components for custom microscopy applications allowing much more flexibility than other packages. Included are both programming tools as well as graphical user interface classes allowing a standard, consistent, and easy to maintain scripting environment. This programming toolkit allows easy access to most commonly used cameras, stages, and shutters through the Micromanager project so the scripter can focus on their custom application instead of boilerplate code generation.
