Rna Duplex Unwinding
Rna Duplex Unwinding - Related Videos
04:10
Helicase Activity Measurement of a Target Protein Using Biotin-Labeled RNA Duplexes
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2025
In this video, we demonstrate the procedure to determine the helicase activity of a target protein to unwind the biotin-labeled dsRNA substrate. The activity of the enzyme was identified by analyzing the electrophoretic mobility shift, followed by a chemiluminescence assay using chemiluminescent enzyme-conjugated streptavidin.
00:58
Eukaryotic RNA Polymerases
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2025
RNA Polymerase (RNAP) is conserved in all animals, with bacterial, archaeal, and eukaryotic RNAPs sharing significant sequence, structural, and functional similarities. Among the three eukaryotic RNAPs, RNA Polymerase II is most similar to bacterial RNAP in terms of both structural organization and folding topologies of the enzyme subunits. However, these similarities are not reflected in their mechanism of action. All three eukaryotic RNAPs require specific transcription factors, of which the...
07:37
Single-Molecule Real-Time Visualization of DNA Unwinding by CMG Helicase
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Cited by 1 •
2024
This protocol demonstrates performing a single-molecule assay for live visualization of DNA unwinding by CMG helicase. It describes (1) preparing a DNA substrate, (2) purifying fluorescently labeled Drosophila melanogaster CMG helicase, (3) preparing a microfluidic flow cell for total internal reflection fluorescence (TIRF) microscopy, and (4) the single-molecule DNA unwinding assay.
11:42
Iterative Optimization of DNA Duplexes for Crystallization of SeqA-DNA Complexes
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2012
Crystal structure of protein–DNA complexes can provide insight into protein function, mechanism, as well as, the nature of the specific interaction. Here, we report how to optimize the length, sequence and ends of duplex DNA for co-crystallization with Escherichia coli SeqA, a negative regulator of replication initiation.
08:45
Duplex Digital PCR for Simultaneous Quantification of Dual Genetic Markers
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2025
In this video, we demonstrate the duplex digital PCR (ddPCR) technique — a modification of the traditional PCR technique that is useful in detecting two different genetic markers simultaneously. A single PCR reaction is partitioned into nanoliter-sized emulsified droplets that are independently amplified, and the detection of differently colored fluorescence amplification signals from the fraction of droplets is used to compute the initial concentration of the target sequences.
FAQs
RNA helicases and related factors couple ATP hydrolysis to structural remodeling of RNA. The released energy helps disrupt base pairing within paired strands or folded double-stranded regions, allowing previously inaccessible sequences to become available. This energy-dependent mechanism is central to regulating when RNA structures change during cellular use and experimental analysis.
RNA helicases provide the primary remodeling activity, while accessory proteins can help determine how and when a target RNA is engaged. Their combined action contributes to the specificity and timing of RNA duplex unwinding rather than producing indiscriminate strand separation. These features are important for controlled regulation of RNA structure and function.
Timing and specificity determine which RNA structures are remodeled and at what stage they become accessible. That control can influence post-transcriptional regulation, including changes associated with translation, RNA processing, or RNA interference. Studying these properties helps connect a molecular remodeling event with its downstream biological role.
Researchers examine the process with biochemical assays and engineered constructs designed to test RNA structure and remodeling activity. These approaches can focus on how a factor acts on a selected RNA arrangement or how altered constructs affect analysis. Together, they provide experimental ways to study mechanism, timing, and specificity.
The process is relevant to RNA interference, translation, RNA processing, and virus replication. In each context, changing RNA structure can alter access to genetic information or influence how RNA is handled. Comparing unwinding behavior across these applications helps researchers relate a shared molecular mechanism to distinct forms of post-transcriptional regulation.
These studies can show how structural accessibility changes the behavior of an RNA molecule and how remodeling is connected to biological regulation. Results from biochemical assays or engineered constructs may clarify the role of specific timing and targeting properties. Such findings also support methods for experimentally manipulating RNA structure and function.