Lecture

RNA-based Probes: Engineering GFP-mimics

Graph Chat

Chat with Graph Search

Ask any question about EPFL courses, lectures, exercises, research, news, etc. or try the example questions below.

DISCLAIMER: The Graph Chatbot is not programmed to provide explicit or categorical answers to your questions. Rather, it transforms your questions into API requests that are distributed across the various IT services officially administered by EPFL. Its purpose is solely to collect and recommend relevant references to content that you can explore to help you answer your questions.

Description

This lecture delves into the engineering and application of RNA-based probes, focusing on GFP-mimics. Starting with a refresher on fluorescent proteins like GFP, it explores the chromophore maturation process, autocatalytic mechanisms, and key structural features. The lecture then discusses the evolution of Spinach as a GFP-mimic for RNA tracking, comparing it with GFP fusion proteins. It covers the structure and mechanism of RNA mimics of GFP, highlighting key papers and the effects of deprotonation on the GFP chromophore. The session concludes with discussions on SELEX technology, binding considerations, and the utility of Spinach in RNA imaging.

Official source

Related concepts (26)

Green fluorescent protein

The green fluorescent protein (GFP) is a protein that exhibits bright green fluorescence when exposed to light in the blue to ultraviolet range. The label GFP traditionally refers to the protein first isolated from the jellyfish Aequorea victoria and is sometimes called avGFP. However, GFPs have been found in other organisms including corals, sea anemones, zoanithids, copepods and lancelets. The GFP from A. victoria has a major excitation peak at a wavelength of 395 nm and a minor one at 475 nm.

Fluorescent tag

In molecular biology and biotechnology, a fluorescent tag, also known as a fluorescent label or fluorescent probe, is a molecule that is attached chemically to aid in the detection of a biomolecule such as a protein, antibody, or amino acid. Generally, fluorescent tagging, or labeling, uses a reactive derivative of a fluorescent molecule known as a fluorophore. The fluorophore selectively binds to a specific region or functional group on the target molecule and can be attached chemically or biologically.

RNA

Ribonucleic acid (RNA) is a polymeric molecule that is essential for most biological functions, either by performing the function itself (Non-coding RNA) or by forming a template for production of proteins (messenger RNA). RNA and deoxyribonucleic acid (DNA) are nucleic acids. The nucleic acids constitute one of the four major macromolecules essential for all known forms of life. RNA is assembled as a chain of nucleotides.

Ribosomal RNA

Ribosomal ribonucleic acid (rRNA) is a type of non-coding RNA which is the primary component of ribosomes, essential to all cells. rRNA is a ribozyme which carries out protein synthesis in ribosomes. Ribosomal RNA is transcribed from ribosomal DNA (rDNA) and then bound to ribosomal proteins to form small and large ribosome subunits. rRNA is the physical and mechanical factor of the ribosome that forces transfer RNA (tRNA) and messenger RNA (mRNA) to process and translate the latter into proteins.

Messenger RNA

In molecular biology, messenger ribonucleic acid (mRNA) is a single-stranded molecule of RNA that corresponds to the genetic sequence of a gene, and is read by a ribosome in the process of synthesizing a protein. mRNA is created during the process of transcription, where an enzyme (RNA polymerase) converts the gene into primary transcript mRNA (also known as pre-mRNA). This pre-mRNA usually still contains introns, regions that will not go on to code for the final amino acid sequence.

Related lectures (37)

Fluorescence in Microscopy: Introduction & Labeling Techniques

Covers the introduction to fluorescence in microscopy, labeling techniques, specificity of markers, multi-color imaging, and factors affecting fluorescence properties.

Chemical Biology Tools: Transcriptome Interrogation

Covers chemical biology tools for RNA study and explores the applications of GFP in biosciences.

Tools for Translatome Analysis

Explores tools for studying the translatome, including mRNA translation and genetic code expansion.

Fluorescence in Microscopy: Important Concepts

Introduces fundamental concepts of fluorescence in microscopy, covering terms like excitation and emission spectra, Stokes shift, quantum efficiency, brightness, and photobleaching.

Nucleic Acids: Structure and Function

Explores the structure and function of nucleic acids, focusing on DNA and RNA, including base pairing rules and the double helix structure.

Related concepts (26)

Green fluorescent protein