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Related Concept Videos

Improving Translational Accuracy02:07

Improving Translational Accuracy

Base complementarity between the three base pairs of mRNA codon and the tRNA anticodon is not a failsafe mechanism. Inaccuracies can range from a single mismatch to no correct base pairing at all. The free energy difference between the correct and nearly correct base pairs can be as small as 3 kcal/ mol. With complementarity being the only proofreading step, the estimated error frequency would be one wrong amino acid in every 100 amino acids incorporated. However, error frequencies observed in...
Initiation of Translation02:33

Initiation of Translation

Initiating translation is complex because it involves multiple molecules. Initiator tRNA, ribosomal subunits, and eukaryotic initiation factors (eIFs) are all required to assemble on the initiation codon of mRNA. This process consists of several steps that are mediated by different eIFs.
First, the initiator tRNA must be selected from the pool of elongator tRNAs by eukaryotic initiation factor 2 (eIF2). The initiator tRNA (Met-tRNAi) has conserved sequence elements including modified bases at...
Leaky Scanning02:28

Leaky Scanning

During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA.  Marilyn Kozak discovered that the sequence RCCAUGG (where R stands for...

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Related Experiment Video

Updated: Jun 10, 2026

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics
09:52

An In Vitro Single-Molecule Imaging Assay for the Analysis of Cap-Dependent Translation Kinetics

Published on: September 15, 2020

High-throughput in vitro translation.

Frank Diehl1, Giovanni Traverso

  • 1Sidney Kimmel Comprehensive Cancer Center, The Johns Hopkins Medical Institutions, 1650 Orleans Street, Baltimore, MD 21231, USA and Trinity College, Cambridge CB2 1TQ, U.K.

Discovery Medicine
|August 14, 2010
PubMed
Summary
This summary is machine-generated.

Cell-free protein expression systems, specifically the in vitro transcription and translation (IVTT) assay, enable sensitive detection of disease-related gene mutations. This method is crucial for developing automatable, nonisotopic diagnostic tools for population screening.

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Related Experiment Videos

Last Updated: Jun 10, 2026

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Published on: September 15, 2020

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09:13

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Published on: August 22, 2017

Rapid In Vivo Fixation and Isolation of Translational Complexes from Eukaryotic Cells
14:29

Rapid In Vivo Fixation and Isolation of Translational Complexes from Eukaryotic Cells

Published on: December 25, 2021

Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genetics

Background:

  • Cell-free protein expression systems are vital tools in research and diagnostics.
  • Detecting gene mutations causing premature translation termination is critical for disease diagnosis.

Purpose of the Study:

  • To highlight the utility of in vitro transcription and translation (IVTT) assays for molecular diagnostics.
  • To emphasize the need for sensitive, automatable, and nonisotopic assay systems for population-wide disease screening.

Main Methods:

  • Utilizes the protein truncation test (in vitro synthesized protein assay).
  • Involves purification of genomic RNA or DNA from clinical samples.
  • Employs PCR or RT-PCR with a forward primer containing T7 promoter, Kozak sequence, and start codon for direct IVTT reaction.

Main Results:

  • The described IVTT assay approach facilitates the detection of specific gene mutations.
  • The method is based on established protein truncation test principles.

Conclusions:

  • The in vitro transcription and translation (IVTT) assay offers a promising platform for molecular diagnostics.
  • Further development of IVTT assays is essential for efficient, population-wide disease screening.