Final answer:
To confirm a nonsense mutation, synthesize and purify RNA, then use sequencing, denaturing urea PAGE, and UV spectrometry to verify the mutation and assess its impact on the RNA product.
Step-by-step explanation:
To confirm a nonsense mutation in which an amino acid coding codon has been converted to a stop codon, resulting in the premature termination of protein synthesis, several steps can be taken. After precipitating and transcribing the DNA template to produce the mutant toxin sensor RNA, in vitro, you would typically purify the synthesized RNA. Confirmation of the mutation can be achieved using sequencing to show the presence of the premature stop codon in the RNA or the translated protein.
Additionally, to evaluate the mutant RNA product, denaturing urea PAGE is commonly used to determine its purity, while the RNA concentration can be ascertained using UV spectrometry. This analysis not only confirms the presence of the mutation but also assesses its impact on the RNA product.
Moreover, phenotypic analysis of the protein product, which will be truncated due to the mutation, can further validate the functional implications of the nonsense mutation. Together, these methods provide a comprehensive confirmation of the nonsense mutation and its effects.