Related Experiment Videos
Localization of genes for the double-stranded RNA killer virus of yeast
Abstract:
The M double-stranded RNA (ds RNA) genome segment of the cytoplasmically inherited killer virus of yeast codes for two polypeptides when denatured and translated in vitro: a previously known 32,000-dalton peptide and a newly discovered 19,000-dalton peptide (NaDodSO4/polyacrylamide gel electrophoresis). An internal 190-base-pair region of the ds RNA is selectively degraded by S1 nuclease treatment at 65 degrees C, resulting in two ds RNA fragments which contain the termini of the original ds RNA. The larger fragment codes for the 32,000-dalton polypeptide and the smaller fragment codes for the 19,000-dalton polypeptide. Thus, the two gene products of M are encoded by distinct regions of this ds RNA.
Insights
The yeast killer virus M ds RNA encodes two distinct proteins from separate regions. S1 nuclease treatment of the ds RNA separates these coding regions, identifying the genes for 32,000-dalton and 19,000-dalton peptides.
Area of Science:
- Molecular Biology
- Virology
- Yeast Genetics
Background:
- The M ds RNA genome of the yeast killer virus is known to encode viral proteins.
- Previous studies identified a 32,000-dalton peptide product.
Purpose of the Study:
- To identify all polypeptide products encoded by the M ds RNA.
- To determine the genetic organization of the M ds RNA genome.
Main Methods:
- In vitro translation of denatured M ds RNA.
- NaDodSO4/polyacrylamide gel electrophoresis for polypeptide analysis.
- S1 nuclease digestion of M ds RNA at 65°C.
- Analysis of ds RNA fragments and their translation products.
Main Results:
- Two polypeptides were identified: a known 32,000-dalton peptide and a newly discovered 19,000-dalton peptide.
- S1 nuclease selectively degraded an internal 190-base-pair region of the M ds RNA.
- This generated two ds RNA fragments, each containing the termini of the original ds RNA.
- The larger fragment encoded the 32,000-dalton polypeptide, and the smaller fragment encoded the 19,000-dalton polypeptide.
Conclusions:
- The M ds RNA genome encodes two distinct polypeptides.
- These two gene products are encoded by separate regions within the M ds RNA molecule.
- This research clarifies the genetic structure and protein-coding capacity of the yeast killer virus M ds RNA.