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Cloning of DNA corresponding to four different measles virus genomic regions
Abstract:
Hybrid plasmids containing sequences corresponding to four different regions of the measles virus genome inserted in pBR322 were obtained by use of polyadenylated 50 S viral RNA as template for reverse transcription. One class of plasmids contains inserts corresponding to the 3' terminal region of the virus genome. The sequence of one of these inserts (605 nucleotides) partially overlaps with the cloned cDNA sequence corresponding to a part of the nucleocapsid protein (N) mRNA (M. Gorecki and S. Rozenblatt (1980). Proc. Natl. Acad. Sci. USA 77, 3686-3690). This insert region shows only one long open reading frame defining the N-terminal part of the nucleocapsid protein. The nucleocapsid protein mRNA starts at about 60 nucleotides from the genome end as revealed by nuclease S1 mapping. Three other classes of plasmid clones contain inserts derived from unidentified regions of the viral genome; they hybridize with viral mRNA species less abundant than those from which cDNA clones have been isolated so far (S. Rozenblatt, C. Gesang, V. Lavie, and F. S. Neumann (1982) J. Virol. 42, 790-797.
Insights
Researchers created hybrid plasmids from measles virus RNA to study its genome. They identified a region encoding part of the nucleocapsid protein, crucial for virus structure and replication.
Area of Science:
- Virology
- Molecular Biology
- Genetics
Background:
- Measles virus is a significant human pathogen.
- Understanding the measles virus genome is crucial for developing antiviral strategies.
- Previous studies have focused on specific viral mRNA species.
Purpose of the Study:
- To construct hybrid plasmids containing measles virus genome fragments.
- To identify and characterize specific regions of the measles virus genome.
- To investigate the nucleocapsid protein (N) mRNA sequence and its genomic origin.
Main Methods:
- Hybrid plasmid construction using reverse transcription of viral RNA.
- Insertion of measles virus genome fragments into pBR322.
- Nucleotide sequencing of plasmid inserts.
- Nuclease S1 mapping to determine mRNA start sites.
Main Results:
- Hybrid plasmids with inserts from four measles virus genome regions were generated.
- One plasmid class contained inserts from the 3' terminal region, including a partial overlap with nucleocapsid protein (N) mRNA.
- A 605-nucleotide insert revealed an open reading frame for the N-terminal part of the nucleocapsid protein.
- Nuclease S1 mapping indicated the nucleocapsid protein mRNA initiates approximately 60 nucleotides from the genome's 3' end.
- Three other plasmid classes contained inserts from unidentified viral genome regions.
Conclusions:
- The study successfully generated hybrid plasmids representing different measles virus genome regions.
- A significant portion of the nucleocapsid protein gene was cloned and characterized.
- The findings provide insights into measles virus gene organization and expression.
- Further investigation of the unidentified genome regions is warranted.