Related Experiment Video
Updated: Aug 8, 2026
Immunofluorescent Labeling of Plant Virus and Insect Vector Proteins in Hemipteran Guts
Published on: May 14, 2021
Identification and characterization of the gene encoding the major structural protein of insect iridescent virus type
1NERC Institute of Virology and Environmental Microbiology, Oxford, United Kingdom.
Abstract:
The major structural protein (MSP--apparent molecular weight 49,000) of insect iridescent virus type 22 (isolated from blackflies--Simulium spp.) was resolved from disrupted, purified virus particles by SDS-PAGE and transferred to nitrocellulose by Western blotting. The portion of the blot containing the MSP was identified and excised. Tryptic peptides, generated by digestion in situ, were purified by HPLC. Three of these peptides were sequenced and an oligonucleotide gene probe was designed using one of them. A SalI clone of IV22 DNA was identified as MSP-specific by hybridization. DNA from this and an overlapping DNA clone was sequenced and a large open reading frame was positively identified as the MSP coding sequence by comparison with the tryptic peptide sequences. The molecular weight of the predicted protein product of this gene is 51,993, comparable with the apparent weight obtained by SDS-PAGE. In infected Spodoptera frugiperda (Sf) cells MSP is synthesized from 12 hr postinfection onwards. The identification of this gene and analysis of its expression opens the way to elucidating the control of late gene expression in an insect iridescent virus.
Insights
Researchers identified the gene for the major structural protein (MSP) of insect iridescent virus type 22. This finding aids in understanding how the virus controls gene expression during infection.
Area of Science:
- Virology
- Molecular Biology
- Insect Pathology
Background:
- Insect iridescent viruses (IIVs) are significant pathogens affecting various insect species.
- Understanding the molecular mechanisms of IIV replication, particularly gene expression, is crucial for controlling insect populations.
Purpose of the Study:
- To identify and characterize the gene encoding the major structural protein (MSP) of insect iridescent virus type 22 (IIV-22).
- To analyze the expression pattern of the MSP gene in infected insect cells.
Main Methods:
- Protein purification and Western blotting to isolate and identify the MSP.
- Peptide sequencing (HPLC) and oligonucleotide probe design.
- DNA cloning, sequencing, and Southern hybridization to identify the MSP gene.
- Analysis of gene expression in infected Spodoptera frugiperda cells.
Main Results:
- The major structural protein (MSP) of IIV-22 was isolated and its N-terminal sequence determined.
- A specific DNA clone containing the MSP gene was identified through hybridization.
- The MSP coding sequence was identified within the viral genome, with a predicted protein product of 51,993 Da.
- MSP synthesis was observed in infected cells starting 12 hours post-infection.
Conclusions:
- The gene encoding the major structural protein of IIV-22 has been successfully identified and characterized.
- This identification provides a foundation for further studies into the regulation of late gene expression in insect iridescent viruses.
- The findings contribute to understanding viral pathogenesis and developing potential control strategies for insect pests.
