New strategies for isolation of low abundance viral and host cDNAs: application to cloning of the hepatitis E virus and analysis of tissue-specific transcription.
The ability to clone viruses molecularly has led to dramatic advances in our understanding of this diverse group of agents at the molecular level. These insights are critical to the development of experimental strategies for the containment and control of the various viral pathogens that cause hepatitis in man. Knowledge of the genes and gene products have, for example, assisted in the design of expression systems that have been useful for the cell-free expression and production of viral constituents in the development of a subunit (recombinant) vaccine for HBV. Of particular advantage to the experimenter would be the ability to clone the virus directly from the infectious source without the need to resort to biologic amplification systems that require costly investments of money and manpower. This would obviate the need for the time-intensive development of a tissue culture propagation or the expensive cost of developing an animal model. The methods described here have applications to the discovery and isolation of low abundance transcripts from host (cellular) genes. These host encoded products may have important roles in virus replication and certainly by themselves constitute a growing area of study. The direct selection protocol has already proven its value in the isolation of novel (previously undescribed) gene sequences from complex cellular sources. The application of these methodologies should, at the single cell level, aid in the delineation of those important host-encoded gene products that are critical for the efficient in vitro propagation of hepatotropic viruses. The identification of these low abundance cellular genes will elucidate the biologic interplay between host and obligate intracellular parasite (virus) and potentially lead to the development of new strategies for virus control that take into consideration the role of the cell as host.
Reyes
G R
GR
Department of Molecular Virology, Genelabs Incorporated, Redwood City, California.
Bradley
D W
DW
Lovett
M
M
D001483:Base Sequence; D003001:Cloning, Molecular / Q000379:methods; D004279:DNA, Viral / Q000235:genetics* / Q000302:isolation & purification; D016752:Hepatitis E virus / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D014018:Tissue Distribution; D014158:Transcription, Genetic
D004279:DNA, Viral