J. Sambrook

Semliki Forest virus temperature-sensitive mutants: Isolation and characterization

Abstract The normal replication of Semliki Forest virus involves RNA synthesis followed by the synthesis of two structural proteins (nucleocapsid and membrane). Temperature-sensitive mutants blocked at different steps of the growth cycle have been isolated from Semliki Forest virus by chemical mutagenesis. A group of the mutants defective in RNA synthesis (RNA − mutants) were prevented from producing viral antigen in cells infected at the restrictive temperature and also were unable to inhibit host cell RNA synthesis. The factor affecting viral RNA synthesis is expressed early in the growth cycle. Other mutants were defective in functions subsequent to viral RNA synthesis (RNA + mutants). Some were defective in the production of nucleocapsids only; others failed to make membrane only. A few RNA + mutants made both structural proteins, but not infectious virus, and may be defective in a maturation function.

Conditional lethal mutants of rabbitpox virus: II. Mutants (p) that fail to multiply in PK-2a cells

Abstract The behaviour of 34 mutants of rabbitpox virus (RP) that fail to multiply in PK-2a cells (p mutants) has been examined. Mutants differ in the stage at which multiplication is blocked in PK-2a cells, as judged by tests for reactivating capacity, viral DNA synthesis, and viral antigen production. Some of the mutants can be rescued from suicide in PK-2a cells by simultaneous infection with RPp+ viruses. No complementation between the p mutants was found. All of them produce white (u) pocks on the chorioallantoic membrane, and recombination experiments show that the p mutants compose one linkage group of the recombination matrix of u mutants. The linkage map of the u mutants has been extended to include a total of 51 mutants.

Conditional lethal mutants of rabbitpox virus: I. Isolation of host cell-dependent and temperature-dependent mutants

Abstract Growth of a cloned stock of rabbitpox virus in the presence of the mutagens 5-bromodeoxyuridine and 2-aminopurine resulted in a reduced yield of infectious virus. Temperature- and host-dependent mutants could be detected in these yields. Methods were devised for the large-scale isolation of both types of conditional lethal mutants and for enriching the yield of temperature-sensitive mutants by the use of the viral inhibitor isatin β-thiosemicarbazone.