Homologous Gene Replacement in Physarum

Overview

Journal Genetics

Publisher Oxford University Press

Specialty Genetics

Date 1995 Jan 1

PMID 7705620

Citations 3

Authors

T G Burland

D Pallotta

Affiliations

Soon will be listed here.

Abstract

The protist Physarum polycephalum is useful for analysis of several aspects of cellular and developmental biology. To expand the opportunities for experimental analysis of this organism, we have developed a method for gene replacement. We transformed Physarum amoebae with plasmid DNA carrying a mutant allele, ardD delta 1, of the ardD actin gene; ardD delta 1 mutates the critical carboxy-terminal region of the gene product. Because ardD is not expressed in the amoeba, replacement of ardD+ with ardD delta 1 should not be lethal for this cell type. Transformants were obtained only when linear plasmid DNA was used. Most transformants carried one copy of ardD delta 1 in addition to ardD+, but in two (5%), ardD+ was replaced by a single copy of ardD delta 1. This is the first example of homologous gene replacement in Physarum. ardD delta 1 was stably maintained in the genome through growth, development and meiosis. We found no effect of ardD delta 1 on viability, growth, or development of any of the various cell types of Physarum. Thus, the carboxy-terminal region of the ardD product appears not to perform a unique essential role in growth or development. Nevertheless, this method for homologous gene replacement can be applied to analyze the function of any cloned gene.

Citing Articles

The one-kilobase DNA fragment upstream of the ardC actin gene of Physarum polycephalum is both a replicator and a promoter.

Pierron G, Pallotta D, Benard M Mol Cell Biol. 1999; 19(5):3506-14.

PMID: 10207074 PMC: 84143. DOI: 10.1128/MCB.19.5.3506.

Chlamydomonas: the cell and its genomes.

Lefebvre P, Silflow C Genetics. 1999; 151(1):9-14.

PMID: 9872944 PMC: 1460453. DOI: 10.1093/genetics/151.1.9.

Mapping of a replication origin within the promoter region of two unlinked, abundantly transcribed actin genes of Physarum polycephalum.

Benard M, Lagnel C, Pallotta D, Pierron G Mol Cell Biol. 1996; 16(3):968-76.

PMID: 8622700 PMC: 231079. DOI: 10.1128/MCB.16.3.968.

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