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An antisense approach to phenotype-based gene cloning in Tetrahymena.

ABSTRACT: We report a pioneering approach using Tetrahymena thermophila that permits rapid identification of genes based on their null or hypomorphic phenotypes. This technique involves cell transformation with a library of plasmids that encode 26S ribosomal subunits containing short insertions. The insertions correspond to antisense sequences for a large number of genes. The majority of cells each acquires a single antisense sequence, which silences a single genomic locus. Because the insertion site within the ribosomal sequence is known, the silenced gene is easily amplified. We demonstrate that this approach can be used to identify genes required for dense core granule exocytosis.

SUBMITTER: Chilcoat ND

PROVIDER: S-EPMC37500 | biostudies-literature | 2001 Jul

REPOSITORIES: biostudies-literature

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An antisense approach to phenotype-based gene cloning in Tetrahymena.

Chilcoat N D ND Elde N C NC Turkewitz A P AP

Proceedings of the National Academy of Sciences of the United States of America 20010703 15

We report a pioneering approach using Tetrahymena thermophila that permits rapid identification of genes based on their null or hypomorphic phenotypes. This technique involves cell transformation with a library of plasmids that encode 26S ribosomal subunits containing short insertions. The insertions correspond to antisense sequences for a large number of genes. The majority of cells each acquires a single antisense sequence, which silences a single genomic locus. Because the insertion site with ...[more]

PMID: 11438705

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