Replacements and knockout strains are extremely valuable experimental tools, including the testing the indispensability of cloned genes and isolation of mutants after random mutagenesis of cloned genes. USA 83, 4369-4373. USA 89, 9196-9200. Germline and somatic transformation of mating Tetrahymena thermophila by particle bombardment. Genetics 146, 135-147.
Hours indicate time after mixing of starved cells to initiate conjugation.
Selection for complete gene replacement by phenotypic assortment enables the distinction between essential and non-essential genes. The initial transformant contains one transformed gene which has been disrupted with an expression cassette containing a drug-selectable marker. Its clonal progeny are serially transferred to higher drug concentrations thus killing cells with fewer copies of the selectable markers. All arrows indicate multiple generations. The genotype of the Mpr locus in Round I exconjugants was not determined.