Cloning of abundant mRNA species present during conjugation of Tetrahymena thermophila: identification of mRNA species present exclusively during meiosis

Abstract

A cDNA library was constructed by using as a template the RNA present during the meiotic prophase of Tetrahymena thermophila. Clones containing cDNA sequences homologous to moderately abundant to abundant transcripts were detected by colony hybridization and confirmed by hybridizing purified cDNA plasmids on filters with labeled RNA probes. Eighteen clones were isolated, and the sizes of their cDNA inserts were determined. Cross-hybridization of individual cDNA plasmid pairs showed that each of these clones contained cDNA that was homologous to one of eight different RNA transcripts. The sizes of the eight RNA transcripts and the stages of the T. thermophila life cycle during which they were present were determined by hybridizing nick-translated cDNA probes against denatured, electrophoresed RNA from various stages. Clones were identified that contained sequences homologous to RNAs present only in early conjugation (meiosis); other clones contained sequences homologous to RNAs which were abundant during conjugation but present at other stages as well. One clone contained a sequence homologous to an RNA that was abundantly present only in nongrowing cells.