Injection of total genomic DNA and restoration of wild type phenotype in trichocyst non-discharge mutant TND of Paramecium caudatum

Abstract

To restore wild type phenotypes in mutants controlled by recessive genes, microinjection of whole genomic DNA into recipient macronucleus in Paramecium caudatum was performed. Donor DNA carrying three wild-type genetic markers (exocytosis, behavior and mating type) was introduced into a recipient cell which was a triple recessive homozygote for the corresponding genes. Of the three phenotypes, a trichocyst non-discharge mutant TND converted to wild type by microinjection of not only total genomic DNA but also fractionated restriction fragments, although the restored phenotype was not complete. This transformation was more efficient when injection was performed by using the recipient cells of S phase, compared with the cells of G1 phase. This successful restoration of wild type phenotype gives us hope for cloning mutated genes, especially when no information about gene products is available. We discuss the advantage of this total genomic DNA injection and the possibility of target gene cloning using this strategy.