Cloning of the two essential yeast genes, PRP6 and PRP9, and their rapid mapping, disruption and partial sequencing using a linker insertion strategy. |
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Authors: | Pierre Legrain Christine Chapon Etienne Schwob Robert Martin Michael Rosbash and Bernard Dujon |
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Institution: | (1) Unité de Génétique Moléculaire des Levures, Department de Biologie Moleculaire, Institut Pasteur, 28 rue du Docteur Roux, F-75724 Paris Cedex 15, France;(2) Institut de Biologie Moleculaire et Cellulaire, 15 rue Descartes, F-67084 Strasbourg, France;(3) Department of Biology, Brandeis University, 2254 Waltham, MA, USA |
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Abstract: | Summary In the yeast Saccharomyces cerevisiae, some thermosensitive (ts) mutants have been shown to be impaired in pre-mRNA splicing (prp mutants). From a yeast genomic library, we have isolated plasmids that complement prp6 or prp9 is mutations. These plasmids also complement the is growth defect of additional independent mutants identified as new prp6 and prp9 is alleles, indicating that the cloned DNAs encode PRP6 and PRP9 genes, respectively. Here, we describe the restriction maps of these loci which are localized on chromosome II and IV, respectively. The limits of open reading frames (ORFs) within the cloned inserts have been determined using a linker insertion strategy combined with the is complementation assay. Double-strand DNA sequencing was also performed directly on the yeast expression vector from the inserted linkers. Gene disruption experiments demonstrate that both genes are essential for viability. |
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Keywords: | Saccharomyces cerevisiae Splicing prp mutants Linker insertion |
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