title: Gene targeting by RNAi-mediated knockdown of potent DNA ligase IV homologue in the cellulase-producing fungus Talaromyces cellulolyticus.
abstract: The genome of the cellulase-producing fungus Talaromyces cellulolyticus (formerly Acremonium cellulolyticus) was screened for a potent DNA ligase IV gene (ligD homologue). Homologous recombination efficiency in T. cellulolyticus is very low. Therefore, suppression of a non-homologous end-joining system was attempted to enable specific gene knockouts for molecular breeding. The transcript levels of ligD homologue were 0.037 of those of the parental YP-4 strain in the Li20 transformant carrying the RNAi construct targeting the ligD homologue. Transformation of the hairpin-type RNAi vector into T. cellulolyticus could be useful in fungal gene knockdown experiments. Cellulase production and protein secretion were similar in the parental YP-4 strain and the Li20 transformant. Knockout transformation of ligD homologue using the Li20 transformant led to 23.1 % double crossover gene targeting. Our results suggest that the potent DNA ligase IV gene of T. cellulolyticus is related to non-homologous end joining and that the knockdown of the ligD homologue is useful in gene targeting.
authors.affiliation: Division of Applied Biosciences, Graduate School of Agriculture, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto, 606-8502, Japan.
authors.forename: Koutarou
authors.initials: K
authors.lastname: Hayata
authors.forename: Seiya
authors.initials: S
authors.lastname: Asada
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Fujii
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Inoue
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Ishikawa
authors.forename: Shigeki
authors.initials: S
authors.lastname: Sawayama
mesh_terms: D002482:Cellulose / Q000096:biosynthesis* / Q000235:genetics; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000235:genetics*; D055785:Gene Knockdown Techniques / Q000379:methods*; D018390:Gene Targeting; D024861:Genetic Enhancement / Q000379:methods*; D034622:RNA Interference / Q000502:physiology*; D011995:Recombination, Genetic / Q000235:genetics; D032901:Talaromyces / Q000235:genetics*
chemical_list: D002482:Cellulose; D011088:DNA Ligases; D000072481:DNA Ligase ATP