Two approaches for induction and isolation of starch mutants in potato (Solanum tuberosum L.): random versus gene targeted mutagenesis = Twee benaderingen voor de inductie en isolatie van zetmeelmutanten in aardappel (Solanum tuberosum L.): ongerichte versus gen gerichte mutagenese

T.J.H. Hoogkamp

Research output: Thesisinternal PhD, WU

Abstract

<p>In this thesis two approaches were used to induce structural mutations in potato starch biosynthesis genes in potato. First production of new monoploid <em>amf</em> genotypes through parthenogenesis made it possible to initiate mutation breeding for <em>amfae</em> double mutants. Two amf monoploids were selected which fulfilled most of the prerequisites. By inducing a mutation in one of the branching enzymes in an <em>amf</em> -mutant it is possible to select a double mutant having less branched amylopectin. This mutation can easily be identified by iodine staining. Amylose-free starch will stain red and less branched amylopectin will stain blue, like amylose containing starch. Mutations were induced by X-ray irradiation of leaf explants followed by adventitious shoot regeneration and microtuber induction or followed by several rounds of multiplication of axillary buds and microtuber induction. In both cases the starch of microtubers was stained with iodine to screen for aberrant types. In 56 tuber samples blue or otherwise aberrant starch granules were found. With this kind of observations, the concept of mutation breeding for starch variants in monoploid potatoes is proven. A second way to induce structural mutations was the use of the <em>Ac</em> ( <em>Activator</em> )/ <em>Ds</em> ( <em>Dissociation</em> ) transposase system of maize in potato where the <em>Ds</em> transposon is activated by a transposase source. In this study the <em>Ds</em> element was linked to the GBSS gene, of which the phenotypic effect of deactivation is known i.e. red staining starch after iodine staining. The known <em>amf</em> mutation was used as a model system to gather more information about the transposition frequency of the <em>Ds</em> transposable elements in potato and to test the tagging of the wildtype GBSS gene. To activate the <em>Ds</em> element four <em>Ds</em> transposon containing plants were combined with the Ac transposase via cross combination or double transformation. Excision rates ranged from 14.8-48.4 %. Three phenotypic starch mutants were found after screening by iodine staining of tuber cut surfaces. These amylose-free mutants were analyzed by <em>in vitro</em> tests, Southern blot hybridization and sequencing. Strong indications were found that inactivation of the GBSS gene was caused by a transposable element.</p>
Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • Wageningen University
Supervisors/Advisors
  • Jacobsen, Evert, Promotor
  • Visser, Richard, Promotor
Award date8 Oct 2001
Place of PublicationS.l.
Publisher
Print ISBNs9789058084538
Publication statusPublished - 2001

Keywords

  • potatoes
  • solanum tuberosum
  • plant breeding
  • induced mutations
  • starch
  • transposable elements
  • haploidy
  • mutants
  • transposition

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