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An evaluation of a new approach to the regeneration of Helichrysum italicum (Roth) G. Don, and the molecular characterization of the variation among sets of differently derived regenerants

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Abstract

A protocol for the induction of regeneration from leaves of Helichrysum italicum was established. Calli were found to form on the basal medium only when it was supplemented with thidiazuron (TDZ) alone or in combination with naphthalene acetic acid (NAA), with a percentage ranking of at least 80%. The hormone-free medium showed the highest percentage of shoot regeneration (62%) even though no callus formed. AFLP markers were employed to verify tissue culture-induced variation in the regenerated plantlets obtained by direct shoot regeneration or the indirect shoot regeneration process (callus formation). Seven out of the eleven AFLP primer pairs yielded polymorphic patterns. The average number of fragments per primer pair was 64.1. Singletons were represented by 12 (2.7%) fragments. Student’s T-test was performed both on the average number of shared fragments and on the nucleotide diversity, and no significant statistical difference was observed between the two regeneration treatments.

Abbreviations

AFLP:

amplified fragment length polymorphism

BM:

basal medium

NAA:

naphthalene acetic acid

RAPD:

random amplified polymorphic DNA

RFLP:

restriction fragment length polymorphism

SSR:

simple sequence repeats

TDZ:

thidiazuron

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Correspondence to Cinzia Montemurro.

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These authors contributed equally to this work.

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Key words

  • Helichrysum italicum
  • in vitro culture
  • Tissue culture-induced variation
  • AFLP markers
  • Nucleotide diversity