Micronucleation by mitosis inhibitors in developing microspores of Spathiphyllum wallisii Regel

Abstract

KEY MESSAGE : We developed an efficient protocol for chromosome scattering in Spathiphyllum microspores. The effects of plant material, developmental age, genotype and antimicrotubular toxin type, exposure and concentration were evaluated. Asymmetric hybridization through microprotoplast-mediated chromosome transfer (MMCT) is a known method for overcoming sexual breeding barriers between distantly related plant species. To obtain microprotoplasts, it is necessary to induce mass micronucleation either in somatic or gametic cells. We have tested the efficiency for micronuclei induction of five mitosis inhibitors, amiprophos-methyl (APM), butamiphos (BUT), chlorpropham (CIPC), oryzalin (ORY) and propyzamide (PRO), on developing microspores of diploid Spathiphyllum wallisii Regel. Besides the used toxins, also the effect of their concentrations and incubation period as well as plant genotypes and material was tested. We observed micronuclei (MNi) in pollen mother cells, dyads and tetrads as well as other abnormalities such as ball metaphases and chromosome bridges. The flower position on the spadix and the type of starting material (dissected anthers vs. complete spadices) did not significantly influence micronucleation frequencies. The highest micronucleation index of 86% was obtained in microspores treated with 10μM ORY during 72h. All six genotypes tested formed micronuclei after this particular treatment, although the efficiency varied between cultivars. Next to ORY, CIPC was also a very efficient MNi inducer. The average number of MNi found in micronucleated cells varied between 1.67-6.44 for CIPC and 0.83-5.50 for ORY. The maximal number of MNi observed was 12 for CIPC and 9 for ORY. Our results demonstrate that CIPC and ORY can be applied for mass micronucleation on developing microspores of S. wallisii as a first step of MMCT in aroid interspecific or intergeneric breeding.