A new and simple protocol for androgenesis in bell pepper is described. The initial medium, a modification of Nitsch and Nitsch's H medium, consisted of a two-phase system of semi-solid and liquid medium and contained maltose as carbon source. The total number of embryos formed was greater with maltose at 40 g·L-1, but embryos developed better at 10 to 20 g·L-1. Depending on the genotype, the number of embryos and plants recovered ranged from 3 to 750 and 0.25 to 8, respectively, per 100 flowers. Further increases in the number of embryos (up to 3561 per 100 flowers) and plants (up to 23 per 100 flowers) could be attained by flushing cultures with air enriched with CO2 at 900 μL·L-1. The ploidy level and the microspore origin of the recovered plants were determined by flow cytometry and zymograms for isocitrate dehydrogenase. Nearly 65% of the acclimated plants had undergone spontaneous doubling of the chromosome number, as confirmed by flow cytometry of leaf nuclei. Isocitrate dehydrogenase zymograms demonstrated that plants originated from microspores and that the two parental alleles were equally represented among the haploid and dihaploid plants.

ADDITIONAL INDEX WORDS. Cucumis sativus, Cucurbitaceae, parthenogenesis, haploid, DHL, pure line, hybrid ABSTRACT. Homozygous doubled haploid lines (DHLs) from new cucumber (Cucumis sativus L.) accessions could be useful to accelerate breeding for resistant varieties. DHLs have been generated by in vitro rescue of in vivo induced parthenogenic embryos. The protocol developed involves the following: 1) induction of parthenogenic embryos by pollinating with pollen irradiated with a Co 60 γ-ray source at 500 Gy; 2) in vitro rescue of putative parthenogenic embryos identifi ed by their morphology and localized using a dissecting scope or X-ray radiography; 3) discrimination of undesirable zygotic individuals from the homozygous plants using cucumber and melon SSR markers; 4) determination of ploidy level from homozygous plants by fl ow cytometry; 5) in vitro chromosome doubling of haploids; and 6) acclimation and selfi ng of selected lines. Codominant markers and fl ow cytometry confi rmed the gametophytic origin of plants regenerated by parthenogenesis, since all homozygous lines were haploids. No spontaneous doubled haploid plants were rescued. Chromosome doubling of haploid plants was accomplished by an in vitro treatment with 500 μM colchicine. Rescue of diploid or chimeric plants was shown by fl ow cytometry, prior to their acclimation and planting in the greenhouse. Selfi ng of colchicine-treated haploid plants allowed for the perpetuation by seed of homozygous lines. The high rate of seed set, 90% of the lines produced seed, facilitated the recovery of inbred lines. Despite some limiting factors, parthenogenesis is routinely used in a cucumber-breeding program to achieve complete homozygosity in one generation. Breeding for new commercial hybrid cultivars will be accelerated. DHLs are ideal resources for genomic analyses.

Melon (Cucumis melo) is one of the principal vegetable crops for fresh market, for which a large number of breeding programs, oriented to generate inbred pure lines and hybrids, is established worldwide. The process to obtain and select these lines has been highly accelerated by the use of biotechnological techniques such as the generation of doubled haploid line (DHL) populations and molecular markers. Moreover, the use of DHLs in genetic studies is a useful tool because of their complete homozygosity and the permanent availability of plant material perpetuated by seed. In this work, the parthenogenetic response of 17 melon genotypes and the F1 hybrid PI 161375 × Spanish cultivar Piel de Sapo (PS) was studied considering three stages along the in vitro DHL generation process. The response of the analyzed melon cultivars was heterogeneous through the DHL generation with different limiting steps for each genotype. The response of the PI 161375 × PS hybrid was more similar to the male (PS) than the female parent (PI 161375), although the response of the maternal genotype was higher for some stages. This points to the important role of alleles from both parents in the different steps of the DHL generation process, and it could explain the identification of six genomic regions with distorted allelic segregation skewed toward PS or PI 161375. This hybrid was used to generate a population of 109 DHLs, the gametophytic origin of which was confirmed by flow cytometry and molecular markers.

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Micropropagation of Pistacia vera `Mateur' was improved by adding MeJA to the multiplication and rooting media. Shoot-tip cultures established from grafted trees were maintained on a modified Murashige and Skoog medium containing 5 μm BA and 0.05 μm IBA. Adding 0.3, 1, or 3.2 μm MeJA improved shoot multiplication rates 2.5, 3.0, and 2.3, respectively. There was a significant interaction between the effects of auxin and temperature on the percentage of shoots forming roots. At 25C, the percentage of shoots forming roots was higher in the presence of NAA than IAA or IBA, whereas, at 28C, there was no difference among the auxins. Adding MeJA to the best auxin treatments-31.6 μm NAA at 25C and 31.6 μm IAA at 28C-increased the percentage of shoots forming roots and number of roots per shoot but decreased root length. More than 80% of the shoots rooted at 25C when 1 μM MeJA was added to the root induction medium, which contained 31.6 μm NAA, and the root elongation medium, without auxin. The large number of short roots induced by MeJA facilitated plantlet transfer to soil and acclimation. Chemical names used: methyl jasmonate (MeJA); N6-benzyladenine (BA), indole-3-butyric acid (IBA), α-naphthaleneacetic acid (NAA), indole3-acetic acid (IAA).