The combined effect of anther incubation time on CP induction medium (12, 14, and 16 days) and 2 concentrations of kinetin in R1 regeneration medium (0.1 and 0.3 mg/L) on the effectiveness of androgenesis was investigated in 17 genotypes of Capsicum grown in Poland. Plant material consisted of breeding lines and intraspecific hybrids of C. annuum; the species of C. frutescens, C. chinense, and C. baccatum var. pendulum; interspecific hybrids F 1 (C. frutescens × C. chinense) and F 1 (C. frutescens × C. baccatum); and doubled haploid lines derived from the hybrids. The results of the tested variants of the experiment were compared with the androgenic response of the control anthers cultured according to the standard protocol previously developed for pepper (12 days incubation on CP, 0.1 mg/L kinetin in R1). Under control conditions, androgenic embryos regenerated only from anthers of 3 of the tested genotypes, whereas application of the selected modifications promoted embryo development in an additional 12 genotypes. The highest effectiveness of androgenesis was observed after 16 days of anther incubation on CP medium combined with 0.1 mg/L kinetin in R1 medium. Twelveand 14-day-long anther incubation was more effective when followed by transferring anthers onto R1 medium supplemented with 0.3 mg/L kinetin.

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The present study evaluated the individual plants reaction of F 2 hybrid generation of C. annuum: ATZ1 9 PO and ATZ1 9 CDT as well as two interspecific hybrids: C. frutescens 9 C. annuum ATM1 and C. frutescens 9 C. chinense on androgenesis conditions in in vitro anther cultures. The experiment was carried out following a modified method of Dumas de Vaulx et al. (Agronomie 1:859-864, 1981). There were demonstrated clear differences in the effectiveness of androgenesis both between the pepper hybrid forms as well as among individual plants of all the genotypes tested. The highest effectiveness of androgenic embryos development was observed for the cultivated form of C. annuum: (ATZ1 9 PO)F 2 . Anthers of most of the plants of this hybrid produced embryos at the level higher than 5%, while in anther cultures of the second C. annuum hybrid (ATZ1 9 CDT)F 2 almost 3-fold fewer embryos and plants were produced. Anthers isolated from flower buds of interspecific hybrids formed much lower number of embryos. A positive reaction was recorded for five hybrid plants of (C. frutescens 9 C. annuum ATM1)F 2 , while in case of (C. frutescens 9 C. chinense)F 2 androgenic embryos were obtained from anthers of two plants. Only in the case of a one of these plants did the effectiveness of androgenesis exceed 5%. The ploidy level of the regenerants was determined by flow cytometry. Among the regenerants there were observed both haploid forms and the plants with the diploid number of chromosomes.

The aim of the research was the analysis of 11 DH-R2 pepper (Capsicum annuum L.) lines, obtained in anther cultures of ‘(ATZ1 × PO)’ F1, ‘(ATZ1 × CDT)’ F1 and ‘(ATZ1 × TG)’ F1 hybrids. To determine the genetic homogeneity of anther-derived lines, the biometrical characteristics of fruits, as well as fruit colour and shape inheritance, were analysed. The biometrical analysis determined the highest phenotype uniformity in androgenic lines of ‘(ATZ1 × PO)’ F1. Based on the fruit shape and colour variation, it was possible to determine the microspore origin of androgenic diploids obtained in anther cultures of ‘(ATZ1 × TG)’ F1. Additionally, enzyme analysis of four isozymes (PGM, PGI, IDH and MDH) showed that isocitrate dehydrogenase identified anther-derived diploids of hybrids between ‘ATZ1’ and ‘TG’, as well as those between ‘ATZ1’ and ‘CDT’, lines.

One of the key factors determining the effectiveness of pepper anther cultures is donor plant genotype. The stock material for androgenic embryos inductions is usually made up of hybrid forms, since the higher the degree of heterozygosity, the greater the chances of producing regenerates with unique genotypes. The aim of the presented research was to evaluate individual plant reaction in anther cultures for the C. annuum hybrid ('ATZ1' × 'TG')F 2 , interspecific hybrid (C. frutescens × C. chinense)F 2 and androgenic DH line AT6. The effectiveness of androgenesis was determined individually for each plant as the percentage of the embryos produced compared to the total number of anthers cultured. In the hybrid ('ATZ1' × 'TG')F 2 , anthers of 19 out of 20 plants evaluated produced embryos at a rate of 0.5 to 16.5%. Anthers of the AT6 DH line formed embryos considerably less frequently. A positive reaction was recorded for 13 out of 20 plants and the effectiveness of androgenesis did not exceed 3% for this genotype. The lowest androgenic response was recorded for the hybrid (C. frutescens × C. chinense)F 2 , where embryo development was observed in only five out of 19 plants and the effectiveness of androgenesis did not exceed 2%. The ploidy level of the regenerates was defined cytometrically. The analysis revealed the presence of haploid and diploid plants among the regenerates of all the genotypes evaluated.

To prove the androgenic origin of the regenerants obtained from the anther cultures of interspecific F 1 C. annuum L. ATZ1 × C. frutescens L.) hybrid, Random Amplified Polymorphic DNA (RAPD) method was applied.Thirty decamer primers were used in the experiment. The size of the amplified products ranged from 138 to 2369 bp, and their number from 3 to 25 per primer. About 274 loci, 47 of which were polymorphic, were analyzed. The results proved the hybrid status of anther donor plants detected by 20 of the applied primers. Of the 25 analyzed androgenic regenerants, 18 were diploids. The RAPD reactions confirmed their polymorphism in comparison with that of the F 1 hybrid generation, which excluded their origin from somatic cells of the anthers. Moreover, the results demonstrated the existence of a genetic variation among the obtained haploids, which illustrated genetic diversity of the microspores developing in hybrid anther cultures. The results of the experiments proved the utility of the RAPD method in detecting polymorphisms between closely related pepper plants, proving at the same time the effectiveness of androgenesis, regeneration of haploid plants, and spontaneously doubled haploids (DH lines).

A b s t r a c tEight anther-derived DH lines of pepper hybrids: two red-fruited (AP14, AP15) and two yellow lines (AP25 and AP32) of C. annuum (ATZ1 × PO)F2, two lines of (C. frutescens × C. annuum)F1 (FA1, FA2), and two of (C. frutescens × C. chinense)F1 (FCH2, FCH3), were studied regarding important morphological plant and fruit characters. C. annuum breeding line 'ATZ1' was used as a standard. The following traits were evaluated: total fruit yield, fruit weight, weight of placenta with seeds, technological matter, pericarp thickness, extract content, dry matter content, weight and number of seeds per fruit. The level of homogeneity within the DH lines was analysed with a one-way analysis of variance, additionally the values of coefficient of variation (CV) were determined for the tested plant characters. The highest phenotypic uniformity was noted for the red-fruited lines obtained from anthers of (ATZ1 × PO)F2 and for the DH lines of (C. frutescens × C. annuum) F1, while the least uniform were FCH2 and FCH3 lines of (C. frutescens × C. chinense)F1. The DH lines AP14 and AP15, as well as FA1 and FA2 were phenotype homogeneous in respect of weight of fruit, technological matter, pericarp thickness and extract content. For these parameters, also the values of CV were the lowest. Statistically significant interline polymorphism between the androgenic lines of the same origin was detected among the lines of C. annuum (ATZ1 × PO)F2.

In research on androhaploids in the progeny of interspecific hybrids within the Capsicum genus, three genetically stable lines of F 7 generation, selected from the C. annuum L. 9 C. chinense Jacq. hybrid, were used. In the first line, only callus tissue was formed as a reaction to the conditions of culture. Cytometric analysis of this tissue revealed the presence of cells with DNA content in nuclei at the level of 1C to 16C. The tissue was mixoploid and non-embryogenic. Anthers of the other line did not respond. In the third one, nine embryos were obtained, and they developed into plants. By means of cytometric analysis, the 1C DNA level was found in eight of them and these were androgenic plants. The origin of one of the diploid plant was not established due to the homozygous character of the donor plants. The experiment results confirm the already known diversity of genotype reaction to the conditions of culture. It moreover point to the possibility of selection of the forms with an androgenic potency from interspecific hybrids.