The efficiency of in vitro polyploidization depends on several variables associated to the plant, the antimicrotubule agent and the interactions between them. In the present work, we have used responsesurface methodology to determine the best operating conditions for plant recovery in polyploidization assays for shoot apices and somatic embryos of two seedless grape cultivars, employing colchicine and oryzalin. Explant type, tubulin-interfering compound and concentration were the critical factors determining plant recovery. Linear reduction in viable plant regeneration via organogenesis and somatic embryogenesis was obtained by increasing oryzalin concentrations and treatment time, whereas the effects of colchicine were better described by a quadratic design for both explants types. The conditions promoting higher rates of plant recovery were used in chromosome doubling experiments with oryzalin and colchicine for shoot apices and somatic embryos of 'Crimson seedless' and 'BRS Clara'. The established protocols allowed the recovery of non-chimerical autotetraploid plants at rates higher than 30 % for both cultivars. Stomata size parameters statistically correlate to the ploidy level of the regenerants and were effective for preliminary polyploidy screening. Autotetraploid lines of seedless grapes were incorporated into the Vitis germplasm bank for further agronomical evaluations. To our knowledge, this is the first report of in vitro oryzalin induced polyploidization of grapevine and of the use of mathematical modeling to optimize chromosome doubling in plants.

Two diploid (2n=22 chromosomes) and self-incompatible coffee species, Coffea canephora Pierre ex Froehner and C. dewevrei De Wild, et Th. Dur., were subjected to 2% acetic orcein and C-banding techniques with the purpose of mitotic chromosome characterizations. Karyological analysis were done in root cell chromosomes. These two diploid species of coffee could be distinguished mainly by some chromosome absolute (u,m) and relative (%) lenght mean values. In both species chromosome 5, 10 and 11 were metacentrics and the others were submetacentries; chromosomes 1 and 3 had secondary constriction beside centromere. The data suggested chromosome 3 had the NOR. Haploid karyotype formula was 1M + 2m + 7sm + lsm s for C. canephora, and 3m + 7sm + lsm s for C. dewevrei. Total karyotype length mean values were 16.97 ± 0.95u,m for C. canephora and 15.60± l.ll|imfor C. dewev rei and the difference was not significant. C-banded karyotypes were very similar in both species with prevailing centromeric/pericentromeric C-bands. The total C-band heterochromatin values expressed in percentage were 15.76 + 0.32% for C. canephora and 16.11 ± 0.26% for C. dewevrei. These values were not significantly different to separate both species. Similarly, no distinction could be made on karyotype asymmetry index mean values between the two species.

Boehmeria nivea Gaud., the popular ramie (or China grass) is a perennial, wind-pollinated, monoecious plant with small diclinous flowers (Rabechault 1951). It is well known for its long and resistent extra-xylary (secondary phloem) textile fibers (Fahn 1975). Ramie has also been exploited as a forage during the winter when the plants grow slowly and are not so good for fiber extraction (Benatti Jr. 1988). The species is native from the temperate asiatic regions and comprises many varieties with asexual propagation by rhizomes. Chromosome numbers was first established as 2n=28 by Krause (1930, cited by Krause 1931, for B. nivea and B. biloba and this number was later confirmed by Medwedewa (1934), even though they did not mention in which variety they worked.Studies on polyploidy induction in ramie with colchicine were done by some researchers as Chi and Lai (1964) and Dong et al. (1988) with the purpose of plant fiber improvement. However, there are few cytological informations from this kind of study related to microsporogenesis, to chromosome behavior or degrees of pollen fertility.So, the present work deals with microsporogenesis analysis in some plants of one ramie variety which were treated with colchicine solution, but without chromosome duplication. Materials and methods Materials and colchicine treatmentSeeds of Boehmeria nivea var. 'Miyasaky' were soaked in 0.1 % colchicine solution for 24 hr, then washed in distilled water, placed in Petri dishes with moist filter paper until germination. Germinated seeds were transferred to Petri dishes with filter paper soaked in 0.1 % colchicine solution for 18 hr. After this, seeds and radicules were carefully washed in distilled water and placed in other 214 Neiva Izabel Pierozzi and Romeu Benatti Jr.Cytologia 63

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-Chromosome studies were performed in V. champinii, V. cinerea, V. girdiana, V. labrusca, V. rotundifolia, V. rupestris and V. vinifera with the purpose of species characterization using chromosome morphometric data and NOR banding. A median ideogram was obtained for each species. The karyotype formula obtained varied from 7m + 12sm to 9m + 11sm. The species showed moderate chromosome asymmetry values according to TF% form, Stebbins, Romero Zarco and Paszko indices. V. champinii and V. girdiana were apart from the other species by CVcl and CVci graphic representation and also formed a group apart in the dendrogram based on Euclidian distances. The chromosome pair number 3 harbors the secondary constriction and a satellite segment in all species analyzed with Giemsa staining and it may be the same observed after NOR banding technique. It seems that the process of speciation in the North American Euvitis species studied involved some discrete changes in chromosome morphometry which have been reflected in the asymmetry index. Index terms: karyotype, NOR-banding, mitotic chromosomes, Vitis, Vitaceae. ESTUDOS CROMOSSOMICOS DE ESPÉCIES DE Vitis RESUMO -Estudos cromossômicos foram efetuados em V. champinii, V. cinerea, V. girdiana, V. labrusca, V. rotundifolia, V. rupestris e V. vinifera com a finalidade de caracterização das espécies, usando dados de morfometria cromossômica e do bandamento NOR. Foi obtido um ideograma médio para cada espécie estudada. A fórmula cariotípica variou de 7m + 12sm a 9m + 11sm. As espécies mostraram assimetria cariotípica moderada pelos índices de TF% forma, Stebbins, Romero Zarco e Paszko. V. champinii e V. girdiana ficaram à parte das demais espécies pela representação gráfica de CVcl e CVci e também pelo dendrograma simplificado obtido pelo método de distância Euclidiana. O par cromossômico nº 3 apresentou constrição secundária e segmento satélite nas espécies analisadas com Giemsa, podendo ser o mesmo evidenciado pelo bandamento NOR. Parece que o processo de especiação das espécies de Euvitis americanas estudadas envolveu mudanças discretas na morfometria dos cromossomos a qual se refletiu no índice de assimetria cromossômica. Termos para indexação: cariótipo, bandamento NOR, cromossomos mitóticos, Vitis, Vitaceae.