Roselaine Cristina Pereira

Duplicação cromossômica de gramíneas forrageiras: uma alternativa para programas de melhoramento genético

The artificial obtaining of duplicate genotypes aims to maximize forage characteristics of agronomic interest, such as nutritional value and forage production, resistance against pests and diseases, abiotic stress tolerance and fertility restoration in sterile hybrids. It also aims to obtain genetic variability in apomictic species through chromosome doubling of the sexual accesses by equaling the ploidy in order to allow crossings and the obtaining of fertile descendents. The polyploidy induction is made using antimitotic substances, and colchicine is the most widely used in forage. However, due to its toxicity, other substances such as herbicides and caffeine has been used successfully for grasses. The efficiency in obtaining the polyploidy artificially depends on various exogenous factors, such as the antimitotic substances used, the kind of explants, the time and the exposure conditions and the antimitotic concentrations. This review aims to present the main methods used to induce polyploidy in grasses and the progress obtained in genetic breeding by the use of artificially duplicate genotypes.

Physiological and cytological aspects of Inga vera subsp. affinis embryos during storage

The remarkably short storability of recalcitrant seeds has prevented their inclusion in programs of ex situ conservation. The causes of their desiccation sensitivity and rapid decline in viability during storage are not fully elucidated yet. In this study the highly recalcitrant, fully viable embryos of the tree species Inga vera subsp. affinis were stored under various conditions and analyzed physiologically and cytologically at intervals, in order to obtain more insights in their loss of viability during storage. Embryos stored fully hydrated at 5oC fully lost viability in 18 d. Sealed storage of partially dehydrated embryos slightly prolonged their viability, with 95% germination being attained after 14 d of storage. However, after 30 d of storage, viability was completely lost. Storage of hydrated embryos in a solution of polyethylene glycol (PEG) at -1.7 MPa water potential was capable of maintaining high germinability until 30 d of storage. When added to the PEG solution, abscisic acid showed a strong temperature-dependent interaction, with a positive effect on the longevity of embryos stored at 20oC and a negative effect on embryos stored at 5oC. In any case, embryo viability could be maintained no longer than 50 d. Embryos from seeds collected from the same trees in the following year showed better storability and still attained 45% germination after 62 d of storage. Analysis of cellular alterations during storage and viability loss of the embryos showed disappearance of starch granules and various damages to the cells, such as cell wall folding and cytoplasm fragmentation.

Obtaining tetraploid plants of ruzigrass (Brachiaria ruziziensis)

The objective of the present work was to induce chromosome doubling in genotypes of Brachiaria ruziziensis with potential for production of good-quality biomass and tolerance/resistance to stress caused by aluminum and spittlebugs. Germinated seeds of B. ruziziensis were immersed in 0.1% colchicine solution for 2 or 3 hours. Ploidy level was determined by flow cytometry and confirmed by chromosome count. Fertility of the tetraploid plants was assessed through evaluation of pollen viability by staining. There was no statistical difference with regard to seedling survival and number of duplicate seedlings between the treatments. Survival rate was 8%, of which 11.45% were tetraploid genotypes. The mean viability of pollen grains from the evaluated tetraploid plants ranged from 51.75 to 55.50%. So, the tetraploids plants obtained from genotypes of a bred population of ruzigrass produce fertile pollen, in percentages that render crosses viable in genetic breeding programs.

Nuclear DNA content and chromosome number in Brachiaria spp. genotypes

Os programas de melhoramento de Brachiaria spp. utilizam os cruzamentos intraespecificos e interespecificos entre plantas sexuais e apomiticas para obtencao de novas cultivares com caracteristicas desejadas. Como existem diferentes niveis de ploidia dentro e entre as especies deste genero, torna-se necessaria a avaliacao dos genotipos utilizados em programas de melhoramento para a orientacao das estrategias de cruzamentos adotadas pelos melhoristas. Neste trabalho, o conteudo de DNA e o numero cromossomico foram determinados para caracterizar o nivel de ploidia de genotipos de Brachiaria spp. Na analise de 15 genotipos, o conteudo de DNA variou de acordo com o nivel de ploidia (2x, 3x e 4x) e entre especie e/ou taxon. O conteudo de medio de DNA foi de 1,74 pg (2x) para B. ruziziensis, 3,74 pg (4x) para B. decumbens e de 3,52 pg (4x) para B. brizantha. Para o genotipo 86 obteve-se 2,57 pg de DNA e 2n = 3x = 27, indicando tratar-se de um acesso triploide, provavelmente um hibrido natural. A variacao no conteudo de DNA total permitiu diferenciar Brachiaria ruziziensis (2n = 2x = 18) das especies tetraploides (2n = 4x = 36) Brachiaria decumbens e Brachiaria brizantha, bem como o provavel hibrido triploide (2n = 3x = 18) dessas especies.

Histogram score contributes for reliability of DNA content estimatives in Brachiaria spp

Flow cytometry allows to estimate the DNA content of a large number of plants quickly. However, inadequate protocols can compromise the reliability of these estimates leading to variations in the values of DNA content the same species. The objective of this study was to propose an efficient protocol to estimate the DNA content of Brachiaria spp. genotypes with different ploidy levels using flow cytometry. We evaluated four genotypes (B. ruziziensis diploid and artificially tetraploidized; a tetraploid B. brizantha and a natural triploid hybrid), three buffer solutions (MgSO4, Galbraith and Tris-HCl) and three species as internal reference standards (Raphanus sativus, Solanum lycopersicum e Pisum sativum). The variables measured were: histogram score (1-5), coefficient of variation and estimation of DNA content. The best combination for the analysis of Brachiaria spp. DNA content was the use of MgSO4 buffer with R. sativus as a internal reference standard. Genome sizes expressed in picograms of DNA are presented for all genotypes and the importance of the histogram score on the results reliability of DNA content analyses were discussed.