Utilização de bulked segregant analysis na identificação de marcadores ligados a genes que controlam a resistência à ferrugem (Puccinia psidii Winter) em Eucalyptus sp

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

大麦雄核发育的基因型效应及相关的分子标记

基因型效应是影响植物雄核发育（Androgenesis）的重要因素，在很多情况下基因型的限制是单倍体育种用于实践的主要障碍。尽管人们对基因型效应进行了大量的遗传学研究，但对其机理仍然了解很少。本研究以具有不同花粉胚胎发生能力的两个大麦基因型为材料，细致地观察了大麦花粉体内发育和雄核发育的过程，确认大麦雄核发育途径包括营养核分裂途径、营养核与生殖核分裂途径和小孢子均等分裂途径三种。在雄核发育过程的每一个阶段，从单核、二核、多核花粉粒到愈伤组织，都可能发生败育。无论基因型是否具有高频率花粉胚胎发生能力，这种败育都是限制愈伤组织产量的重要因素。对两个基因型离体培养花药中不同类型花粉频率的连续统计表明，从培养开始到大量多核花粉粒形成，三种花粉－单核花粉、分裂花粉和死亡花粉的频率变化在两基因型中是相似的，发育途径都是以小孢子均等分裂途径为主，因此两基因型的早期雄核发育过程无明显区别。然而两基因型最终出愈率是有显著差异的，这种差异从时间上讲是在小孢子形成多核粉以后产生的。推测大麦不同基因型之间，花药培养反应的差异不是受小孢子脱分化启动能力控制的，更可能由其多核花粉发育成愈伤组织或胚状体的能力决定，可能与细胞壁的形成有关。从这种意义上来说，雄核发育过程中的愈伤组织形成又可以分为两个阶段：愈伤组织形成的诱导和愈伤组织形成的维持，并且它们分别受不同的基因控制，有不同的遗传机制。因此，大麦愈伤组织形成维持能力的差异是造成不同基因型对花药培养反应不同的主要原因。 利用BSA（Bulked Segregant Analysis）法对60个单株构成的F2分离群体进行分析。在520条随机引物中，有7个在高频池和低频池中的扩增产物有差异，其中引物S500重复性最好，其扩增带S500_(－650)经过单株验证后，确认与高频率胚胎发生能力连锁。聚丙烯酰胺凝胶电泳银染证明这一条带只存在于高频池中，可以初步确认是一个与高频率胚胎发生能力连锁的RAPD标记。

Isolation of Y- and X-linked SCAR markers in yellow catfish and application in the production of all-male populations

P>Sex controls have been performed in some farmed fish species because of significant growth differences between females and males. In yellow catfish (Pelteobagrus fulvidraco), adult males are three times larger than female adults. In this study, six Y- and X-linked amplified fragment length polymorphism fragments were screened by sex-genotype pool bulked segregant analysis and individual screening. Interestingly, sequence analysis identified two pairs of allelic genes, Pf33 and Pf62. Furthermore, the cloned flanking sequences revealed several Y- and X-specific polymorphisms, and four Y-linked or X-linked sequence characterized amplified region (SCAR) primer pairs were designed and converted into Y- and X-linked SCAR markers. Consequently, these markers were successfully used to identify genetic sex and YY super-males, and applied to all-male population production. Thus, we developed a novel and simple technique to help commercial production of YY super-males and all-male populations in the yellow catfish.

Discovery of a male-biased mutant family and identification of a male-specific SCAR marker in gynogenetic gibel carp Carassius auratus gibelio

Gibel carp ( Carassius auratus gibelio) is a uniquely gynogenetic species with a minor ratio of males in natural habitats, but its male origin and sex determination mechanisms have been unknown. In this study, a male-biased mutant family was discovered from the gynogenetic gibel carp, and a male-specific SCAR marker was identified from the mutant family. Normal spermatogenesis was observed in the male testes by immuno. fluorescence histochemistry. Nearly identical AFLP profiles were observed between males and females, but a male-specific 86 bp AFLP fragment was screened by sex-pool bulked segregant analysis and individual screening. Based on the male-specific AFLP fragment, a total of 579 bp sequences were cloned by genome walking. Subsequently, a male-specific SCAR marker was designed, and the male-specific DNA fragment was confirmed to be steadily transmitted to the next generation and consistently detected only in males. (C) 2009 National Natural Science Foundation of China and Chinese Academy of Sciences. Published by Elsevier Limited and Science in China Press. All rights reserved.

Mapeamento genómico de um gene de resistência ao míldio em Brassica oleracea L.

Tese dout., Biologia, Universidade do Algarve, 2006

Validation of microsatellite markers for assisted selection of soybean genotypes resistant to powdery mildew

Powdery mildew is one of the most serious diseases of soybean and is found in all producing countries. The purpose of this study was to validate microsatellite markers previously identified as associated with resistance to powdery mildew in soybean. The study was conducted in two F, parent populations with contrasting resistance to powdery mildew, In the analysis 10 SSR primers were used for the populations. and tour polymorphic markers were identified for cross I (MGBR95-20937 x IAC-Foscarin 31) and three for cross 2 (MGBR-46 x Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA) 48). The Chi-square analysis of the phenotypic evaluation confirmed the expected segregation (3: 1) of a dominant gene related to resistance. The polymorphic markers also segregated as expected (1:2:1). The markers Sat 366 and Sat 393 in the crosses 1 and 2. respectively, located at 9.41 and 12.45 cM from the gene. were considered promising for marker-assisted selection for resistance to powdery mildew in soybean. at a selection efficiency of 92.7% and 60.3% respectively.

Marcadores RAPD para detecção de resistência à ferrugem-asiática-da-soja

Os objetivos deste trabalho foram confirmar a herança da resistência da PI 459025 (Rpp4) à ferrugem-asiática-da-soja e identificar marcadores moleculares do tipo RAPD, ligados a este gene de resistência, em populações de soja. Pelo cruzamento dos genitores contrastantes PI 459025 x Coodetec 208 obteve-se uma população, cujas populações das gerações F2 e F2:3 foram artificialmente infectadas e avaliadas quanto à reação ao fungo Phakopsora pachyrhizi, pelo tipo de lesão (RB - resistente e TAN - suscetível). Com os resultados da avaliação fenotípica, dois bulks foram obtidos com DNA de plantas homozigóticas resistentes e suscetíveis, respectivamente, pela análise de bulks segregantes. de 600 iniciadores RAPD aleatórios, foram identificados três com fragmentos polimórficos entre os bulks e parentais contrastantes quanto à resistência. Pela análise do qui-quadrado, confirmaram-se: a herança monogênica, com dominância completa quanto à resistência ao patógeno, e a segregação 3:1 para a presença de banda dos três marcadores. Os três marcadores são ligados respectivamente a 5,1, 6,3 e 14,7 cM de distância do loco de resistência, em fase de repulsão no grupo de ligação G, o que foi confirmado pela utilização do marcador microssatélite Satt288. Estes marcadores são promissores na seleção assistida para resistência à ferrugem-asiática-da-soja.

Utilização de análise de segregantes agrupados na identificação de marcadores ligados a genes que controlam a resistência à ferrugem (Puccinia psidii Winter) em Eucalyptus sp.

Devido a grande importância da cultura de Eucalyptus no Brasil, empresas do setor florestal têm buscado através de programas de melhoramento genético, reduzir as perdas de produção e atender a demanda do mercado de papel e celulose. Um exemplo, é a busca por genes de resistência a doenças, principalmente a ferrugem causada por Puccinia psidii Winter, que resulta em redução da produtividade em plantas altamente suscetíveis. No presente trabalho, mudas de Eucalyptus pertencentes a uma geração F1, provenientes do cruzamento controlado entre parentais híbridos E. grandis X E. urophylla, sendo eles resistente e suscetível, foram inoculadas com Puccinia psidii em casa de vegetação e acompanhadas até o aparecimento dos sintomas da ferrugem. Foram classificadas, em dois grupos: resistentes (ausência de sintomas) e suscetíveis (presença de sintomas e esporulação). As amostras de DNA foram comparadas com o uso de marcadores moleculares associado ao método de BSA (Bulked Segregant Analysis). O polimorfismo entre os grupos foi geneticamente relacionado ao loco que determina a característica de resistência ou sucetibilidade. Dentre os 720 primers testados, 19 foram polimórficos, porém, apenas o marcador AK 01 manteve-se presente, quando testado em todos os indivíduos da população, mostrando-se a uma distância genética estimada de 20 cM em repulsão ao gene de resistência.

Mapping QTLs for aluminum tolerance in maize

Aluminum toxicity is one of the major constraints for plant development in acid soils, limiting food production in many countries. Cultivars genetically adapted to acid soils may offer an environmental compatible solution, providing a sustainable agriculture system. The aim of this work was to identify genomic regions associated with Al tolerance in maize, and to quantify the genetic effects on the phenotypic variation. A population of 168 F-3:4 families derived from a cross between two contrasting maize inbred lines for Al tolerance was evaluated using the NSRL and RSRL parameters in nutrient solution containing toxic level of aluminum. Variance analyses indicated that the NSRL was the most reliable phenotypic index to measure Al tolerance in the population, being used for further QTL mapping analysis. RFLP and SSR markers were selected for bulked segregant analysis, and additional SSR markers, flanking the polymorphisms of interest, were chosen in order to saturate the putative target regions. Seven linkage groups were constructed using 17 RFLP and 34 SSR markers. Five QTLs were mapped on chromosomes 2, 6 and 8, explaining 60% of the phenotypic variation. QTL(4) and marker umc043 were located on chromosomes 8 and 5, close to genes encoding for enzymes involved in the organic acids synthesis pathways, a widely proposed mechanism for Al tolerance in plants. QTL(2) was mapped in the same region as Alm2, also associated with Al tolerance in maize. In addition, dominant and additive effects were important in the control of this trait in maize.

Identification of a molecular marker linked to apomixis in Brachiaria humidicola (Poaceae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

QTL identification for tolerance to fruit set in tomato by FAFLP markers

This study aimed to detect quantitative trait loci (QTL) by fALP (Fluorescent Amplified Fragment Length Polymorphism) markers associated to the trait tomato fruit set at high temperatures. A biparental cross between line Jab-95 (heat-tolerant) and cultivar Caribe (heat-susceptible) was made. A total of 192 plants of the F2 generation were evaluated, generating 172 polymorphic markers through six primer combinations previously identified by the Bulked Segregant Analysis technique. To construct the genetic map, 106 of the 172 markers that segregated in the expected Mendelian segregation proportion (3:1) were used. The map covered 1191.46 cM of the genome. Six trait-linked QTL were identified in the analysis of simple markers and three others by the interval-mapping methodology. These results could be highly useful in improvement programs, since heat-tolerant plants can be selected rapidly, which improves tomato fruit set.

Caracterização de uma região genômica relacionada a uma anomalia de viveiro em Eucalyptus

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Seleção assistida por marcadores moleculares microssatélites para resistência ao oídio em soja

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Herança quali-quantitativa e marcadores moleculares para seleção assistida de genótipos de soja resistentes à ferrugem asiática

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)