Thesis (MSc)--Stellenbosch University, 2003. / ENGLISH ABSTRACT: Apple (Malus x domestica Borkh.) is an important horticultural crop worldwide and in the Western Cape. The income generated from apple and other deciduous fruit production amounts to approximately 25% of the gross total value of horticultural production in the Western Cape. Unfortunately diseases and pests adversely affect fruit production in this region. Woolly apple aphids (Eriosoma lanigerum L. (Hausmann» have a significant effect on the apple industry in the Western Cape. Damage caused is two-fold, occurring aerially and terrestrially. Insects colonise the plants, feeding off the phloem sap. Aphid infestation around the root system results in repeated infestation of the foliage as it serves as a reservoir of aphids. In extreme cases, the apple cores are also infested, thus affecting the sale of apples. In 1962, Northern Spy was identified as a woolly apple aphid resistant rootstock and has since then formed the basis for traditional rootstock breeding programmes. The Er1 gene in Northern Spy confers resistance. According to one report, the natural resistance of Northern Spy was overcome in South Africa in 1968, but this was not confirmed in an independent study. The main objectives of this study was to firstly identify molecular markers more closely linked to the woolly apple aphid resistance gene, Er1, than existing markers, by applying AFLP technology to selected seedlings, identified to be resistant by conventional phenotyping. If identified, these markers can be incorporated into existing breeding programmes. Secondly, previously identified RAPD and SCAR markers were tested to determine their applicability in local populations for use in breeding programmes. Ultimately the segregation of the Er1 gene in South African populations can be determined if tightly linked markers are identified. Three families were derived from crosses of each of three resistant genotypes, namely Northern Spy, Rootstock 5 and Russian Seedling and a susceptible cultivar, Braeburn. For the three successive years of the study, each resistant genotype was allowed to cross-pollinate in isolation with the susceptible parent. Two hand-pollinated families, Russian Seedling x Liberty and Russian Seedling x Northern Spy, were also included in the study. The amplified fragment length polymorphism (AFLP) technique was used in an attempt to identify markers in the resistant and susceptible seedlings. No markers were identified using this technique. Known sequence characterised amplified regions (SCAR) and random amplified polymorphic DNA (RAPD) markers were used due to their suitability in marker-assisted selection for woolly apple aphid resistance. Varying results were obtained with these markers and no conclusive information was acquired with regard to the segregation of the Er] gene in any of these rootstocks and crosses. This underlines the need for the development of markers that can readily be applied in local breeding programmes. The identification and integration of such markers will greatly benefit the local and world wide apple industries. / AFRIKAANSE OPSOMMING: Appels (Malus x domestica Borkh.) is wêreldwyd en in die Wes-Kaap 'n belangrike landbougewas. Inkomste gegenereer deur appels en ander sagtevrugte vorm bykans 25% van die bruto inkomste uit vrugte in die Wes-Kaap. Siektes en insekpeste verlaag egter die produksie van vrugte in hierdie streek. Appelbloedluise (Eriosoma lanigerum L. (Hausmann» het 'n groot invloed op appelproduksie in die Wes-Kaap. Skade word bogronds en ondergronds aangerig. Insekte koloniseer die plant en leef op floeëmsap. Besmetting van die wortels lei tot herhaalde besmetting van bogrondse dele aangesien die insekte aanteelop die wortels. In uiterste gevalle word die vrugte geaffekteer, wat vrug-verkope beïnvloed. 'Northern Spy' is in 1962 geïdentifiseer as 'n onderstam met natuurlike weerstand teen appelbloedluis en het vir lank die basis gevorm vir tradisionele telingsprogramme. Weerstand word verleen deur die Erf geen. Volgens een verslag is die natuurlike weerstand van Northern Spy egter in 1968 in Suid-Afrika oorkom, maar dit is nog nie in 'n onafhanklike studie bevestig word nie. Die hoof doelstellings van hierdie studie was om eerstens deur middel van die AFLP tegniek molekulêre merkers te identifiseer wat nouer gekoppel is aan die appelbloedluis weerstandsgeen, En, as bestaande merkers. Hierdie tegniek is toegepas op saailinge wat deur konvensionele fenotipering geselekteer is. Indien merkers suksesvol geïdentifiseer is, kan dit in bestaande telingsprogramme geïntegreer word. Tweedens is bestaande RAPD en SCAR merkers ook getoets om hul toepaslikheid te bepaal vir gebruik in plaaslike teelprogramme. Oplaas sal die segregasie van die Erf geen in Suid- Afrikaanse populasies ook deur middel van nou gekoppelde merkers bepaal kan word. Kruisings van elk van die drie weerstandbiedende genotipes, naamlik 'Northern Spy', 'Rootstock 5' en 'Russian Seedling', en die vatbare kultivar, 'Braeburn' , het drie families daargestel. Elke weerstandbiedende genotipe is toegelaat om in isolasie te kruisbestuif met die vatbare ouer. Twee hand-bestuifde families, 'Russian Seedling' x 'Liberty' en 'Russian Seedling' x 'Northern Spy', is in 'n latere stadium van die studie ingesluit. Die AFLP tegniek is gebruik vir die identifikasie van polimorfiese merkers tussen vatbare en weerstandbiedende populasies. Geen merkers is egter geïdentifiseer nie. Bestaande SCAR en RAPD merkers is vervolgens gebruik om te bepaal of hulle geskik is vir gebruik in merker-bemiddelde seleksie vir appelbloedluis weerstand. Wisselende resultate is verkry ten opsigte van amplifikasie, herhaalbaarheid van resultate was swak en geen onweerlegbare bewyse oor die segregasie van die Erfgeen is bekom nie. Dit beklemtoon die noodsaaklikheid om merkers wat geredelik in plaaslike teelprogramme toegepas kan word, te ontwikkel. Die identifikasie en integrasie van sulke merkers sal die plaaslike en wêreld-wye appel industrieë aansienlik bevoordeel.

Thesis (MSc)--University of Stellenbosch, 2004. / ENGLISH ABSTRACT: Viruses are some of the most important pathogens of grapevines. There are no effective chemical treatments, and no grapevine- or other natural resistance genes have been discovered against grapevine infecting viruses. The primary method of grapevine virus control is prevention by biological indexing and molecular- and serological screening of rootstocks and scions before propagation. Due to the spread of grapevine viruses through insect vectors, and in the case of GRSPaV the absence of serological screening, these methods of virus control are not always effective. In the past several methods, from cross-protection to pathogen derived resistance (PDR), have been applied to induce plant virus resistance, but with inconsistent results. In recent years the application of post-transcriptional gene silencing (PTGS), a naturally occurring plant defense mechanism, to induce targeted virus resistance has achieved great success. The Waterhouse research group has designed plant transformation vectors that facilitate specific virus resistance through PTGS. The primary focus of this study was the production of virus specific transformation vectors for the introduction of grapevine virus resistance. The Waterhouse system has been successfully utilised for the construction of three transformation vectors with the pHannibal vector as backbone. Each vector contains homologous virus coat protein (CP) gene segments, cloned in a complementary conformation upstream and downstream of an intron sequence. The primary vector (pHann-SAScon) contains complementary CP gene segments of both GRSPaV and GLRaV-3 and was designed for the introduction of multiple-virus resistance. For the construction of the primary vector the GRSPaV CP gene was isolated from RSP infected grapevines. A clone of the GLRaV-3 CP gene was acquired. The second vector (pHann- LR3CPsas) contains complementary CP gene segments of GLRaV-3. The third vector (pHann-LR2CPsas) contains complementary CP gene segments of GLRaV-2. The cassette containing the complementary CP gene segments of both GRSPaV and GLRaV-3 was cloned into pART27 (pART27-HSAScon), and used to transform N tabacum cv. Petit Havana (SRI), through A. tumefaciens mediated transformation. Unfortunately potential transformants failed to regenerate on rooting media; hence no molecular tests were performed to confirm transformation. Once successful transformants are generated, infection with a recombinant virus vector (consisting of PYX, the GFP gene as screenable marker and the complementary CP gene segments of both GRSPaV and GLRaV-3) will be used to test for the efficacy of the vectors to induce resistance. A secondary aim was added to this project when a need was identified within the South African viticulture industry for GRSPaV specific antibodies to be used in serological screening. To facilitate future serological detection of GRSPaV, the CP gene was isolated and expressed with a bacterial expression system (pETI4b) within the E. coli BL2I(DE3)pLysS cell line. The expressed protein will be used to generate GRSPaV CP specific antibodies. / AFRIKAANSE OPSOMMING: Virusse is van die belangrikste patogene by wingerd. Daar bestaan geen effektiewe chemiese beheer nie, en geen wingerd- of ander natuurlike weerstandsgene teen wingerdvirusse is al ontdek nie. Die primêre metode van beheer t.o.v. wingerdvirusse is voorkoming deur biologiese indeksering, en molekulêre- en serologiese toetsing van onderstokke en entlote voor verspreiding. As gevolg van die verspreiding van wingerdvirusse deur insekvektore, en in die geval van GRSPa V die tekort aan serologiese toetsing, is dié metodes van virusbeheer nie altyd effektief nie. In die verlede is metodes soos kruis-beskerming en patogeen-afgeleide weerstand (PDR) gebruik om virusweerstand te induseer, maar met inkonsekwente resultate. In onlangse jare is post-transkripsionele geenonderdrukking (PTGS), 'n natuurlike plantbeskermingsmeganisme, met groot sukses toegepas om geteikende virusweerstand te induseer. Die Waterhouse-navorsingsgroep het planttransformasievektore ontwerp wat spesifieke virusweerstand induseer d.m.v. PTGS. Die vervaardiging van virus spesifieke tranformasievektore vir die indusering van wingerdvirusweerstand was die primêre doelwit van hierdie studie. Die Waterhouse-sisteem was gebruik vir die konstruksie van drie transformasievektore, met die pHannibal vektor as basis. Elke vektor bevat homoloë virus kapsiedproteïen (CP) geensegmente, gekloneer in 'n komplementêre vorm stroom-op en stroom-af van 'n intronvolgorde. Die primêre vektor (pHann-SAScon) bevat komplementêre CP geensegmente van beide GRSPaV en GLRaV-3, en was ontwerp vir die indusering van veelvoudige-virusweerstand. Die CP-geen van GRSPa V was vanuit RSP-geïnfekteerde wingerd geïsoleer, vir die konstruksie van die primêre vektor. 'n Kloon van die GLRa V-3 CP-geen was verkry. Die tweede vektor (pHann-LR3CPsas) bevat komplementêre CP geensegmente van GLRaV-3. Die derde vektor (pHann-LR2CPsas) bevat komplementêre CP geensegmente van GLRa V-2. Die kasset bestaande uit die komplementêre CP geensegmente van beide GRSPaV en GLRaV-3, was gekloneer in pART27 (pART27-HSAScon), en gebruik om N tabacum cv. Petit Havana (SRI) te transformeer d.m.v. A. tumefaciens bemiddelde transformasie. Ongelukkig het potensiële transformante nie geregenereer op bewortelingsmedia nie; gevolglik was geen molekulêre toetse gedoen om transformasie te bevestig nie. Na suksesvolle transformante gegenereer is, sal infeksie met 'n rekombinante-virusvektor (bestaande uit PYX, die GFP geen as waarneembare merker en die komplementêre CP geensegmente van beide GRSPa V en GLRa V-3) gebruik word om die effektiwiteit van die vektore as weerstandsinduseerders te toets. 'n Sekondêre doelwit is by die projek gevoeg toe 'n behoefte aan GRSPaV spesifieke teenliggame binne die Suid-Afrikaanse wynbedryf geïdentifiseer is, vir gebruik in serologiese toetsing. Om toekomstige serologiese toetsing van GRSPa V te bemiddel, was die CP-geen geïsoleer en in 'n bakteriële uitdrukkingsisteem (PETI4b) uitgedruk, in die E. coli BL21(DE3)pLysS sellyn. Die uitgedrukte proteïne sal gebruik word vir die vervaardiging van GRSPa V CP spesifieke antiliggame.

Thesis ( MScAgric) -- Stellenbosch University, 1997. / ENGLISH ABSTRACT: Stem rust is considered the most damaging of the wheat rusts causing yield losses of more than 50% in epidemic years. Similarly, Russian wheat aphids (RWA) can be regarded as one ofthe most devastating insect pests of wheat. Yield losses due to R W A primarily result from a reduction in plant resources (sucking plant sap). Secondary losses are incurred by viruses transmitted during feeding. Mapping disease and insect resistance genes that are effective against prevailing pathotypes and biotypes of South Africa will optimize their utilization in breeding programmes. The wheat line, 87M66-2-l, is homozygous for a single dominant stem rust resistance gene located on chromosome lD. This stem rust resistance gene has been derived from Triticum tauschii accession RL5289 and is here referred to as Srtau. The aim of this study was to determine the chromosome arm involved. Following the chromosome arm allocation of Srtau, its possible linkage with the genes Rg2, Lr 21 , Sr X and Sr 33 was studied. A telosomic analysis has shown that Srtau is located on chromosome arm 1 DS and is linked to the centromere with a recombination frequency of 21 ± 3 .40%. Glume blotch and a heavy mildew infection of segregating families planted in the field in 1996 made the linkage study between Lr 21 (leaf rust resistance) and Rg2 (glume colour) impossible. However, estimated linkages of 9 ± 1.9 map units between Sr33 (stem rust resistance) and Srtau, ± 6 map units between Sr X (stem rust resistance) and Sr 3 3 and ± 1 0 map units between Sr X and Srtau suggested that SrX, Sr33 and Srtau are closely linked on I DS. Taking existing map data into consideration, it seems that the most likely order of the genes is: centromere - Srtau - Sr 3 3 - Sr X. A single dominant R W A resistance gene, Dn5, was identified in the T aestivum accession 'SA 463' and is located on chromosome 7D. The aim ofthis study was to determine the chromosome arm involved. The possible linkage of Dn5 with the endopeptidase locus, Ep-D1 b. and chlorina mutant gene, cn-D1, was then studied. Endopeptidase zymograms of 'SA 463' revealed two unknown polymorphisms. F 2 monosomic analyses involving the chromosomes 7 A, 7B and 7D were performed in an attempt to identify the loci associated with these polymorphisms. Dn5 was mapped on chromosome arm 7DL. A recombination frequency of60 ± 4.53% between Dn5 and the centromere suggested the absence of linkage. Linkage between Ep-Dl and cn-Dl could not be calculated as a result of similar isoelectric points of the 7DL encoded endopeptidases of the parental material studied. Recombination frequencies of32 ± 4.97% between Dn5 and EpDl and 37 ± 6.30% between Dn5 and cn-Dl were, however, encountered. The two novel endopeptidase alleles encountered in 'SA 463' were designated as Ep-Dle and Ep-Ald. A RWA resistance gene was transferred from the rye accession ' Turkey 77' to wheat and in the process the RWA resistant wheat lines 91M37-7 and 91M37-51 were derived. No rye chromatin could be detected in these plants following C-banding. The aim of this study was to determine (i) on which chromosome the gene(s) is located, and (ii) whether the resistance can be the result of a small intercalary translocation of rye chromatin. A monosomic analysis of the RWA resistance gene in 91M37-51 has shown that a single dominant resistance gene occurs on chromosome 7D. The use of rye-specific dispersed probes did not reveal any polymorphisms between the negative controls and RW A resistant lines 91M3 7- 7 and 91M37-51 which would suggest that it is unlikely that the resistance was derived from rye. / AFRIKAANSE OPSOMMING: Stamroes word as die mees vemietigende graanroessiekte beskou en het in epidemiese jare oesverliese van meer as 50% tot gevolg. Russiese koringluise is eweneens een van die emstigste insekplae van koring. Russiese koringluise veroorsaak oesverliese deurdat dit plantsap uitsuig en die plant van voedingstowwe beroof. Dit tree egter ook as 'n virusvektor op en kan so indirekte oesverliese veroorsaak. Kartering van siekte- en insekweerstandsgene wat effektief is teen die Suid-Afrikaanse patotipes en biotipes, sal hulle gebruik in teelprogramme optimiseer. Die koringlyn, 87M66-2-l , is homosigoties vir 'n dominante stamroes-weerstandsgeen wat op chromosoom ID voorkom. Hierdie weerstandsgeen is uit die Triticum tauschii aanwins, RL5289, afkomstig en word hiema verwys as Srtau. Daar is gepoog om te bepaal op watter chromosoomarm Srtau voorkom, waama sy koppeling met betrekking tot die gene Rg2, Lr21 , SrX en Sr33 bepaal is. 'n Telosoomanalise het getoon dat Srtau op chromosoom-arm 1 DS voorkom en gekoppel is aan die sentromeer met 'n rekombinasie-frekwensie van 21 ± 3.40%. Segregerende populasies wat in 1996 in die land geplant is, is hewig deur aarvlek en poeieragtige meeldou besmet en dit het die moontlike bepaling van koppeling tussen Lr21 (blaarroesweerstand) en Rg2 (aarkaffie kleur) belemmer. Koppelingsafstande van 9 ± 1. 9 kaart-eenhede tussen Sr 33 (stamroesweerstand) en Srt au, ± 6 kaart -eenhede tussen Sr X ( stamroesweerstand) en Sr 3 3 en ± 1 0 kaart -eenhede tussen SrX en Srtau is geraam en toon dat SrX, Sr33 en Srtau nou gekoppel is. Die waarskynlikste volgorde van die gene op lDS is: sentromeer- Srtau- Sr33- SrX. 'n Enkele dominante Russiese koringluis-weerstandsgeen, Dn5, is in dieT aestivum aanwins 'SA 463 ' ge"identifiseer en kom op chromosoom 7D voor. Die studie het ten doel gehad om te bepaal op watter chromosoom-arm Dn5 voorkom, asook wat die koppeling van Dn5 met die endopeptidase lokus, Ep-Dl, en die chlorina mutante geen, cn-Dl , is. Endopeptidase simograrnme van 'SA 463' het twee onbekende polimorfismes getoon. Die gene wat kodeer vir hierdie twee polimorfismes is met behulp van F2 monosoom-analises wat die chromosome 7 A, 7B en 7D betrek, gei:dentifiseer. Dn5 is op chromosoom 7DL gekarteer. 'n Rekombinasie-frekwensie van 60 ± 4.53% is gevind vir die sentromeer en Dn5 en dui op die afwesigheid van koppeling. Koppeling tussen Ep-Dl en cn-Dl kon nie bepaal word nie omdat die endopeptidase bande geproduseer deur die ouerlike materiaal wat in die studie gebruik is, nie met sekerheid in die nageslag onderskei kon word nie. Rekombinasie-frekwensies van 32 ± 4.97% tussen Dn5 en Ep-Dl en 37 ± 6.30% tussen Dn5 en cn-Dl is egter bereken. Dit word voorgestel dat daar na die twee onbekende endopeptidase-allele wat in 'SA 463 ' voorkom, verwys word as Ep-Dle en Ep-Ald. 'n Russiese koringluis-weerstandsgeen is uit die rog-aanwins, 'Turkey 77', oorgedra na koring en in die proses is die Russies koringluis weerstandbiedende lyne, 91M37-7 en 91M37-51 , geproduseer. Geen rog-chromatien kon egter met behulp van C-bande in hierdie lyne waargeneem word nie. Die doel van die studie was om te bepaal (i) op watter chromosoom die geen(e) voorkom, en (ii), of die Russiese koringluis weerstandsgeen die gevolg kan wees van 'n klein interkalere translokasie van rog- chromatien. 'n Monosoom-analise van die Russiese koringluis-weerstandsgeen in 91M37-51 het getoon dat 'n enkele dominante weerstandsgeen op chromosoom 7D voorkom. Rog-spesifieke herhalende peilers het geen polimorfismes tussen negatiewe kontroles en die Russiese koringluis weerstandbiedende lyne 91M37-7 en 91M37-51 getoon nie. Dit is dus onwaarskynlik dat die weerstand in die lyne uit rog verhaal is.

Made available in DSpace on 2015-08-20T17:10:05Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-03-26. Added 1 bitstream(s) on 2015-08-20T17:25:53Z : No. of bitstreams: 1 000844642.pdf: 1476912 bytes, checksum: 23696bd53d8521611ee359e3a454c41b (MD5) / Amplamente disseminada pelo Brasil, causando enormes prejuízos em viveiros e plantios comerciais, a ferrugem do eucalipto, incitada por Puccinia psidii, é atualmente umas das principais doenças da cultura do eucalipto. A busca por progênies resistentes é facilitada quando existem informações a respeito do controle genético da resistência e estratégias de seleção, como a seleção assistida por marcadores moleculares. Assim, objetivou-se estimar parâmetros genéticos, avaliar o ganho genético por meio da seleção para resistência parcial à ferrugem causada por P. psidii em progênies F1 de Eucalyptus grandis, bem como a identificação de SNPs (Single Nucleotide Polymorphisms) nos genes quitinase e 1,3 β-glucanase relacionados à resistência. O trabalho foi dividido em dois capítulos. O primeiro capítulo foi realizado no município de Itararé-SP, onde foram instalados dois experimentos com 88 progênies F1 de Eucalyptus grandis. O delineamento experimental utilizado foi o de blocos casualizados, com quatro repetições, sendo cinco plantas por parcela, totalizando 1760 plantas por campo experimental. Aos 30 dias após o plantio, foi realizada a primeira avaliação quanto à ocorrência da ferrugem, mediante a escala de notas. Foram realizadas quatro avaliações mensais no período de setembro a dezembro de 2014. As estimativas dos componentes de variância e parâmetros genéticos foram obtidas pelo método REML/BLUP, empregando-se o programa genético-estatístico SELEGEN-REML/BLUP. Os parâmetros genéticos encontrados demonstraram pouca variabilidade genética e alta resistência à ferrugem. Os ganhos de seleção foram satisfatórios para essa fase da seleção. O segundo capítulo foi realizado no Laboratório de Biotecnologia e Genética Molecular, pertencente ao Departamento de Genética do Instituto de Biociências, da UNESP, câmpus de Botucatu ... / Widely disseminated in Brazil, causing huge losses in nurseries and commercial plantations, eucalyptus rust, incited by Puccinia psidii, is currently one of the main diseases of eucalyptus plantation. The search for resistant progenies is facilitated when information regarding the genetic control of resistance and selection trategies as an assisted selection by molecular markers are available. This study objective was to estimate genetic parameters, assess the genetic gain by selection for partial resistance to rust caused by P. psidii in Eucalyptus grandis of F1 progenies, as well as identification of SNPs (single nucleotide polymorphisms) in genes chitinase and β-1,3-glucanase related to resistance. This study was divided into two chapters. The first chapter was conducted in the city of Itararé-SP, which were installed two experiments with 88 F1 progenies of Eucalyptus grandis. The experimental design utilized was a randomized complete block design with four replications and five plants per plot, totaling 1760 plants per experimental field. At 30 days after planting was carried out the first assessment regarding the occurrence of rust, by the rating scale. Four monthly evaluations were carried out from September to December ...

Made available in DSpace on 2015-01-26T13:21:13Z (GMT). No. of bitstreams: 0 Previous issue date: 2014-08-19Bitstream added on 2015-01-26T13:30:58Z : No. of bitstreams: 1 000806750.pdf: 405498 bytes, checksum: d7b936b62c9737ef848ca34aff10a7e8 (MD5) / A alface (Lactuca sativa L.), pertencente à família Asteraceae, é uma das hortaliças mais consumidas no Brasil. Um dos principais problemas fitossanitários para essa cultura são as fitoviroses, em especial o Lettuce mosaic virus (LMV), que causa elevados prejuízos aos produtores em diversas regiões do país, além do Lettuce mottle virus (LeMoV), que vem crescendo em importância nos últimos anos. Como forma de controle, o uso de variedades resistentes é maneira mais eficaz de contornar os danos causados por vírus. Na maioria das variedades resistentes de alface faz-se o uso de genes recessivos, em sua maioria envolvidos na interação planta-vírus, são os chamados codificadores de fatores de iniciação de tradução eucarióticos (eIFs). Diante disso, este trabalho teve como objetivo avaliar dois genótipos de alface (169501 e 169501C), quanto a resistência ao LMV e ao LeMoV. Foram realizadas transmissões via extrato vegetal nos dois genótipos de alface com LMV e LeMoV em diferentes intervalos de tempo. Para LMV também foram realizadas transmissões por afídeos. As duas variedades testadas se mostraram resistentes ao LMV nas duas formas de transmissão, mas apresentaram sintomas quando inoculadas com LeMoV. Em reações de RT-PCR com oligonucleotídeos específicos, foram comprovadas a ausência de LMV e a infecção por LeMoV nas plantas inoculadas. O isolado de LMV após análise do sequenciamento da porção N’ terminal da ... / Lettuce (Lactuca sativa L.) belongs to the Asteraceae family and is one of the most consumed vegetables in Brazil. One of the major disease problems in this crop are plant viruses, especially the Lettuce mosaic virus (LMV) that cause losses to producers in various regions of the country, but also Lettuce mottle virus (LeMoV). As a form of control, the use of resistant varieties is the most effective way to overcome the damage caused by plant viruses. Most of the resistant varieties of lettuce makes the use of recessive genes, mostly involved in plant-virus interactions and characterized as eukaryotic translation initiation factor (eIFs). Thus, this study aimed to evaluate two lettuce genotypes (169501 and 169501C) for resistance to LMV and LeMoV. LMV and LeMoV were sap transmitted at different time of intervals. For LMV also transmissions with aphids were carried out. The two varieties tested were resistant to LMV in both forms of transmission, but showed symptoms when inoculated with LeMoV. In RT-PCR with specific primers, LMV was not detected but LeMoV yes, confirming the infection in the inoculated plants. The isolate of LMV used in the tests was not classified as belonging to subgroups Most ...

Resumo: O manejo de Bemisia tabaci biótipo B tem se tornado um desafio aos agricultores, uma vez que este inseto apresenta elevada capacidade em desenvolver resistência a todas as classes de inseticidas. Na busca por táticas alternativas e menos agressivas para o manejo das moscas-brancas, o uso de genótipos resistentes e a utilização de extratos de vegetais com atividades inseticida e/ou insetistática, têm revelado resultados promissores no combate à B. tabaci biótipo B. Dessa forma, o presente trabalho teve como objetivo avaliar a resistência de genótipos de aboboreiras (Cucurbita spp.) frente a B. tabaci biótipo B, estudar os tipos de resistência envolvidos e avaliar a possível interação no uso desses materiais com extratos aquosos provenientes de espécies da família Meliaceae. Realizou-se também análise bioquímica para confirmação da resistência de alguns genótipos. Para a instalação dos bioensaios, foi mantida uma criação de B. tabaci biótipo B em casa de vegetação. Vinte genótipos de Cucurbita spp. („Alicia AF 9354‟, „Aline AF 9353‟, „Golden Delight‟, „Nova Caravela‟, „Menina Morena‟, „Novita‟, „AF- 6741‟, „Atlas‟, „Bárbara 305‟, „Menina Brasileira‟, „Caserta‟, „Itapuã 301‟, „Tâmara‟, „Samira‟, „Canhão Seca Gigante‟, „Exposição‟, „Novita Plus‟, „Daiane‟, „Formosa‟, „Sandy‟) foram avaliados em ensaios (atratividade, oviposição com e sem chance de escolha, preferência alimentar e antibiose), sob condições de laboratório e casa de vegetação. A atratividade (adultos/cm2) foi avaliada 12, 24, 48 e 72 horas após a liberação. Após a última contagem, foram coletadas folhas para a contagem do número de ovos (com chance de escolha). As... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The management de Bemisia tabaci biotype B has become a challenge to agriculturalists, since this insect presents an elevated capacity to develop resistance to all classes of insecticides. In the search for less aggressive alternative tactics to control whiteflies, both resistant genotypes and vegetal extracts with insecticidal and / or insectistatic activities have revealed promising results for combating B. tabaci biotype B. Thus the present work aimed to evaluate the resistance of squash genotypes (Cucurbita spp.) challenged with B. tabaci biotype B, to study the types of resistance involved and to evaluate possible interactions in the use of these materials with aqueous extracts from species of the family Meliaceae. Biochemical analysis was also performed to confirm the resistance of some genotypes. To install bioassays, a creation of B. tabaci biotype B was maintained in a greenhouse. Twenty genotypes of Cucurbita spp. („Alicia AF 9354‟, „Aline AF 9353‟, „Golden Delight‟, „Nova Caravela‟, „Menina Morena‟, „Novita‟, „AF- 6741‟, „Atlas‟, „Bárbara 305‟, „Menina Brasileira‟, „Caserta‟, „Itapuã 301‟, „Tâmara‟, „Samira‟, „Canhão Seca Gigante‟, „Exposição‟, „Novita Plus‟, „Daiane‟, „Formosa‟, „Sandy‟) were evaluated in assays (attractiveness, oviposition, with and without free choice, feeding preference and antibiosis), under greenhouse and laboratory conditions. Attractiveness (adults/cm2) was evaluated 12, 24, 48 and 72 hours after release. After the final count, leaves were collected to enumerate the number of eggs (with free choice). Areas of leaves were measured to obtain the number of eggs/cm2 and the number of adults/ cm2. An oviposition preference assay was also performed without free choice. Antibiosis assays evaluated... (Complete abstract click electronic access below) / Orientador: Edson Luiz Lopes Baldin / Coorientador: Carlos Frederico Wilcken / Banca: Paulo Cesar Bogorni / Banca: Giuseppina Pace Pereira Lima / Banca: André Luiz Lourenção / Banca: Geraldo Andrade Carvalho / Doutor

Orientador: Edson Luiz Furtado / Coorientadora: Celso Luis Marino / Banca: Marcia Regina Costa / Banca: Quelmo Silva de Novaes / Banca: Adriana Zanin Kronka / Banca: Ana Carolina Firmino / Resumo: Amplamente disseminada pelo Brasil, causando enormes prejuízos em viveiros e plantios comerciais, a ferrugem do eucalipto, incitada por Puccinia psidii, é atualmente umas das principais doenças da cultura do eucalipto. A busca por progênies resistentes é facilitada quando existem informações a respeito do controle genético da resistência e estratégias de seleção, como a seleção assistida por marcadores moleculares. Assim, objetivou-se estimar parâmetros genéticos, avaliar o ganho genético por meio da seleção para resistência parcial à ferrugem causada por P. psidii em progênies F1 de Eucalyptus grandis, bem como a identificação de SNPs (Single Nucleotide Polymorphisms) nos genes quitinase e 1,3 β-glucanase relacionados à resistência. O trabalho foi dividido em dois capítulos. O primeiro capítulo foi realizado no município de Itararé-SP, onde foram instalados dois experimentos com 88 progênies F1 de Eucalyptus grandis. O delineamento experimental utilizado foi o de blocos casualizados, com quatro repetições, sendo cinco plantas por parcela, totalizando 1760 plantas por campo experimental. Aos 30 dias após o plantio, foi realizada a primeira avaliação quanto à ocorrência da ferrugem, mediante a escala de notas. Foram realizadas quatro avaliações mensais no período de setembro a dezembro de 2014. As estimativas dos componentes de variância e parâmetros genéticos foram obtidas pelo método REML/BLUP, empregando-se o programa genético-estatístico SELEGEN-REML/BLUP. Os parâmetros genéticos encontrados demonstraram pouca variabilidade genética e alta resistência à ferrugem. Os ganhos de seleção foram satisfatórios para essa fase da seleção. O segundo capítulo foi realizado no Laboratório de Biotecnologia e Genética Molecular, pertencente ao Departamento de Genética do Instituto de Biociências, da UNESP, câmpus de Botucatu ... / Abstract: Widely disseminated in Brazil, causing huge losses in nurseries and commercial plantations, eucalyptus rust, incited by Puccinia psidii, is currently one of the main diseases of eucalyptus plantation. The search for resistant progenies is facilitated when information regarding the genetic control of resistance and selection trategies as an assisted selection by molecular markers are available. This study objective was to estimate genetic parameters, assess the genetic gain by selection for partial resistance to rust caused by P. psidii in Eucalyptus grandis of F1 progenies, as well as identification of SNPs (single nucleotide polymorphisms) in genes chitinase and β-1,3-glucanase related to resistance. This study was divided into two chapters. The first chapter was conducted in the city of Itararé-SP, which were installed two experiments with 88 F1 progenies of Eucalyptus grandis. The experimental design utilized was a randomized complete block design with four replications and five plants per plot, totaling 1760 plants per experimental field. At 30 days after planting was carried out the first assessment regarding the occurrence of rust, by the rating scale. Four monthly evaluations were carried out from September to December ... / Doutor

Orientador: Renate Krause Sakate / Banca: Marcelo Agenor Pavan / Banca: Massaharu Marubayashi / Resumo: A alface (Lactuca sativa L.), pertencente à família Asteraceae, é uma das hortaliças mais consumidas no Brasil. Um dos principais problemas fitossanitários para essa cultura são as fitoviroses, em especial o Lettuce mosaic virus (LMV), que causa elevados prejuízos aos produtores em diversas regiões do país, além do Lettuce mottle virus (LeMoV), que vem crescendo em importância nos últimos anos. Como forma de controle, o uso de variedades resistentes é maneira mais eficaz de contornar os danos causados por vírus. Na maioria das variedades resistentes de alface faz-se o uso de genes recessivos, em sua maioria envolvidos na interação planta-vírus, são os chamados codificadores de fatores de iniciação de tradução eucarióticos (eIFs). Diante disso, este trabalho teve como objetivo avaliar dois genótipos de alface (169501 e 169501C), quanto a resistência ao LMV e ao LeMoV. Foram realizadas transmissões via extrato vegetal nos dois genótipos de alface com LMV e LeMoV em diferentes intervalos de tempo. Para LMV também foram realizadas transmissões por afídeos. As duas variedades testadas se mostraram resistentes ao LMV nas duas formas de transmissão, mas apresentaram sintomas quando inoculadas com LeMoV. Em reações de RT-PCR com oligonucleotídeos específicos, foram comprovadas a ausência de LMV e a infecção por LeMoV nas plantas inoculadas. O isolado de LMV após análise do sequenciamento da porção N' terminal da ... / Abstract: Lettuce (Lactuca sativa L.) belongs to the Asteraceae family and is one of the most consumed vegetables in Brazil. One of the major disease problems in this crop are plant viruses, especially the Lettuce mosaic virus (LMV) that cause losses to producers in various regions of the country, but also Lettuce mottle virus (LeMoV). As a form of control, the use of resistant varieties is the most effective way to overcome the damage caused by plant viruses. Most of the resistant varieties of lettuce makes the use of recessive genes, mostly involved in plant-virus interactions and characterized as eukaryotic translation initiation factor (eIFs). Thus, this study aimed to evaluate two lettuce genotypes (169501 and 169501C) for resistance to LMV and LeMoV. LMV and LeMoV were sap transmitted at different time of intervals. For LMV also transmissions with aphids were carried out. The two varieties tested were resistant to LMV in both forms of transmission, but showed symptoms when inoculated with LeMoV. In RT-PCR with specific primers, LMV was not detected but LeMoV yes, confirming the infection in the inoculated plants. The isolate of LMV used in the tests was not classified as belonging to subgroups Most ... / Mestre

Com o aumento da população mundial, são necessárias estratégias criativas para aumentar a produção de alimentos. Para superar esse imenso desafio, novos cultivares vem sendo desenvolvidos, através de diferentes técnicas, e com diferentes características, para alcançar uma maior produção de alimentos. Para conservação dessa produção vários métodos podem ser empregados, e a irradiação com cobalto constitui um método seguro e eficaz para o aumento no tempo de vida útil dos alimentos. Devido à importância comercial e nutricional da soja, possíveis alterações promovidas devem ser estudadas. O presente trabalho visa determinação dessas possíveis alterações causadas pela irradiação (com doses de 2, 4 e 8 kGy) em dois diferentes cultivares de soja (BRS 213 e Embrapa48) realizando análises químicas de perfil lipídico, isoflavonas, polifenóis totais, livres e ligados a proteínas, análise sensorial descritiva quantitativa, ensaio biológico para determinação da qualidade proteíca e ensaio com insetos para determinação de quebra de ressitência dos grãos e desinfestação. Os dois cultivares apresentaram perfil lipídico semelhante, e a irradiação promoveu pequena redução nas quantidades de ácidos linolênico e linoléico. Com relação as isoflavonas, o tratamento de radiação promoveu aumento nas quantidades das glicosiladas com dose de 2 kGy e diminuição com doses de 4 e 8 kGy enquanto aumentou a capacidade antioxidante e reduziu as proteínas ligadas à polifenóis. As alterações nas qualidades sensoriais dos grãos foram com relação a aparência e textura. Foi comprovada a eficiência da irradiação na desinfestação dos grãos e perda da resistência natural. As amostras irradiadas com 4 e 8 kGy se aproximaram dos índices de qualidade protéica aparente do padrão caseína. Os resultados obtidos indicam que o tratamento de radiação se oferece como um processo efetivo de desinfestação, sem prejudicar a qualidade da valiosa proteína de soja e com mínima alteração sensorial / With the increasement of the world population, criative strategies will be necessary to increase food prodution. To achieve this chalenge, new cultivars have been development, though different tecniques and with characteristics to improve the production. To maintain food conservation, several methods can be used. The use of Cobalto-60 radiation is a secure and useful method to increase the life time of foods. Due to the commercial and nutritional importance of soybean, some alterations must be studied. This study has the objective to determinate this alterations caused by irradiation (with doses of 2, 4 and 8 kGy) in grain of two different cultivars of soybean (BRS 213 e Embrapa 48), this study includes chemical analysis of lipid profile, isoflavones, total, free and protein-bound polyphenols, analysis sensory descriptive quantitative, biological assay for the determination of protein quality and insects assay for determination of break of resistance in the grains and desinfestation. Two cultivars presented similar lipid profile, and the irradiation promoted small reduction in the amounts of linoleic and linolenic acids. Regard to the isoflavones, the radiation treatment promoted increase in the amounts of the glycosides with dose of 2 kGy and decrease with doses of 4 and 8 kGy promoted increase in the antioxidant capacity and reduced protein-bound polyphenols. The alterations in the sensory qualities of the grains were regarding appearance and texture. It was proven the efficiency of the irradiation in the desinfestation of the grains and loss of the natural resistance. Samples irradiated with 4 and 8 kGy approached of the indices for apparent protein quality to the pattern casein. The results of this study indicating that radiation treatment of soybean offers considerable promise as an effective desinfestations process, without impairing the quality of the valuable soybean protein and with low sensory alteration

D.Phil. / The fungus, Verticillium dahliae, is the causative agent of Verticillium wilt, which results in significant cotton (Gossypium hirsutum) crop losses worldwide. This study contributes to the elucidation of cotton defence responses against V. dahliae. The identification, cloning and characterization of three genes that were differentially expressed in response to elicitation with a cell wall-derived (CWD) V. dahliae elicitor are described. It was hypothesized that the molecular architectures of the three characterized genes are supportive of a role in cotton defence against V. dahliae. As one of these genes was present as two homoeologous copies, this study also reports on the molecular characterization of both homoeologs, thus providing further insight into the processes of genomic evolution between homoeologous loci in allotetraploid cotton. The three genes were initially represented as expressed sequence tags (ESTs), obtained from a previous differential display reverse transcription polymerase chain reaction (DDRT-PCR) study by Zwiegelaar (2003), as part of an MSc project. These ESTs, designated C1B10, C4B5 and C4B4, were differentially induced upon elicitation with a CWD V. dahliae elicitor (Zwiegelaar, 2003). In the present study, the genes represented by the three ESTs were identified and characterized by genome walking and 5‘/3‘ rapid amplification of cDNA ends (RACE). Additionally, PCR and reverse-transcription PCR (RT-PCR) were utilized, where necessary, to obtain internal sequences, not covered by the genome walking and RACE reactions. Through the use of these molecular techniques, the full transcript and genomic sequences of each of the three genes was obtained, including their promoters. The promoter of each gene was analyzed for cis-elements driving gene transcription, through bioinformatic analysis. Furthermore, the copy number of each gene was determined through Southern blot analysis. The genes were translated to reveal their encoded protein sequences. The amino acid sequences were submitted to a basic local alignment (BLAST) search of the NCBI database to identify, and align them with, homologous proteins from other plant species (and those from G. hirsutum, if any). An in silico analysis of the encoded protein of each gene was also performed. This examination included domain architecture, post-translational modification, subcellular location and tertiary structure predictions. This study also involved the isolation of the elicitor from the cell walls of V. dahliae fungal cultures. The potency of the freshly-isolated elicitor was investigated with a triphenyltetrazolium chloride (TTC) viability assay on cotton cell suspensions. Its potential to induce PR-proteins was also explored but these results were inconclusive. In addition, expression studies were performed with real-time PCR (q-PCR), to confirm the up- or down-regulation of each gene upon elicitation of cotton cell suspensions with the CWD V. dahliae elicitor, and to investigate the time frame/kinetics of induction. The gene corresponding to the C1B10 EST was designated GhLIPN as this study revealed that it encodes a lipin protein. Lipins are novel proteins with phosphatidate phosphatase 1 (PAP1) activity, exclusive to eukaryotes. They play a fundamental role in the lipid metabolism of organisms ranging in complexity from yeast to animals and plants. In plants, this role includes lipid membrane remodelling during phosphate (Pi) deficiency. During the study of the GhLIPN gene, it was discovered that it occurred as two distinct homoeologous copies from the A- and D-co-resident genomes of allopolyploid G. hirsutum. The GhLIPN homoeologs were named GhLIPN I and N for Insert present and No insert, respectively, based on the presence or absence of a 13 base pair (bp) insertion/deletion (indel) site in intron 6.