The molecular basis for the gene-for-gene relationship of Vm-resistance in apple to Venturia inaequalis was investigated. Incompatible reactions involved a hypersensitive response (HR), which was accompanied by the accumulation of dark brown pigments and autofluorescent materials in epidermal and mesophyll cells at the site of invasion. Cell-free culture filtrates of the avirulent isolate elicited an HR in the Vm host (h5) leaves, but not in the susceptible host (h1). The elicitor activity was resistant to boiling but was abolished by proteinase K digestion. Elicitation of HR was used to monitor purification of the avirulence factor, AVRVm, from liquid cultures of the avirulent isolate following ultrafiltration, acetone precipitation and ion-exchange chromatography. The purest fraction contained three major proteins all with low isoelectric points (pI 3.0-4.5). The fraction also elicited HR on the differential host h4, but not on other resistant hosts (h2, h3 and h6) tested. Three candidate AVRVm proteins were identified and amino acid sequences were obtained using Edman degradation and mass spectrometry. Nucleotide sequences corresponding to these proteins were found in databases of V. inaequalis expressed sequence tags. There were no polymorphisms evident between avirulent and virulent isolates (representing races 1 and 5 respectively) either at genomic DNA or cDNA level of the full open reading frames. RT-PCR revealed that all genes were expressed in both avirulent and virulent isolates during in vitro and in planta growth. All three genes showed similar levels of expression between avirulent and virulent isolates during their in vitro growth. However, preliminary RT-PCR experiments showed that two of these genes were likely to be expressed at lower levels in the virulent compared with the avirulent isolate during compatible infection. Implications of this difference in expression and the future experiments to identify the genuine AvrVm gene were discussed.

The molecular basis for the gene-for-gene relationship of Vm-resistance in apple to Venturia inaequalis was investigated. Incompatible reactions involved a hypersensitive response (HR), which was accompanied by the accumulation of dark brown pigments and autofluorescent materials in epidermal and mesophyll cells at the site of invasion. Cell-free culture filtrates of the avirulent isolate elicited an HR in the Vm host (h5) leaves, but not in the susceptible host (h1). The elicitor activity was resistant to boiling but was abolished by proteinase K digestion. Elicitation of HR was used to monitor purification of the avirulence factor, AVRVm, from liquid cultures of the avirulent isolate following ultrafiltration, acetone precipitation and ion-exchange chromatography. The purest fraction contained three major proteins all with low isoelectric points (pI 3.0-4.5). The fraction also elicited HR on the differential host h4, but not on other resistant hosts (h2, h3 and h6) tested. Three candidate AVRVm proteins were identified and amino acid sequences were obtained using Edman degradation and mass spectrometry. Nucleotide sequences corresponding to these proteins were found in databases of V. inaequalis expressed sequence tags. There were no polymorphisms evident between avirulent and virulent isolates (representing races 1 and 5 respectively) either at genomic DNA or cDNA level of the full open reading frames. RT-PCR revealed that all genes were expressed in both avirulent and virulent isolates during in vitro and in planta growth. All three genes showed similar levels of expression between avirulent and virulent isolates during their in vitro growth. However, preliminary RT-PCR experiments showed that two of these genes were likely to be expressed at lower levels in the virulent compared with the avirulent isolate during compatible infection. Implications of this difference in expression and the future experiments to identify the genuine AvrVm gene were discussed.

The molecular basis for the gene-for-gene relationship of Vm-resistance in apple to Venturia inaequalis was investigated. Incompatible reactions involved a hypersensitive response (HR), which was accompanied by the accumulation of dark brown pigments and autofluorescent materials in epidermal and mesophyll cells at the site of invasion. Cell-free culture filtrates of the avirulent isolate elicited an HR in the Vm host (h5) leaves, but not in the susceptible host (h1). The elicitor activity was resistant to boiling but was abolished by proteinase K digestion. Elicitation of HR was used to monitor purification of the avirulence factor, AVRVm, from liquid cultures of the avirulent isolate following ultrafiltration, acetone precipitation and ion-exchange chromatography. The purest fraction contained three major proteins all with low isoelectric points (pI 3.0-4.5). The fraction also elicited HR on the differential host h4, but not on other resistant hosts (h2, h3 and h6) tested. Three candidate AVRVm proteins were identified and amino acid sequences were obtained using Edman degradation and mass spectrometry. Nucleotide sequences corresponding to these proteins were found in databases of V. inaequalis expressed sequence tags. There were no polymorphisms evident between avirulent and virulent isolates (representing races 1 and 5 respectively) either at genomic DNA or cDNA level of the full open reading frames. RT-PCR revealed that all genes were expressed in both avirulent and virulent isolates during in vitro and in planta growth. All three genes showed similar levels of expression between avirulent and virulent isolates during their in vitro growth. However, preliminary RT-PCR experiments showed that two of these genes were likely to be expressed at lower levels in the virulent compared with the avirulent isolate during compatible infection. Implications of this difference in expression and the future experiments to identify the genuine AvrVm gene were discussed.

The molecular basis for the gene-for-gene relationship of Vm-resistance in apple to Venturia inaequalis was investigated. Incompatible reactions involved a hypersensitive response (HR), which was accompanied by the accumulation of dark brown pigments and autofluorescent materials in epidermal and mesophyll cells at the site of invasion. Cell-free culture filtrates of the avirulent isolate elicited an HR in the Vm host (h5) leaves, but not in the susceptible host (h1). The elicitor activity was resistant to boiling but was abolished by proteinase K digestion. Elicitation of HR was used to monitor purification of the avirulence factor, AVRVm, from liquid cultures of the avirulent isolate following ultrafiltration, acetone precipitation and ion-exchange chromatography. The purest fraction contained three major proteins all with low isoelectric points (pI 3.0-4.5). The fraction also elicited HR on the differential host h4, but not on other resistant hosts (h2, h3 and h6) tested. Three candidate AVRVm proteins were identified and amino acid sequences were obtained using Edman degradation and mass spectrometry. Nucleotide sequences corresponding to these proteins were found in databases of V. inaequalis expressed sequence tags. There were no polymorphisms evident between avirulent and virulent isolates (representing races 1 and 5 respectively) either at genomic DNA or cDNA level of the full open reading frames. RT-PCR revealed that all genes were expressed in both avirulent and virulent isolates during in vitro and in planta growth. All three genes showed similar levels of expression between avirulent and virulent isolates during their in vitro growth. However, preliminary RT-PCR experiments showed that two of these genes were likely to be expressed at lower levels in the virulent compared with the avirulent isolate during compatible infection. Implications of this difference in expression and the future experiments to identify the genuine AvrVm gene were discussed.

The expression, accumulation and kinetic properties of 1 -aminocyclopropane- 1 -carboxylic acid (ACC) oxidase (ACO), the enzyme which catalyses the final step in the ACC-dependent pathway of ethylene biosynthesis in plants, is examined. The investigation is divided into three sections: (i) identification of two ACO genes in apple leaf tissue, designated MD-ACO2 and MD-ACO3, (ii) kinetic analyses of each of the three isoforms of ACO in apple (MD-ACO1, MD-ACO2 and MD-ACO3) expressed in E. coli, and (iii) temporal and developmental expression in vivo of each of the ACO genes and accumulation of the corresponding gene products in leaf and fruit tissue. The coding regions of putative ACO gene transcripts were generated from leaf tissue using RT-PCR. Sequence alignments and interrogation of the expressed sequence tags (ESTs) database indicated that the entire open reading frame (ORF) sequences were encoded by two distinct genes, and these are designated MD-ACO2 and MD-ACO3. A third ACO gene had been identified in apple by other research workers previously, and designated MD-ACO1. Differences are obvious in the number of base-pairs (bp) constituting the entire ORF of MD-ACO1 (942 bp), MD-ACO2 (990 bp) and MD-ACO3 (966 bp). MD-ACO1 and MD-ACO2 share a close nucleotide sequence identity of 93.9 % in the ORF but diverge in the 3' untranslated regions (3' -UTR) (69.5 %). In contrast, MD-ACO3 shares a lower sequence identity with both MD-ACO1 (78.5 %) and MD-ACO2 (77.8 %) in the ORF, and in the 3'-UTR (MD-ACO1, 68.4 %; MD-ACO2, 71 %). A comparison of the gene structures show that the endonuclease restriction sites are unique to each individual MD-ACO sequence. Genomic Southern analysis, using probes spanning the 3'-UTR and the 3'-end of the coding region confirmed that MD-ACO3 is encoded by a distinct gene. However, while the distinction between MD-ACO1 and MD-ACO2 is not as definitive, different gene expression patterns adds credence to their distinctiveness. Each of the three deduced amino acid sequences contain all of the residues hitherto reported to be necessary for maximal ACO activity. Expression of MD-ACO1, MD-ACO2 and MD-ACO3 as fusion proteins in E. coli was induced using isopropy1-β-D-thiogalactopyranoside (IPTG), the recombinant proteins purified by nickel-nitrilotriacetic acid (NiNTA) affinity chromatography and the products had predicted masses determined from the nucleotide sequences, including the His-tag of 38.53 kDa (MD-ACO1), 40.39 kDa (MD-ACO2) and 39.3 kDa (MD-ACO3). Polyclonal antibodies were raised against the MD-ACO3 fusion in rabbit for western blot analysis. Antibodies had been raised previously against recombinant MD-ACO1, and while it was considered likely the MD-ACO2 would be recognized by the MD-ACO1 antibodies, MD-ACO2 does not appear to be recognized in vivo by the antibody. Analyses of the kinetic properties of the three apple ACOs was undertaken. Apparent Michaelis constants (Km) of 89.39 μM (MD-ACO1), 401.03 μM (MD-ACO2) and 244.5 μM (MD-ACO3) have been determined which suggests differences in the affinity of each enzyme for the substrate ACC. Maximum velocity (Vmax) was determined for MD-ACO1 (15.15 nmol), MD-ACO2 (12.94 nmol) and MD-ACO3 (18.94 nmol). The catalytic constant (Kcat) was determined for MD-ACO1 (6.6 x 10-2), MD-ACO2 (3.44 x 10-2) and for MD-ACO3 (9.14 x 10-2), with kcat/Km(μM s-1) values of 7.38 x 10-4 μM s-1 (MD-ACO1), 0.86 x 10-4 M s-1 (MD-ACO2) and 3.8 x 10-4 μM s-1 (MD-ACO3). The optimal pH for MD-ACO1 was 7.0 - 7.5, MD-ACO2 7.5 - 8.0 and MD-ACO3 7.0 - 8.0. All three isoforms exhibited absolute requirements for the co-substrate ascorbate in vitro with optimal activity at 30 mM. Similarly, ferrous iron (FeSO4.7H20; of 15 - 25 μM) and sodium bicarbonate (NaHCO3; of 30 mM) were required for optimal activity, and were the same for all isoforms. No significant difference in thermostability was found in this study between the MD-ACO isoforms at the P = 0.05 level. However, the activities of the enzyme differed significantly between temperatures over time. In vivo expression of each of the ACO genes in leaf tissue determined using RT-PCR and cDNA Southern analysis reveal that both MD-ACO2 and MD-ACO3 are expressed in young leaf tissue and in mature leaf tissue. While MD-ACO3 is expressed predominantly in young leaf tissue, MD-ACO2 (in common with MD-ACO1) is expressed predominantly in mature fruit tissue. None of the MD-ACOs were observed to be senescence associated genes (SAG). MD-ACO3 protein accumulated predominantly in young leaf tissue and less intensely in both mature leaf tissue and young fruit tissue, while MD-ACO1 accumulated only in mature fruit tissue. For the developmental studies, samples were collected at approximately 11 am in this study. MD-ACO2 and MD-ACO3 were also expressed in leaf tissue collected over a 24 h period in the spring and also in the autumn. For both genes transcripts accumulated in the presence of fruit but tended to disappear in the absence of fruit. These results show that MD-ACO1, MD-ACO2 and MD-ACO3 are differentially expressed, and that MD-ACO3 is encoded by a gene distinct from MD-ACO1 and MD-ACO2. MD-ACO1 and MD-ACO2 are either allelic forms of the same gene or closely clustered. Although there is some variation in kinetic properties which may reflect different physiological environments, they do not vary greatly.

Thesis (Master)--İzmir Institute of Technology, İzmir, 2006 / Keywords:Semiotics, logo, brand emotion, apple computers, IBM. Includes bibliographical references (leaves. 111-122).

The molecular basis for the gene-for-gene relationship of Vm-resistance in apple to Venturia inaequalis was investigated. Incompatible reactions involved a hypersensitive response (HR), which was accompanied by the accumulation of dark brown pigments and autofluorescent materials in epidermal and mesophyll cells at the site of invasion. Cell-free culture filtrates of the avirulent isolate elicited an HR in the Vm host (h5) leaves, but not in the susceptible host (h1). The elicitor activity was resistant to boiling but was abolished by proteinase K digestion. Elicitation of HR was used to monitor purification of the avirulence factor, AVRVm, from liquid cultures of the avirulent isolate following ultrafiltration, acetone precipitation and ion-exchange chromatography. The purest fraction contained three major proteins all with low isoelectric points (pI 3.0-4.5). The fraction also elicited HR on the differential host h4, but not on other resistant hosts (h2, h3 and h6) tested. Three candidate AVRVm proteins were identified and amino acid sequences were obtained using Edman degradation and mass spectrometry. Nucleotide sequences corresponding to these proteins were found in databases of V. inaequalis expressed sequence tags. There were no polymorphisms evident between avirulent and virulent isolates (representing races 1 and 5 respectively) either at genomic DNA or cDNA level of the full open reading frames. RT-PCR revealed that all genes were expressed in both avirulent and virulent isolates during in vitro and in planta growth. All three genes showed similar levels of expression between avirulent and virulent isolates during their in vitro growth. However, preliminary RT-PCR experiments showed that two of these genes were likely to be expressed at lower levels in the virulent compared with the avirulent isolate during compatible infection. Implications of this difference in expression and the future experiments to identify the genuine AvrVm gene were discussed.

This thesis adds to the discussion surrounding how Apple Inc. has been able to to garner and maintain such a loyal brand following. With the help of background from theory on branding, it examines the uses of Michel Foucault’s heterotopia and religious allegory within Apple’s branding strategies. With the help of past discussions surrounding these topics, the presence of these uses in present day Apple branding is exemplified in the description and analysis of Apple’s 2014 “Your Verse Anthem” and the Apple Store as a branded space. Through this analysis, this thesis also makes an original argument for a consistent Apple heterotopia that is inherently religious. This heterotopia mirrors the story of Adam and Eve as, within its story, Apple promises that if their consumers will become loyal to the Apple brand by taking a bit out of Apple’s “forbidden fruit,” signified in their logo, Apple will help them see the truth of the Tree of Knowledge, which will help them understand that they need not conform but can instead use Apple products to realize their aspirations and create a life that represents their own version of utopia. This characteristically Apple religious heterotopia and the tactics used by Apple as a whole come together to draw the connection between Apple and the strategies used by cult leaders. This connection ultimately helps explain the loyalty and fever behind Apple’s consumer following.

The diploma thesis concerns with influence of Apple brand on consumer buying behavior. The aim of thesis is to qualify the influence of brand on consumer buying behavior on smartphone market. From this conditions will arise the recommendations to Apple company, how is possible to bolster its position on mobile phone market. The theoretical part of thesis place emphasis on description of marketing mix, brand theory and consumer buying behavior. Practical part is based on studied secondary data. There is detailed description of Apple company its brand building strategy, situation on smartphone market with marketing instruments, that are used on this specific market. Quantitative questionnaire survey performed in electronical form creates a core of the practical part of the thesis and is preceded by qualitative pre-survey on seven iPhone users. Precisely 154 respondents participated on questionnaire survey that observed thoughts and judgements of questioned people about Apple brand. There are included opinions of owners, that own various smartphone brands and also respondents from the group of Apple iPhone users. The responses of both groups are in the chapter of appraisal results and conditions compared along with studied secondary data. These outcomes of questionnaire survey and secondary data are used as a framework to completion several recommendations for Apple brand and the whole company on this market. Almost three quarters of owners, that use all chosen smartphone brands, perceive the prices of Apple smartphones as overly high. Just as the first group evaluates the situation a bit more than half of the iPhone users, although inside this group is a lower sensitivity on Apple products price perception. One of the important recommendation for Apple company is to be more efficient in communication about the adjustment of its smartphone prices and pricing policy towards to consumers across the countries. It is necessary to obtain more consumers from a group of new Apple brand users and students. These expects the offer of cheaper smartphone in high quality medium category. In case of their satisfaction is there scope for Apple to offer them its premium smartphones in the top category.

Os aspectos que envolvem a discussão acerca da diminuição da fome e da miséria em todo o mundo passam pelo entendimento de como se comportam os produtores de alimentos e, por consequência, da sua manutenção no campo. Em se tratando de produtores de alimentos, o entendimento de como se estruturam os canais de distribuição se faz importante à compreensão de como se organiza o poder econômico nessas configurações inter-organizacionais e, por conseguinte, as consequências sociais daí depreendidas. Considerando a cultura da maçã dentre as mais importantes produções de frutas in natura do estado do Rio Grande do Sul e das mais consumidas no Brasil e no mundo, vislumbrar essa estrutura do canal de distribuição e o modo como está estruturado e composto mostrou-se o desafio a ser vencido na pesquisa realizada. Para tanto, o uso e aplicação de métodos de pesquisa que passam desde questionários até entrevistas não-estruturadas permitiu, a partir de uma concepção inicial da estrutura de um canal de comercialização genérico, buscar um desenho e o entendimento das relações entre os players do canal de distribuição da maçã produzida no município de Vacaria (maior produtor gaúcho de maçã e entre os maiores do Brasil). Os resultados indicaram que as relações entre os membros do canal são estabelecidas uma a uma e foram se fortalecendo ao longo dos anos para aqueles players que conseguiram manter-se no mercado. A substituição e renovação daqueles que não conseguiram se manter fortes economicamente aparentaram ser efetivos, como uma Lei de Seleção Natural desenvolvida por Charles Darwin. O divisor entre a manutenção e a extinção passa pelo o uso de tecnologias refrigeradas de transporte e armazenamento. O uso dessas tecnologias reconfiguraram o setor e representaram avanços significativos ao processo de crescimento e internacionalização da produção de maçã brasileira, especificamente de Vacaria. Finalmente, a característica dos grandes players existentes no canal o diferencia de outros canais de distribuição. / Aspects involved in the global discussion regarding hunger and extreme poverty reduction must be addressed using a framework that considers the behavior of food producers and consequently their conditions in the field. When dealing with such a sample, understanding the structure of distribution channels is mandatory for the comprehension of economic power´s organization in these interorganizational configurations and its inherent social outcomes. Considering the apple as one of the most relevant in natura cultures in the State of Rio Grande do Sul, Brazil, as well as one with the highest levels of consumption in Brazil and in the world, analyzing its distribution channel structure and the way it is organized was the central challenge that was dealt with in this research. Hence, the use of methodology based on questionnaires and non-structured interviews allowed – through the use of generic distribution channel structures´ concept – to achieve a robust design and comprehension of relationships between agents of the apple´s distribution channel in the township of Vacaria (main apple producer in the State of Rio Grande do Sul and one of the most important nationwide). Results suggest that relationships between channel members are established on a one-by-one basis and were strengthened over time for those agents who were able to maintain their market position. Substitution and renewal were effective processes for those who did not manage to sustain economic performance – as in an analogy of the Theory of Natural Selection developed by Charles Darwin. The core determinant for market position preservation (or its loss) seems to be the use of refrigerated transport and storage. The use of such technologies reshapes this sector´s structure and represents significant advances in the growth and internationalization processes of Brazilian apple production, specifically for the case of Vacaria. Lastly, characteristics of large agents in this distribution channel pose some specificities when comparing to other channels and sectors.