No description available.

LD2668 .T4 1947 F56 / Master of Science

1. A seed vigour test provides a more sensitive measurement of seed quality than the standard germination test. The cold test is one of the most widely used vigour tests for maize but procedures had not been standardized. In this study different cold test methods were compared for their usefulness as a seed vigour test. 2. Field emergence of 12 hybrid seed lots and 10 parent line seed lots were determined in four different field trials, representing different environmental conditions. Conditions were cold and wet, cool and wet, and favourable. Percentage emergence and emergence rates were determined for the different trials. 3. Considerable differences were found between percentage emergence in the different trials and the standard germination test results. The lowest emergence counts as well as emergence rates were obtained with the oldest seed lots. The differences found in emergence counts between the different seed lots appear to be the result of ageing and dete¬rioration and thus differences in vigour. The ranking order of the different seed lots according to percentage emergence were similar for the different trials. 4. Six different cold test methods and the standard germination test were conducted in the laboratory and the results of the different tests were correlated with field emergen¬ce. The deep-box and rolled towel methods were used and the germination substrates used were sand, a sand-soil mixture, vermiculite, a vermiculite-soil mixture and paper towels with and without soil. 5. Considerable differences were found in percentage emergence between the different laboratory tests. The lowest vigour percentages were obtained with the older seed lots, thus a decline in percentage emergence with an increase in age of the seed lot. The same tendency was found with the field emergence trials. The ranking order of the seed lots did not show large differences between the different tests. 6. Lower vigour percentages were obtained when soil was included in the germination medium and this effect was more pronounced with the older seed lots. The sand + soil cold test was found to be the strictest of the cold tests with the lowest percentage emergence. 7. Correlations were determined between the results of the different cold tests and emergence in the different field trials. All correlation coefficients of the hybrid seed lots were found to be significant at the 1 % level. However, the correlation coefficients obtained between the standard germination test and field emergence were generally of a lower order than those obtained with the cold tests. No significant correlations were 61 found between emergence and the germination percentages of the parent line seed lots. 8. Although the correlations between the different laboratory test results and field emergence do not provide ground for a clear recommendation of adoption of one cold test procedure over another, inclusion of soil in the germination medium resulted in a small increase in correlation coefficients. Correlation coefficients between emergence and the sand + soil cold test were the highest in most of the trials . 9. Although any of the cold tests which include soil in the medium appear to be suitable for adoption by South African seed laboratories, the vermiculite + soil test has advantages over the sand + soil and paper + soil tests. / Dissertation (MSc (Botany))--University of Pretoria, 2006. / Plant Science / unrestricted

This item was digitized as part of the Million Books Project led by Carnegie Mellon University and supported by grants from the National Science Foundation (NSF). Cornell University coordinated the participation of land-grant and agricultural libraries in providing historical agricultural information for the digitization project; the University of Arizona Libraries, the College of Agriculture and Life Sciences, and the Office of Arid Lands Studies collaborated in the selection and provision of material for the digitization project.

Graduation date: 1994

O molibdênio é um dos micronutrientes essenciais para a soja, atuando diretamente no metabolismo do nitrogênio como cofator benzimático da nitrogenase. Usualmente, este nutriente é fornecido às plantas via tratamento de sementes ou aplicação foliar. O objetivo do presente estudo foi avaliar os efeitos do molibdênio via foliar no potencial fisiológico de sementes de soja e verificar sua interferência na atividade das enzimas envolvidas no metabolismo do nitrogênio. Foram utilizadas sementes de soja da cultivar BMX Turbo, produzidas em Erechim, RS, safra 2013, provenientes de plantas tratadas com as seguintes concentrações de Mo: 0; 25; 50 e 75 g ha-1, fornecidas por meio de dois produtos comerciais (Biomol e Molybdate) e armazenadas durante 0 e 6 meses em condições não controladas. O primeiro experimento foi conduzido no Laboratório de Análise de Sementes Seedtes em Pato Branco, PR. O delineamento utilizado foi inteiramente casualizado em esquema fatorial 4 x 2 x 2 com quatro repetições cada. O potencial fisiológico das sementes foi avaliado por meio do teste de germinação, crescimento de plântulas, envelhecimento acelerado e emergência em solo. O segundo experimento foi realizado em casa de vegetação onde foram cultivadas em vasos as sementes oriundas dos tratamentos com diferentes concentrações de Mo: 0; 25; 50 e 75 g ha-1 fornecidas por meio de dois produtos comerciais (Biomol e Molybdate). O delineamento utilizado foi inteiramente casualizado em esquema fatorial 4 X 2 com quatro repetições. As avaliações foram realizadas quando as plantas atingiram o estadio fenológico R1 quanto à nodulação, massa seca de raiz e parte aérea das plantas e determinação da atividade das enzimas glutamina sintetase e glutamato sintetase e o teor de proteínas solúveis totais. Os dados foram submetidos à análise de variância e quando significativos foram avaliados pelo teste de Tukey para comparação dos produtos e do armazenamento de sementes e com estudo de regressão para as concentrações ao nível de 5% de probabilidade. As análises foram realizadas com o software estatístico SISVAR. O armazenamento de sementes de soja em condições não controladas afetou o vigor das sementes produzidas com Mo, independente do produto comercial utilizado durante a produção. A aplicação de Mo via foliar influencia positivamente a produção de sementes de soja que apresentaram respostas crescentes na germinação e no vigor com a aplicação de Mo acima de 25 g ha-1. O enriquecimento de Mo via foliar não afetou a nodulação das plantas da geração seguinte, porém, a utilização de Mo acima de 25 g ha-1 proporcionou aumento na atividade das enzimas envolvidas no metabolismo do nitrogênio, bem como no teor de proteínas totais. / Molybdenum is one of the essential micronutrients for soybeans, acting directly on nitrogen metabolism as enzyme cofactor of nitrogenase. Usually, this nutrient is supplied to the plants through seed treatment or foliar application. The aim of this study was to evaluate the molybdenum effects by foliar in the physiological potential of soybean seeds and verify its interference in the enzyme activities involved in nitrogen metabolism. Soybean seeds of BMX Turbo cultivar were used, produced in Erechim, RS, harvest 2013, from plants treated with the following Mo concentrations: 0; 25; 50 and 75 g ha-1, supplied through two commercial products (Biomol and Molybdate) and stored during 0 and 6 months in uncontrolled conditions. The first experiment was conducted in Seedtes Seed Analysis Laboratory in Pato Branco, PR. The used design was completely randomized in a factorial analysis 4 x 2 x 2 with four replications each. The physiological potential of the seeds was evaluated by the germination test, seedling growth, accelerated aging and emergence on the soil. The second experiment was conducted in a greenhouse, where the seeds derived from treatments with different concentrations of Mo: 0; 25; 50 and 75 g ha-1 supplied through two commercial products (Biomol and Molybdate) were grown in vases. The used design was completely randomized in a factorial analysis 4 x 2 with four replications. Evaluations were performed when the plants reached the R1 phenological stage concerning the nodulation, dry matter of root and shoot of the plants and the determination of the activity of the enzymes glutamine synthetase and glutamate synthetase and the content of total soluble proteins. The data were submitted to variance analysis and when significant they were assessed by Tukey’s test for comparison of products and seed storage and with regression study to the concentrations at 5% probability. Analyses were performed using SISVAR statistical software. The soybean seed storage under uncontrolled conditions affected the seed vigour produced with Mo, regardless of the commercial product used during production. The application of Mo through foliar positively influences the production of soya beans which presented increasing responses in the germination and vigour with the application of Mo above 25 g ha-1 . The enrichment of Mo through foliar did not affect the nodulation of plants of the next generation, however, the use of Mo above 25 g ha-1 provided an increase in the activity of enzymes involved in nitrogen metabolism as well as on the total protein content.

As fruteiras nativas brasileiras mesmo apresentando potencial de uso na agricultura, têm sido pouco exploradas comercialmente. Neste sentido, devem-se estimular estudos que permitam fomentar futuros programas de melhoramento e tecnologias para sua produção. O objetivo deste trabalho foi estudar o comportamento fisiológico de sementes de fruteiras do Bioma Floresta com Araucária, bem como, testar métodos para conservação das mesmas. O trabalho foi conduzido no Laboratório de Fisiologia Vegetal e no Viveiro de Produção de Mudas da Universidade Tecnológica Federal do Paraná – Câmpus Dois Vizinhos – Paraná. Foram utilizadas sementes de frutos maturos fisiologicamente das fruteiras nativas guabijuzeiro, guabirobeira, pitangueira, jabuticabeira de cabinho, jabuticabeira híbrida, cerejeira-do-mato, ameixeira da mata e sete capoteiro. Os frutos foram coletados conforme época de suas respectivas frutificações, em produtores da região Sudoeste do Paraná. Foi realizado quatro experimentos, sendo o primeiro para determinação do teor de umidade das sementes, onde adotou-se o delineamento inteiramente ao acaso, com 12 tratamentos e quatro repetições de 100 sementes. O segundo foi para verificar a longevidade das sementes durante 18 meses, por meio do armazenamento das mesmas em sacos de papel e garrafas PET® como embalagens e, adotando-se o uso ou não de fécula de mandioca como biofilme. O experimento foi instalado em delineamento inteiramente casualizado, em trifatorial 2 x 2 x 18 (embalagem de armazenamento x biofilme x período de armazenamento), com quatro repetições de 100 sementes. No terceiro fez-se estudo da dormência, testando técnicas para quebra da dormência em sementes destas oito fruteiras nativas, utilizando o uso de luz e sem luz. O experimento foi instalado em delineamento inteiramente casualizado, em esquema fatorial 2 x 7 (luminosidade x tratamento para quebra da dormência), com quatro repetições de 100 sementes. O quarto trabalho foi realizado para validação do teste de tetrazólio em sementes de jabuticabeira e cerejeira-da-mata armazenadas. A maior e mais rápida redução do teor de umidade diminuiu a capacidade germinativa das sementes de guabijuzeiro, jabuticabeiras híbrida e de cabinho e ameixeira-da-mata. A garrafa PET® permitiu maior conservação aumentando o número de sementes germinadas. O biofilme pode ser considerada técnica promissora para aumentar a eficiência no processo germinativo de cerejeira-da-mata. As sementes das espécies nativas estudadas não apresentam dormência, exceção para sete capoteiro que apresentaram dormência fisiológica controlada pelo fotoblastismo negativo, já que somente germinaram na ausência de luz. As sementes de cerejeira-da-mata (E. involucrata) podem ser conservadas em temperatura controlada (6°C) por pelo menos 45 dias. Sementes de jabuticabeira de cabinho (P. trunciflora) embaladas a vácuo conservam sua viabilidade por pelo menos 35 dias. O teste Tetrazólio apresentou-se viável e mais rápido para avaliar a viabilidade das sementes de cerejeira-da-mata e de jabuticabeira de Cabinho. / The Brazilian native fruits have potential use in agriculture, but it is little exploited commercially. In this sense, it should be encouraged studies for use in future breeding programs and obtain of technologies for its cultive. The aim of this work was to study the seed physiological characterization from native fruits Araucaria Forest Biome, as well as, it to test methods for seed storage. The work was carried out at Plant Physiology Laboratory and Nursery Sector of UTFPR - Câmpus Dois Vizinhos, Paraná State, Brazil. It was use seeds physiologically mature fruits Brazilian native from guabiju tree, guabiroba tree, Surinam Cherry tree, Cabinho and Hybrid jabuticaba tree, Native cherry tree, Native plum tree and sete capote tree. The fruits were harvested according period yield, in Southwest region of Paraná State. It was carried out four experiments, it being the first to identify the critical moisture content that affect seed germination, through the experimental design completely randomized, with 12 treatments and 4 replications of one hundred seeds by plot. The second was to evaluated the seed viability during 18 months through the PET® bottle (300 mL) and white waxed paper bag as storage packing and, the cassava starch biofilm application or not. The experimental design was blocks completely randomized, in factorial 2. x 2 x 18 (storage packaging x biofilm x storage time), with 4 replications of one hundred seeds by plot. In the third was dormancy study through it technical test for seeds break dormancy from eight fruit Brazilian trees, with or without light. The fourth experiment was realized for Tetrazolium test validation with jabuticaba fruit and Native cherry seeds stored. The greater and more rapid moisture content reduction decreased the guabiju seed tree, Hybrid and Cabinho seeds jabuticaba tree and Native plum tree germination process. However, new studies will be necessary to assess the seed viability on condition of lower moisture content. The PET® bottle permitted greater conservation, it increased the seed germination. The biofilm can be consider promising technique to increase germination efficiency of Native fruit cherry. The native Brazilian fruit seed species didn't have dormancy, except for sete capote fruit tree that presented morf were considered capoteiro showed physiological dormancy controlled by photoblastic negative response, it have germination only in the light absence. The Cabinho jabuticaba fruit (P. trunciflora) seeds vacuum packed maintains its viability about thirty five days. The Tetrazolium test presented viable and faster to evaluated the Native cherry and Cabinho jabuticaba fruit seeds viability.

O molibdênio é um dos micronutrientes essenciais para a soja, atuando diretamente no metabolismo do nitrogênio como cofator benzimático da nitrogenase. Usualmente, este nutriente é fornecido às plantas via tratamento de sementes ou aplicação foliar. O objetivo do presente estudo foi avaliar os efeitos do molibdênio via foliar no potencial fisiológico de sementes de soja e verificar sua interferência na atividade das enzimas envolvidas no metabolismo do nitrogênio. Foram utilizadas sementes de soja da cultivar BMX Turbo, produzidas em Erechim, RS, safra 2013, provenientes de plantas tratadas com as seguintes concentrações de Mo: 0; 25; 50 e 75 g ha-1, fornecidas por meio de dois produtos comerciais (Biomol e Molybdate) e armazenadas durante 0 e 6 meses em condições não controladas. O primeiro experimento foi conduzido no Laboratório de Análise de Sementes Seedtes em Pato Branco, PR. O delineamento utilizado foi inteiramente casualizado em esquema fatorial 4 x 2 x 2 com quatro repetições cada. O potencial fisiológico das sementes foi avaliado por meio do teste de germinação, crescimento de plântulas, envelhecimento acelerado e emergência em solo. O segundo experimento foi realizado em casa de vegetação onde foram cultivadas em vasos as sementes oriundas dos tratamentos com diferentes concentrações de Mo: 0; 25; 50 e 75 g ha-1 fornecidas por meio de dois produtos comerciais (Biomol e Molybdate). O delineamento utilizado foi inteiramente casualizado em esquema fatorial 4 X 2 com quatro repetições. As avaliações foram realizadas quando as plantas atingiram o estadio fenológico R1 quanto à nodulação, massa seca de raiz e parte aérea das plantas e determinação da atividade das enzimas glutamina sintetase e glutamato sintetase e o teor de proteínas solúveis totais. Os dados foram submetidos à análise de variância e quando significativos foram avaliados pelo teste de Tukey para comparação dos produtos e do armazenamento de sementes e com estudo de regressão para as concentrações ao nível de 5% de probabilidade. As análises foram realizadas com o software estatístico SISVAR. O armazenamento de sementes de soja em condições não controladas afetou o vigor das sementes produzidas com Mo, independente do produto comercial utilizado durante a produção. A aplicação de Mo via foliar influencia positivamente a produção de sementes de soja que apresentaram respostas crescentes na germinação e no vigor com a aplicação de Mo acima de 25 g ha-1. O enriquecimento de Mo via foliar não afetou a nodulação das plantas da geração seguinte, porém, a utilização de Mo acima de 25 g ha-1 proporcionou aumento na atividade das enzimas envolvidas no metabolismo do nitrogênio, bem como no teor de proteínas totais. / Molybdenum is one of the essential micronutrients for soybeans, acting directly on nitrogen metabolism as enzyme cofactor of nitrogenase. Usually, this nutrient is supplied to the plants through seed treatment or foliar application. The aim of this study was to evaluate the molybdenum effects by foliar in the physiological potential of soybean seeds and verify its interference in the enzyme activities involved in nitrogen metabolism. Soybean seeds of BMX Turbo cultivar were used, produced in Erechim, RS, harvest 2013, from plants treated with the following Mo concentrations: 0; 25; 50 and 75 g ha-1, supplied through two commercial products (Biomol and Molybdate) and stored during 0 and 6 months in uncontrolled conditions. The first experiment was conducted in Seedtes Seed Analysis Laboratory in Pato Branco, PR. The used design was completely randomized in a factorial analysis 4 x 2 x 2 with four replications each. The physiological potential of the seeds was evaluated by the germination test, seedling growth, accelerated aging and emergence on the soil. The second experiment was conducted in a greenhouse, where the seeds derived from treatments with different concentrations of Mo: 0; 25; 50 and 75 g ha-1 supplied through two commercial products (Biomol and Molybdate) were grown in vases. The used design was completely randomized in a factorial analysis 4 x 2 with four replications. Evaluations were performed when the plants reached the R1 phenological stage concerning the nodulation, dry matter of root and shoot of the plants and the determination of the activity of the enzymes glutamine synthetase and glutamate synthetase and the content of total soluble proteins. The data were submitted to variance analysis and when significant they were assessed by Tukey’s test for comparison of products and seed storage and with regression study to the concentrations at 5% probability. Analyses were performed using SISVAR statistical software. The soybean seed storage under uncontrolled conditions affected the seed vigour produced with Mo, regardless of the commercial product used during production. The application of Mo through foliar positively influences the production of soya beans which presented increasing responses in the germination and vigour with the application of Mo above 25 g ha-1 . The enrichment of Mo through foliar did not affect the nodulation of plants of the next generation, however, the use of Mo above 25 g ha-1 provided an increase in the activity of enzymes involved in nitrogen metabolism as well as on the total protein content.

As fruteiras nativas brasileiras mesmo apresentando potencial de uso na agricultura, têm sido pouco exploradas comercialmente. Neste sentido, devem-se estimular estudos que permitam fomentar futuros programas de melhoramento e tecnologias para sua produção. O objetivo deste trabalho foi estudar o comportamento fisiológico de sementes de fruteiras do Bioma Floresta com Araucária, bem como, testar métodos para conservação das mesmas. O trabalho foi conduzido no Laboratório de Fisiologia Vegetal e no Viveiro de Produção de Mudas da Universidade Tecnológica Federal do Paraná – Câmpus Dois Vizinhos – Paraná. Foram utilizadas sementes de frutos maturos fisiologicamente das fruteiras nativas guabijuzeiro, guabirobeira, pitangueira, jabuticabeira de cabinho, jabuticabeira híbrida, cerejeira-do-mato, ameixeira da mata e sete capoteiro. Os frutos foram coletados conforme época de suas respectivas frutificações, em produtores da região Sudoeste do Paraná. Foi realizado quatro experimentos, sendo o primeiro para determinação do teor de umidade das sementes, onde adotou-se o delineamento inteiramente ao acaso, com 12 tratamentos e quatro repetições de 100 sementes. O segundo foi para verificar a longevidade das sementes durante 18 meses, por meio do armazenamento das mesmas em sacos de papel e garrafas PET® como embalagens e, adotando-se o uso ou não de fécula de mandioca como biofilme. O experimento foi instalado em delineamento inteiramente casualizado, em trifatorial 2 x 2 x 18 (embalagem de armazenamento x biofilme x período de armazenamento), com quatro repetições de 100 sementes. No terceiro fez-se estudo da dormência, testando técnicas para quebra da dormência em sementes destas oito fruteiras nativas, utilizando o uso de luz e sem luz. O experimento foi instalado em delineamento inteiramente casualizado, em esquema fatorial 2 x 7 (luminosidade x tratamento para quebra da dormência), com quatro repetições de 100 sementes. O quarto trabalho foi realizado para validação do teste de tetrazólio em sementes de jabuticabeira e cerejeira-da-mata armazenadas. A maior e mais rápida redução do teor de umidade diminuiu a capacidade germinativa das sementes de guabijuzeiro, jabuticabeiras híbrida e de cabinho e ameixeira-da-mata. A garrafa PET® permitiu maior conservação aumentando o número de sementes germinadas. O biofilme pode ser considerada técnica promissora para aumentar a eficiência no processo germinativo de cerejeira-da-mata. As sementes das espécies nativas estudadas não apresentam dormência, exceção para sete capoteiro que apresentaram dormência fisiológica controlada pelo fotoblastismo negativo, já que somente germinaram na ausência de luz. As sementes de cerejeira-da-mata (E. involucrata) podem ser conservadas em temperatura controlada (6°C) por pelo menos 45 dias. Sementes de jabuticabeira de cabinho (P. trunciflora) embaladas a vácuo conservam sua viabilidade por pelo menos 35 dias. O teste Tetrazólio apresentou-se viável e mais rápido para avaliar a viabilidade das sementes de cerejeira-da-mata e de jabuticabeira de Cabinho. / The Brazilian native fruits have potential use in agriculture, but it is little exploited commercially. In this sense, it should be encouraged studies for use in future breeding programs and obtain of technologies for its cultive. The aim of this work was to study the seed physiological characterization from native fruits Araucaria Forest Biome, as well as, it to test methods for seed storage. The work was carried out at Plant Physiology Laboratory and Nursery Sector of UTFPR - Câmpus Dois Vizinhos, Paraná State, Brazil. It was use seeds physiologically mature fruits Brazilian native from guabiju tree, guabiroba tree, Surinam Cherry tree, Cabinho and Hybrid jabuticaba tree, Native cherry tree, Native plum tree and sete capote tree. The fruits were harvested according period yield, in Southwest region of Paraná State. It was carried out four experiments, it being the first to identify the critical moisture content that affect seed germination, through the experimental design completely randomized, with 12 treatments and 4 replications of one hundred seeds by plot. The second was to evaluated the seed viability during 18 months through the PET® bottle (300 mL) and white waxed paper bag as storage packing and, the cassava starch biofilm application or not. The experimental design was blocks completely randomized, in factorial 2. x 2 x 18 (storage packaging x biofilm x storage time), with 4 replications of one hundred seeds by plot. In the third was dormancy study through it technical test for seeds break dormancy from eight fruit Brazilian trees, with or without light. The fourth experiment was realized for Tetrazolium test validation with jabuticaba fruit and Native cherry seeds stored. The greater and more rapid moisture content reduction decreased the guabiju seed tree, Hybrid and Cabinho seeds jabuticaba tree and Native plum tree germination process. However, new studies will be necessary to assess the seed viability on condition of lower moisture content. The PET® bottle permitted greater conservation, it increased the seed germination. The biofilm can be consider promising technique to increase germination efficiency of Native fruit cherry. The native Brazilian fruit seed species didn't have dormancy, except for sete capote fruit tree that presented morf were considered capoteiro showed physiological dormancy controlled by photoblastic negative response, it have germination only in the light absence. The Cabinho jabuticaba fruit (P. trunciflora) seeds vacuum packed maintains its viability about thirty five days. The Tetrazolium test presented viable and faster to evaluated the Native cherry and Cabinho jabuticaba fruit seeds viability.

The pattern and sequence of zein degradation in the endosperm of germinating maize seeds were investigated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting. The proteases involved in the degradation of various zein components (α, ß and γ) were extracted with three buffer systems and partially characterized with respect to their ability to degrade various zein components. They were also investigated in vivo by germinating the seeds in the presence of protease inhibitors used singly and in combination. Of the various zein components, γ-zein (27-kD) was the first to be degraded and its degradation was complete by the third day after germination (DAG). Beta-zeins (17- and 18-kD) began to be degraded on the second DAG, degradation being complete by the seventh day for the l7-kD polypeptide, and the fourth day for the 18-kD polypeptide. The degradation of 10-kD- zein began on the fourth DAG and was complete by the eighth day. The α-zein fraction (22-and 24-kD) was degraded beginning on the faith day and continued gradually until after the tenth day. From the results of these studies, the arrangement of various zein fractions within the protein bodies can be deduced and this was consistent with the immunocytochemical data published by others. Gamma-zein is situated in the peripheral region of the protein bodies and could be a structural component of the protein body membrane or it may be directly anchored in the membrane. Beta-zeins are internal to γ-zein with the l0-kD in the interface between the 17-kD and γ-zein. The 10- kD zein is located between the 17-kD and α-zein or interlacing with α-zein in the protein body core. Finally, a-zeins are in the protein body core. Based on these observations the proteolysis of the protein in protein bodies of maize would start from the periphery and proceed towards their core. The proteases involved in degradation of various zein components were synthesized de novo. The mRNAs pre-existing in dry seeds appeared to direct the synthesis of active proteases required for zein degradation at least during the initial stages of germination. Serine protease was responsible for the degradation of a- and ß-zeins while aspartic (acid) protease may play some role in ß-zein degradation. Serine and cysteine (thiol) proteases worked synergistically in γ-zein degradation. Enzymes extractable from the endosperm of germinating seeds with 0.2 M acetate buffer (pH 3.8) were able to degrade the α-, ß-, and γ-zeins in an in vito assay. / Ph. D.