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No description available.

衰老是一個複雜的發病過程，它由多種因素引發，主要包括基因和遺傳。已經有很多研究報告表明能量限制（ER）可以延長果蠅的壽命，而芝麻素和黑米具有抗氧化活性。但是，基本的機制仍然不明。因此本研究檢驗ER，芝麻素和黑米萃取物（BRE）的抗衰老活性，並探討他們如何與CuZnSOD (SOD1)，MnSOD (SOD2)，catalase (CAT)，Methuselah（Mth）和Rpn11五個基因相互作用，參與果蠅的抗氧化防禦和衰老。 / 第一部分研究能量限制的抗衰老機制。果蠅隨機分成三組並餵養低能量 (0.393 kcal/ml), 標準能量 (SE; 0.784 kcal/ml) 和高能量(HE; 2.351 kcal/ml) 的食物。 能量減半的食物可以延長果蠅的平均壽命16%，這種作用伴隨著SOD1, SOD2 和Rpn11基因表達的上升。味覺試驗表明，能量限制的延長壽命作用與它顯著減少食物攝入量有關。急性paraquat實驗表明，能量限制可以延長野生型果蠅的壽命，而對超氧化物歧化酶突變果蠅的死亡無影響。同時，低能量的食可以緩解，而高能量的食則加速與年齡相關的攀爬能力的喪失。結果表明，能量限制可以延緩果蠅衰老，這種作用至少部分是由上調抗氧化基因的表達調控的。 / 第二部分研究芝麻素對果蠅壽命的影響。結果顯示，添加2和 5 mg/ml的芝麻素分別增加果蠅的平均存活時間13%和5%。進一步的研究表明，2 mg/ml芝麻素的抗衰老作用是通過上調SOD1，SOD2，CAT和Rpn11的基因表達實現的。另外，芝麻素可以延緩野生型果蠅的由paraquat引發的神經退行性疾病的進展，並且上調SOD1，SOD2和Rpn11的基因表達。急性paraquat實驗結果表明，芝麻素可以延長野生型和Aβ42 33769突變果蠅的壽命。由此得出結論，芝麻素可以延長果蠅壽命，並且減輕野生型果蠅的由paraquat引發的神經退行性疾病的症狀，這些作用至少部分是由基因SOD1，SOD2，CAT和Rpn11，而不是Mth調控的。 / 第三部分探討黑米萃取物的抗衰老作用。添加30mg/ml 的黑米萃取物可以延長果蠅的平均壽命14%，這是通過上調SOD1，SOD2，CAT和Rpn11，下調Mth基因的表達實現的。同時，黑米萃取物可以延緩野生型果蠅的由paraquat引發的神經退行性疾病的進展，伴隨著上調SOD1，SOD2和Rpn11的基因表達。 此外，補充黑米萃取物可以增加野生型和Aβ42 33769突變果蠅的生存時間。結果表明，黑米萃取物可以延長果蠅壽命，並且延緩野生型果蠅的由paraquat引發的神經退行性疾病的進程，這些作用至少部分是由基因SOD1，SOD2，CAT，Mth和Rpn11調控的。 / 總之，本研究揭示能量限制，補充抗氧化劑芝麻素和黑米萃取物可以改變黑腹果蠅的壽命。這些作用部分是由基因SOD1, SOD2, CAT, Rpn11 和Mth調控的。 / Aging is a complicated pathogenesis that is triggered by multiple factors mainly including genetics and environment. There have been numerous reports that demonstrate Energy Restriction (ER) could extend the lifespan of fruit fly, and sesamin and black rice possess antioxidant activity. However, the underlying mechanism remains unknown. The present study was therefore to examine the anti-aging activity of ER, sesamin and black rice extracts (BRE) and to investigate how they interacted with genes of CuZnSOD (SOD1), MnSOD (SOD2), catalase (CAT), Methuselah (Mth) and Rpn11 involved in the antioxidant defense and aging of Drosophila melanogaster. / Part I was to investigate the mechanism by which ER prolonged the lifespan of fruit fly. Fruit flies were divided into three groups and given one of three diets namely ER diet (0.393 kcal/ml), standard energy diet (SE; 0.784 kcal/ml) and high energy diet (HE; 2.351 kcal/ml). It was found that ER extended the mean lifespan by 16%, with elevated expressions of SOD1, SOD2 and Rpn11. Gustatory assay showed that the lifespan extension of ER was not related to the significantly less food intake. In addition, ER prolonged the lifespan of OR wild type fly, but not that of SOD mutant in the intensive paraquat test. Meanwhile, the ER diet could improve, while HE diet accelerated the age-dependent loss of climbing activity in OR wild type fly. Results confirmed that ER could delay the aging of fruit fly, mediated at least in part by up-regulating the genes of antioxidant enzymes. / Part II was to study the effect of sesamin supplementation on the lifespan of fruit fly. Results showed that sesamin at doses of 2 and 5 mg/ml diet increased the mean survival time by 13% and 5%, respectively. Further experiments demonstrated that the lifespan-prolonging activity of 2 mg sesamin/ml diet was accompanied by up-regulation of SOD1, SOD2, CAT and Rpn11. It was further observed that sesamin attenuated the paraquat-induced neurodegeneration in OR wild type fly, with up-regulation of SOD1, SOD2 and Rpn11. Sesamin also increased the survival time of OR wild type fly and Alzheimer mutant fly Aβ42 33769 with intensive paraquat treatment. It was therefore summarized that sesamin extended the lifespan and alleviated the neurodegeration in fruit fly, at least in part resulting from the interactions with genes SOD1, SOD2, CAT and Rpn11, not Mth. / Part III was to investigate the life-prolonging activity of BRE. Addition of 30 mg BRE into 1 ml diet (BRE30) could prolong the mean lifespan of fruit flies by 14%, accompanied with up-regulation of mRNA SOD1, SOD2, CAT and Rpn11, and down-regulation of Mth. It was also found that BRE30 could attenuate the paraquat-induced neurodegeneration in OR wild type fly, with up-regulation of SOD1, SOD2, CAT and Rpn11. In addition, BRE30 diet increased the survival time of OR wild type fly and Alzheimer mutant fly Aβ42 33769 exposed to paraquat. It was concluded that BRE could extend the lifespan and alleviate the neurodegeration in fruit fly, most likely by regulating the genes of SOD1, SOD2, CAT, Mth and Rpn11. / In summary, the present study found that the lifespan of fruit fly could be altered by ER and addition of antioxidants sesamin and BRE. The effect was in part regulated by genes SOD1, SOD2, CAT, Rpn11 and Mth. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Zuo, Yuanyuan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 122-135). / Abstracts also in Chinese. / Acknowledgments --- p.I / Abstract --- p.II / List of abbreviations --- p.VII / Table of Contents --- p.IX / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Human aging models --- p.2 / Chapter 1.2.1 --- Yeasts --- p.3 / Chapter 1.2.2 --- Nematodes --- p.3 / Chapter 1.2.3 --- Drosophila melanogaster --- p.4 / Chapter 1.2.4 --- Rodents --- p.5 / Chapter 1.2.5 --- Primates --- p.6 / Chapter 1.3 --- The aging process and behavioral senesce in D. melanogaster --- p.7 / Chapter 1.4 --- Anti-aging pathways in D. melanogaster --- p.8 / Chapter 1.4.1 --- Inhibition of respiration --- p.8 / Chapter 1.4.2 --- Metabolic rate --- p.9 / Chapter 1.4.3 --- Oxidative stress --- p.10 / Chapter 1.4.4 --- Apoptotic pathways --- p.10 / Chapter 1.4.5 --- The Insulin/IGF-1 like signaling (IIS) pathway --- p.11 / Chapter 1.4.7 --- The sirtuin pathway --- p.12 / Chapter 1.4.8 --- The olfactory system --- p.12 / Chapter 1.5 --- Free radical theory of aging --- p.13 / Chapter 1.5.1 --- Free radicals --- p.14 / Chapter 1.5.2 --- Antioxidant system --- p.19 / Chapter 1.5.2.1 --- Antioxidant enzymes --- p.19 / Chapter 1.5.2.2 --- Non-enzymatic antioxidants --- p.21 / Chapter 1.6 --- Energy restriction --- p.24 / Chapter 1.6.1 --- ER or fasting in humans --- p.24 / Chapter 1.6.2 --- ER in D. melanogaster --- p.25 / Chapter 1.6.3 --- Response of oxidative stress to ER in D. melanogaster --- p.25 / Chapter 1.7 --- Exogenous antioxidants --- p.27 / Chapter 1.7.1 --- Sesamin --- p.30 / Chapter 1.7.2 --- Black rice extracts --- p.30 / Chapter 1.8 --- Age-related biomarkers in D. melanogaster --- p.31 / Chapter 1.8.1 --- Behavioral changes in D. melanogaster --- p.31 / Chapter 1.8.2 --- Paraquat-induced mortality in D. melanogaster --- p.32 / Chapter 1.8.3 --- Age-related genes --- p.32 / Chapter Chapter 2 --- Lifespan extension, oxidative stress and energy restriction in fruit flies --- p.36 / Chapter 2.1 --- Introduction --- p.36 / Chapter 2.2 --- Objectives --- p.39 / Chapter 2.3 --- Materials and methods --- p.39 / Chapter 2.3.1 --- Fly stocks --- p.39 / Chapter 2.3.2 --- Diet --- p.39 / Chapter 2.3.3 --- Lifespan assay --- p.42 / Chapter 2.3.4 --- Measurement of body weight --- p.42 / Chapter 2.3.5 --- Gustatory assay --- p.42 / Chapter 2.3.6 --- Climbing assay --- p.43 / Chapter 2.3.7 --- Paraquat treatment --- p.43 / Chapter 2.3.8 --- SOD activity --- p.44 / Chapter 2.3.9. --- CAT activity --- p.45 / Chapter 2.3.10 --- Real-Time PCR --- p.46 / Chapter 2.3.11 --- Western blot analysis --- p.48 / Chapter 2.3.12 --- Statistics --- p.49 / Chapter 2.4 --- Results --- p.49 / Chapter 2.4.1 --- Lifespan extension of ER in fruit flies --- p.49 / Chapter 2.4.2 --- Changes of food intake and locomotor function in fruit flies --- p.50 / Chapter 2.4.3 --- Resistance to paraquat-induced oxidative stress in fruit flies --- p.50 / Chapter 2.4.4 --- Influence of ER on enzymatic activity, gene expression and protein expression in fruit flies --- p.51 / Chapter 2.5 --- Discussion --- p.60 / Chapter Chapter 3 --- Sesamin extends lifespan of fruit flies --- p.63 / Chapter 3.1 --- Introduction --- p.63 / Chapter 3.2 --- Objectives --- p.69 / Chapter 3.3 --- Materials and methods --- p.69 / Chapter 3.3.1 --- Chemicals --- p.69 / Chapter 3.3.2 --- Fly stocks --- p.69 / Chapter 3.3.3 --- Diet --- p.70 / Chapter 3.3.4 --- Lifespan assay --- p.70 / Chapter 3.3.5 --- Measurement of body weight --- p.71 / Chapter 3.3.6 --- Gustatory assay --- p.71 / Chapter 3.3.7 --- Intensive paraquat treatment --- p.72 / Chapter 3.3.8 --- Chronic paraquat treatment --- p.72 / Chapter 3.3.9 --- Climbing assay --- p.73 / Chapter 3.3.10 --- Diets switch experiment --- p.73 / Chapter 3.3.11 --- SOD activity --- p.74 / Chapter 3.3.12 --- CAT activity --- p.74 / Chapter 3.3.13 --- Real-time PCR --- p.74 / Chapter 3.3.14 --- Western blot analysis --- p.74 / Chapter 3.3.15 --- Statistics --- p.75 / Chapter 3.4 --- Results --- p.75 / Chapter 3.4.1 --- Lifespan extension of sesamin in fruit flies --- p.75 / Chapter 3.4.2 --- Results of diets switch --- p.76 / Chapter 3.4.3 --- Effect of sesamin on intensive paraquat treatment in OR, SOD{U+207F}¹°⁸, Aβ42 32038 and Aβ42 33769 flies --- p.77 / Chapter 3.4.4 --- Effect of sesamin on chronic paraquat treatment in OR flies --- p.78 / Chapter 3.5 --- Discussions --- p.92 / Chapter Chapter 4 --- Black rice extract extends lifespan of fruit flies --- p.96 / Chapter 4.1 --- Introduction --- p.96 / Chapter 4.2 --- Objectives --- p.97 / Chapter 4.3 --- Materials and methods --- p.97 / Chapter 4.3.1 --- Chemicals --- p.97 / Chapter 4.3.2 --- Fly stocks --- p.97 / Chapter 4.3.3 --- Diet --- p.98 / Chapter 4.3.4 --- Lifespan assay --- p.98 / Chapter 4.3.5 --- Measurement of body weight --- p.99 / Chapter 4.3.6 --- Gustatory assay --- p.99 / Chapter 4.3.7 --- Intensive paraquat treatment --- p.99 / Chapter 4.3.8 --- Chronic paraquat treatment --- p.99 / Chapter 4.3.9 --- Climbing assay --- p.100 / Chapter 4.3.10 --- Diets switch experiment --- p.100 / Chapter 4.3.11 --- SOD activity --- p.100 / Chapter 4.3.12 --- Real-time PCR --- p.100 / Chapter 4.3.13 --- Western blot analysis --- p.101 / Chapter 4.3.14 --- Statistics --- p.101 / Chapter 4.4 --- Results --- p.101 / Chapter 4.4.1 --- Lifespan extension of BRE in fruit flies --- p.101 / Chapter 4.4.2 --- Results of diets switch --- p.102 / Chapter 4.4.3 --- Effect of BRE on intensive paraquat treatment in OR, SOD{U+207F}¹°⁸, Aβ42 32038 and Aβ42 33769 flies --- p.103 / Chapter 4.4.4 --- Effect of BRE on chronic paraquat treatment in OR flies --- p.104 / Chapter 4.5 --- Discussion --- p.118 / References --- p.122

by Xie Weijun. / Publication date from spine. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1995. / Includes bibliographical references (leaves 107-122). / ABSTRACT --- p.i / ACKNOWLEDGMENTS --- p.iii / TABLE OF CONTENTS --- p.iv / LIST OF FIGURES --- p.vii / LIST OF TABLES --- p.x / Chapter 1. --- INTRODUCTION --- p.1 / Chapter 2. --- LITERATURE REVIEW --- p.4 / Chapter 2.1 --- Economic and biotechnological significance of Lentinula edodes --- p.4 / Chapter 2.2 --- Biological background --- p.8 / Chapter 2.2.1 --- Life cycle --- p.8 / Chapter 2.2.2 --- Morphological changes during fruit body development --- p.9 / Chapter 2.2.3 --- Biochemistry of fruit body development --- p.13 / Chapter 2.2.4 --- Molecular studies --- p.17 / Chapter 2.3 --- RNA AP-PCR --- p.23 / Chapter 3. --- MATERIALS AND METHODS --- p.26 / Chapter 3.1 --- Biological materials --- p.26 / Chapter 3.2 --- Media --- p.26 / Chapter 3.3 --- RNA AP-PCR with RNAs isolated from four different developmental stages --- p.27 / Chapter 3.3.1 --- Isolation of total RNAs --- p.27 / Chapter 3.3.2 --- RNA AP-PCR --- p.30 / Chapter 3.3.3 --- PCR reamplification --- p.33 / Chapter 3.4 --- PCR cloning and screening --- p.33 / Chapter 3.4.1 --- PCR product purification --- p.34 / Chapter 3.4.2 --- Cloning --- p.34 / Chapter 3.4.3 --- Transformation --- p.36 / Chapter 3.4.4 --- Screening --- p.36 / Chapter 3.4.4.1 --- Restriction enzyme digestion of plasmid DNA --- p.36 / Chapter 3.4.4.2 --- PCR Screening --- p.38 / Chapter 3.5 --- DNA sequencing --- p.38 / Chapter 3.5.1 --- Preparation of double-stranded template --- p.38 / Chapter 3.5.2 --- Sequencing reaction using double-stranded templates --- p.39 / Chapter 3.5.3 --- Electrophoresis --- p.40 / Chapter 3.5.4 --- Autoradiography and analysis --- p.41 / Chapter 3.6 --- Dot-blot analysis --- p.41 / Chapter 3.6.1 --- Dot-blot --- p.41 / Chapter 3.6.2 --- Probe preparation --- p.42 / Chapter 3.6.3 --- Prehybridization and hybridization --- p.43 / Chapter 3.6.4 --- Autoradiography and Analysis --- p.43 / Chapter 3.7 --- Study on one clone: the gene encoding ubiquitin --- p.44 / Chapter 3.7.1 --- "Design of primers:UBR5A, UBR3A and Polyub5A, polyub3A" --- p.44 / Chapter 3.7.2 --- Isolation of DNA from mycelium and fruit body --- p.46 / Chapter 3.7.3 --- Specific PCR with Polyub primers --- p.46 / Chapter 3.7.4 --- RT-PCR --- p.47 / Chapter 4. --- RESULTS --- p.48 / Chapter 4.1 --- Total RNA isolation --- p.48 / Chapter 4.2 --- RNA AP-PCR --- p.52 / Chapter 4.3 --- PCR reamplification --- p.58 / Chapter 4.4 --- Molecular cloning of RAP products --- p.58 / Chapter 4.5 --- Screening --- p.61 / Chapter 4.6 --- DNA sequencing --- p.65 / Chapter 4.7 --- Sequence analyses --- p.80 / Chapter 4.8 --- Dot blot --- p.84 / Chapter 4.9 --- Further study on pMrG290a --- p.84 / Chapter 5. --- DISCUSSION --- p.92 / Chapter 5.1 --- RNA AP-PCR --- p.92 / Chapter 5.2 --- PCR cloning --- p.94 / Chapter 5.3 --- Nucleotide sequencing --- p.95 / Chapter 5.4 --- Dot blot --- p.96 / Chapter 5.5 --- Effect of polyphenol in RNA or DNA samples to the efficiency of reverse transcription and PCR --- p.98 / Chapter 5.6 --- Ubiquitin and fruit body development --- p.99 / Chapter 5.7 --- "Mitochondrial biogenesis, bioenergetics and fruit body development" --- p.102 / CONCLUSION --- p.105 / REFERENCES --- p.107

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This study was aimed at determining the expression and activity of polyphenoloxidase (PPO) during apricot fruit development together with the biochemical characteristics of the enzyme extract at different development stages. Biochemical factors considered include substrate, pH, NaCl level, inhibitor type, high temperature inactivation and sulphur dioxide level. Changes in apricot (Prunus armeniaca L., cv. 'Moorpark') polyphenoloxidase (PPO) were measured during fruit development from a few days after full bloom until over-ripe at 92 days after full bloom. Cold ground amples in McIllvaine's buffer were analysed for PO activity over a range of pH (5.0, 6.0, 6.8 and 7.2); for response to intact fruit sample pre-heating (25, 35, 45, 55 and 65 °C); for sulphite and NaCl inhibition (0.2, 0.5, 2 and 5mM) and other inhibitors (SHAM 0.2mM, cinnamic acid 2.5mM and tropolone 0.5mM). PPO activity was measured at 25°C using a Clark-type oxygen electrode with 4-methyl catechol (20mM) as substrate. As fruit ripened PPO activity increased under all conditions tested. The increase in activity was not even with fruit development. Three common peaks of PPO activity occurred at ages 22-29 days, 57 days and for fully-ripe fruit at 85-92 days. Optimum pH was found to be 6.8 with a wide range for all ages of fruit. PPO activity tended to be higher for more mature fruit at a higher pH of 7.2 to 8.0, whereas activity tended to be higher in less developed fruit at the lower pH of 6.0. Catechol and chlorogenic acid showed reduced PPO activity compared with 4-methyl catechol over all development ages, however, there was a different pattern of response. Both catechol and chlorogenic acid showed greater PPO activity in the fully mature, day 92 fruit and less in the very young day 8 fruit, relative to the control 4-methyl catechol substrate. L-DOPA, as a substrate, showed a reaction lag as previously reported, and quite depressed PPO activity with no particular variation with development age compared to the control. Pre-heating of fruit samples in air for 30 minutes resulted in increased inactivation with holding temperature (35°C - 31%, 45°C - 82%, 55°C - 97%, 65°C - 99%). Sulphite and NaCl acted as inhibitors with increasing effect as concentration increased. Added sulphite depressed PPO activity by about 30% at the level (2mM) used. This was less than the literature would suggest and it appeared that fully-ripe fruit were less inhibited than mature, non-ripe fruit. NaCl has a greater inhibitory effect on apricot PPO activity at the lower pH 5.0 tested. As NaCl added increases PPO activity decreases after an initial small rise. Again, less sensitivity to NaCl inhibition is shown by fruit of greater development age. Sensitivity to inhibition by SHAM, cinnamic acid and tropolone decreased with development age. Tropolone was the most effective inhibitor of apricot PPO. The pattern of change in PPO activity, was consistent with physiological and biochemical changes reported by other workers as fruit develop from hard, green to soft, ripe. Regarding the existence of different PPO isozymes during development, no evidence of a isozyme based differential response with age was found within the constraints of the parameters tested. / Thesis (M.App.Sc.)--School of Agriculture & Wine, 2002.

Thesis (M.S.)--Oregon State University, 1963. / Typescript. Includes bibliographical references (leaves 104-106). Also available on the World Wide Web.

Thesis (Ph. D.)--Ohio State University, 1982. / Includes bibliographical references (leaves 153-156). Available online via OhioLINK's ETD Center

Determination of phylogenetic relationships among ancestral species of Rubus has been elusive. Most Rubus species (including blackberries and raspberries), representing nine of the 12 subgenera, occur in a large, well supported clade named 'A' for reference). The remaining nine species are excluded from this group and represent three subgenera: subg. Anoplobatus (R. bartonianus, R. deliciosus, R. neomexicanus, R. odoratus, R. parviflorus, R. trilobus), subg. Chamaemorus (R. chamaemorus), and subg. Dalibarda (R. lasiococcus, R.pedatus). In addition, Rubus dalibarda L. is often treated in its own monotypic genus as Dalibarda repens L. Phylogenetic analyses of DNA sequence data from chloroplast regions and the nuclear ribosomal DNA internal transcribed spacer ITS 1 - 5.8S - ITS 2; ITS) region have not resolved basal relationships in Rubus and the position of Dalibarda repens has varied from being the sister group to Rubus to nested within it. However, monophyly of American subg. Anoplobatus species is supported by both genomic regions. Our goal was to clarify ancestral relationships, investigate the position of Dalibarda repens relative to Rubus, and examine the origin of the circumboreal, octoploid species R. chamaemorus using sequence data from one additional chloroplast DNA region, trnS-trnG, and the singlecopy nuclear gene Granule-Bound Starch Synthase (GBSSI-1). Parsimony analyses of trnS-trnG sequences indicate a basal trichotomy, while R.chamaemorus is strongly supported as sister to R. pedatus. A combined cpDNA (trnS-trnG and three other regions) parsimony analysis indicates that subg. Anoplobatus is sister to clade A, and strongly supports Dalibarda repens as sister to R. lasiococcus. This suggests that Dalibarda repens be classified as R. dalibarda consistent with Linnaeus (1762) and Focke (1910). Parsimony analyses of GBSSI-1 sequences result in a large polytomy and do not recover clade A. The presence of three (GBSSI-la, GBSSI-1 (3 and GBSSI-ly) putative forms of the gene is observed. However, separate parsimony analysis of GBSSIly sequences demonstrates strong support for clade A and the monophyly of ubg. Anoplobatus. In this analysis, two different alleles are present in R. chamaemorus; one occurs outside clade A (sister to R. lasiococcus) and the other nests within clade A (sister to R. arcticus). Thus these data suggest that R. chamaemorus may be an ancient allopolyploid. The phylogenetic position of Dalibarda repens relative to Rubus cannot be resolved by existing GBSSI-1 data.

Thesis (Ph. D.)--Oregon State University, 2001. / Typescript (photocopy). Includes bibliographical references. Also available on the World Wide Web.