Mak Wai Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (leaves 95-115). / Abstracts in English and Chinese. / Declaration --- p.i / Abstract --- p.ii / Acknowledgements --- p.viii / Table of Contents --- p.ix / List of Tables --- p.xiii / List of Figures --- p.xiv / List of Abbreviations --- p.xvi / Chapter Chapter 1 --- Introduction and Literature Review / Chapter 1.1 --- Introduction --- p.1 / Chapter 1.2 --- Endocrine control of reproduction in crustaceans --- p.2 / Chapter 1.3 --- The X-organ sinus gland (XOSG) complex --- p.3 / Chapter 1.3.1 --- CHH/MIH/GIH family of neuropeptides --- p.4 / Chapter 1.3.2 --- Gonad inhibiting hormone (GIH) --- p.6 / Chapter 1.3.3 --- Molt inhibiting hormone (MIH) --- p.7 / Chapter 1.3.4 --- Crustaceans hyperglycemic hormone (CHH) --- p.9 / Chapter 1.3.5 --- The chormatophorotropins (RPCH and PDH neuropeptide family) --- p.10 / Chapter 1.4 --- Other mechanisms of reproduction control --- p.11 / Chapter 1.4.1 --- Methyl farnesoate (MF) and mandibular organ inhibiting hormone (MOIH) --- p.11 / Chapter 1.4.2 --- Gonad stimulating hormone (GSH) --- p.13 / Chapter 1.4.3 --- Serotonin (5HT) --- p.13 / Chapter 1.4.4 --- Dopamine --- p.14 / Chapter 1.4.5 --- Enkephalin (ENK) --- p.15 / Chapter 1.4.6 --- 17β-Estradiol --- p.16 / Chapter 1.4.7 --- Progesterone --- p.17 / Chapter 1.5 --- Androgenic hormone (AH) --- p.18 / Chapter 1.6 --- Objective and methodology of present research --- p.19 / Chapter 1.7 --- Reproductive biology of the shrimp Metapenaeus ensis --- p.20 / Chapter Chapter 2 --- Materials and Methods / Chapter 2.1 --- Introduction --- p.26 / Chapter 2.2 --- Extraction of eyestalk RNA from the shrimp --- p.27 / Chapter 2.3 --- Construction of the eyestalk cDNA library --- p.28 / Chapter 2.3.1 --- First-strand cDNA synthesis --- p.28 / Chapter 2.3.2 --- cDNA amplification by LD PCR --- p.29 / Chapter 2.3.3 --- Proteinase K digestion --- p.29 / Chapter 2.3.4 --- Sfi I digestion --- p.30 / Chapter 2.3.5 --- cDNA size fractionation --- p.30 / Chapter 2.3.6 --- Ligation of cDNA to λ TriplEx2 Vector --- p.31 / Chapter 2.3.7 --- Packaging --- p.31 / Chapter 2.3.8 --- Titering the unamplified library --- p.32 / Chapter 2.3.9 --- Library amplification --- p.33 / Chapter 2.3.10 --- Titering the amplified library --- p.34 / Chapter 2.4 --- Mass Excision of the eyestalk cDNA library --- p.34 / Chapter 2.5 --- PCR screening of inserted sequence --- p.35 / Chapter 2.6 --- RNA arbitrarily primed polymerase chain reaction (RAP-PCR) --- p.35 / Chapter 2.7 --- Dot blot hybridization --- p.37 / Chapter 2.7.1 --- Probe Labelling --- p.37 / Chapter 2.7.2 --- Dotting of membrane --- p.38 / Chapter 2.7.3 --- Hybridization --- p.38 / Chapter 2.7.4 --- Washing and chemiluminescent detection --- p.39 / Chapter 2.7.5 --- Probe stripping for re-hybridization --- p.40 / Chapter 2.8 --- Sequencing --- p.40 / Chapter 2.9 --- BLAST search --- p.41 / Chapter 2.10 --- Northern blot analysis --- p.41 / Chapter 2.10.1 --- Probe labelling --- p.41 / Chapter 2.10.2 --- RNA formaldehyde denaturing gel electrophoresis --- p.41 / Chapter 2.10.3 --- Northern blot --- p.42 / Chapter 2.10.4 --- Pre-hybridization --- p.43 / Chapter 2.10.5 --- Hybridization --- p.43 / Chapter 2.10.6 --- Washing and chemiluminescent detection --- p.43 / Chapter 2.11 --- Real-time RT-PCR --- p.44 / Chapter 2.11.1 --- DNaseI treatment --- p.44 / Chapter 2.11.2 --- First strand synthesis --- p.44 / Chapter 2.11.3 --- Primer design and verification --- p.45 / Chapter 2.11.4 --- Real-time PCR --- p.45 / Chapter 2.11.5 --- Statistical analysis --- p.45 / Chapter Chapter 3 --- Results / Chapter 3.1 --- Experimental animals --- p.50 / Chapter 3.2 --- Total RNA extraction --- p.50 / Chapter 3.3 --- Library construction --- p.50 / Chapter 3.4 --- PCR screening of inserted sequences --- p.50 / Chapter 3.5 --- RNA arbitrarily primed polymerase chain reaction (RAP-PCR) --- p.51 / Chapter 3.6 --- Dot blot hybridization --- p.51 / Chapter 3.7 --- Sequencing and BLAST search --- p.52 / Chapter 3.8 --- Northern blot analysis --- p.53 / Chapter 3.8.1 --- Housekeeping gene - elongation factor la --- p.53 / Chapter 3.8.2 --- Insulin-like growth factor binding protein (IGFBP) --- p.54 / Chapter 3.8.3 --- Arrestin --- p.54 / Chapter 3.8.4 --- Opsin --- p.54 / Chapter 3.9 --- Real-time RT-PCR --- p.55 / Chapter 3.9.1 --- β-Actin --- p.55 / Chapter 3.9.2 --- Farnesoic acid O-methyltranferase (FAMeT) --- p.56 / Chapter Chapter 4 --- Discussion / Chapter 4.1 --- Application of RAP-PCR and dot blot analysis in identifying differential expressed genes --- p.79 / Chapter 4.1.1 --- Abundant genes --- p.79 / Chapter 4.1.2 --- Appearance of 16S rRNA in eyestalk cDNA library of M. ensis --- p.80 / Chapter 4.1.3 --- False positive --- p.80 / Chapter 4.2 --- Investigation on common housekeeping genes --- p.81 / Chapter 4.3 --- Potential functions of identified differential expressed genes in reproduction of shrimp --- p.82 / Chapter 4.3.1 --- Arrestin --- p.84 / Chapter 4.3.2 --- Opsin --- p.86 / Chapter 4.3.3 --- Insulin-like growth factor binding protein (IGFBP) --- p.88 / Chapter 4.3.4 --- β-Actin --- p.89 / Chapter 4.4 --- Investigation for farnesoic acid O-methyltransferase (FAMeT) in shrimp --- p.89 / Chapter Chapter 5 --- General Conclusion --- p.91 / References --- p.95 / Appendix --- p.116
| Identifer | oai:union.ndltd.org:cuhk.edu.hk/oai:cuhk-dr:cuhk_325485 |
| Date | January 2006 |
| Contributors | Mak, Wai Yan., Chinese University of Hong Kong Graduate School. Division of Molecular Biotechnology. |
| Source Sets | The Chinese University of Hong Kong |
| Language | English, Chinese |
| Detected Language | English |
| Type | Text, bibliography |
| Format | print, xvi, 126 leaves : ill. ; 30 cm. |
| Rights | Use of this resource is governed by the terms and conditions of the Creative Commons “Attribution-NonCommercial-NoDerivatives 4.0 International” License (http://creativecommons.org/licenses/by-nc-nd/4.0/) |
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