Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L.:
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
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2011
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| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | VI, 100 Bl. Ill., graph. Darst. |
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| 245 | 1 | 0 | |a Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. |c vorgelegt von Ahmed Mustafa Mohammed Hassanein |
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Datensatz im Suchindex
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| adam_text | Titel: Micro-propagation and genetic manipulation of Artemisia annua L. expression analysis of different
Autor: Hassanein, Ahmed Mustafa Mohammed
Jahr: 2011
Index
I Introduction..................................................................................................................1
1.1 Artemisia annua L..................................................................................................................1
1.1.1 Artemisinin and its derivatives.......................................................................3
1.1.2 Sequestration of artemisinin...........................................................................6
1.1.3 Glandular trichomes and the glabrous genes..................................................7
1.1.3.1 Glandular trichome........................................................................................7
1.1.3.2 Glabra genes...................................................................................................7
1.1.4 Medicinal uses of artemisinin and its derivatives...........................................8
1.1.5 Mode of action of artemisinin as antimalarial and anti cancer.......................9
1.1.6 Bisynthesis of artemisinin............................................................................10
1.2 Nigel la saliva L....................................................................................................................12
1.2.1 Active principles (composition and phytochemicals)..................................14
1.2.2 Pharmacological Activities...........................................................................15
1.2.3 Triterpene biosynthesis.................................................................................16
1.2.4 Nigella sativa ß-amyrin synthase (NsßASl) gene........................................18
1.2.5 Prediction of a unique N-terminal domain of NsßASl................................18
1.2.6 mRNA splicing.............................................................................................19
1.3 Aims of the work.................................................................................................................20
77 Materials and methods...............................................................................................21
II. 1 Chemicals and consumables...............................................................................................21
11.1.1 Chemicals ....................................................................................................21
11.1.2 Equipments...................................................................................................22
11.1.3 Organisms.....................................................................................................24
11.1.4 Vectors ........................................................................................................24
11.1.5 Yeast strain...................................................................................................25
11.1.6 Plants ........................................................................................................25
11.1.7 Oligonucleotides...........................................................................................25
11.2 Media, solutions and buffers...............................................................................................27
II.2.1 Media ........................................................................................................27
11.2.1.1 Growth media for E. coli and Agrobacteria...................................27
11.2.1.2 MS-Medium...................................................................................27
II.2.1.3-6 Medium for callus induction..........................................................28
II.2.1.7-18 Medium for shoot induction...........................................................28
11.2.1.19 Medium for shoot elongation.........................................................29
11.2.1.20 Medium for root induction.............................................................29
11.2.1.21 Medium for root induction.............................................................29
11.2.1.22 Media for the regeneration of Artemisia........................................29
11.2.1.23 Media for Saccharomyces cerevisae..............................................30
H.2.2 Solutions and buffers ...................................................................................31
11.2.2.1 Solutions.........................................................................................31
11.2.2.2 Buffers............................................................................................32
11.3 Methods ......................................................................................................................32
11.3.1 Molecular biological methods for cloning....................................................32
11.3.1.1 Restriction.......................................................................................32
11.3.1.2 Ligation...........................................................................................32
11.3.1.3 Plasmid DNA isolation from E. coli...............................................32
11.3.1.4 Plasmid DNA isolation from E. coli according to..........................
1, 2 and 3 solutions method............................................................32
11.3.2 Quantification of DNA.................................................................................33
11.3.3 DNA Sequencing..........................................................................................33
Index
11.3.4 Polymerase chain reaction (PCR).................................................................33
11.3.5 Agrarose gel electrophoresis........................................................................34
11.3.6 Preparative agarose gel electrophoresis........................................................35
11.3.7 Transformation ofE. coli by electroporation...............................................35
11.3.8 Preparation of competent yeast.....................................................................35
11.3.9 Transformation of competent yeast by the Li-Acetate/PEG method............36
11.3.10 Preservation of yeast cells............................................................................36
11.3.11 Transformation of A. tumefaciens by electroporation..................................37
11.3.12 Growth of recombinant A. tumefaciens and preparations of glycerol
stock ........................................................................................................37
11.3.13 Agrobacteria-mediated leaf disc transformation of Artemisia ....................37
11.3.14 Tabak infiltration (N.benthamiana)..............................................................38
11.4 Isolation of genomic DNA from plants using CTAB protocol...........................................38
11.5 Isolation of RNA ........................................................................................................39
11.6 Reverse transcription- semi-quantitative PCR (RT-SQ-PCR)............................................39
11.7 Sterilization of the seeds.....................................................................................................40
11.8 Qualitative and quantitative analysis...................................................................................40
II. 8.1 Silylation.......................................................................................................40
II. 8.2 Gas chromatography and mass spectrophotometery....................................40
III Results........................................................................................................................42
ULLI Regeneration ........................................................................................................42
111.1.1.1 Seed sterilization and germination ..............................................................42
111.1.1.2 Callus induction............................................................................................42
111.1.1.3 Shoot induction.............................................................................................43
111.1.2 Regeneration of another cultivar of Artemisia annua L. of highly yielding strain
Artemis® F2 ........................................................................................................44
111.1.2.1 Callus and shoot induction...........................................................................44
111.1.2.2 Shoot elongation medium.............................................................................45
111.1.2.3 Root induction medium................................................................................45
111.1.3 Summary of regeneration protocols for A annua...........................................................46
111.1.4 Expression of Arabidopsis thaliana GL1 and GL3 genes in Artemisia annua................46
111.1.5 Agrobacterium tumefaciens- mediated leaf expiants transformation..............................49
111.1.6 Expression of GL 1 m A.annua (Art 04X)........................................................................50
111.1.7 Transformation using vacuum infiltration method..........................................................50
111.1.8 Agrobacterium tumefaciens-mediated leaf disc transformation using Hygromycin
Cassette ........................................................................................................51
III.II Analysis of the expression of different types of NsßASl in Nigella sativa flower..............
followed by the analysis in different tissues..................................................................52
111.11.1 Localization study of NsßASl/cwg...............................................................55
111.11.2 Functional characterization of NsßASl long.................................................57
111.11.3 NsßASl/o«g expression analysis of related species ofNigella sativa.........58
IV Discussion..................................................................................................................63
IV.1 Regeneration ........................................................................................................63
IV.1.1 Transformation ............................................................................................64
IV.2 Alternative transcription initiation of NsßASl..................................................................64
IV.3 Localization study of NsßASl long...................................................................................65
IV.4 Tissue specific expression of NsßASl/o«g........................................................................68
IV.5 Functional characterization of NsßASl long......................................................................68
IV.6 NsßASl/ong expression analysis of related species ofNigella sativa..............................69
II
Index
V Outlook.......................................................................................................................70
VI Summary....................................................................................................................71
VII Appendix.....................................................................................................................73
VIII References..................................................................................................................77
III
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| language | English |
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| spelling | Hassanein, Ahmed Mustafa Mohammed 1974- Verfasser (DE-588)1011277565 aut Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. vorgelegt von Ahmed Mustafa Mohammed Hassanein 2011 VI, 100 Bl. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Münster (Westfalen), Univ., Diss., 2011 Einjähriger Beifuß (DE-588)4682039-5 gnd rswk-swf Transkription Genetik (DE-588)4185906-6 gnd rswk-swf Schwarzkümmel (DE-588)4278192-9 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Einjähriger Beifuß (DE-588)4682039-5 s Schwarzkümmel (DE-588)4278192-9 s Transkription Genetik (DE-588)4185906-6 s DE-604 HBZ Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=022553338&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Hassanein, Ahmed Mustafa Mohammed 1974- Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. Einjähriger Beifuß (DE-588)4682039-5 gnd Transkription Genetik (DE-588)4185906-6 gnd Schwarzkümmel (DE-588)4278192-9 gnd |
| subject_GND | (DE-588)4682039-5 (DE-588)4185906-6 (DE-588)4278192-9 (DE-588)4113937-9 |
| title | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. |
| title_auth | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. |
| title_exact_search | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. |
| title_full | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. vorgelegt von Ahmed Mustafa Mohammed Hassanein |
| title_fullStr | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. vorgelegt von Ahmed Mustafa Mohammed Hassanein |
| title_full_unstemmed | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. vorgelegt von Ahmed Mustafa Mohammed Hassanein |
| title_short | Micro-propagation and genetic manipulation of Artemisia annua L. & expression analysis of different NsbetaAS1 gene transcripts in Nigella sativa L. |
| title_sort | micro propagation and genetic manipulation of artemisia annua l expression analysis of different nsbetaas1 gene transcripts in nigella sativa l |
| topic | Einjähriger Beifuß (DE-588)4682039-5 gnd Transkription Genetik (DE-588)4185906-6 gnd Schwarzkümmel (DE-588)4278192-9 gnd |
| topic_facet | Einjähriger Beifuß Transkription Genetik Schwarzkümmel Hochschulschrift |
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