Molecules and mechanisms involved in the nematode oxidative stress response:
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | German |
| Veröffentlicht: |
1997
|
| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | 121 S. Ill., graph. Darst. |
Internformat
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| 100 | 1 | |a Tawe, Wilson N. |d 1967- |e Verfasser |0 (DE-588)120068834 |4 aut | |
| 245 | 1 | 0 | |a Molecules and mechanisms involved in the nematode oxidative stress response |c by Wilson N. Tawe |
| 264 | 1 | |c 1997 | |
| 300 | |a 121 S. |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 502 | |a Hamburg, Univ., Diss. | ||
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| 655 | 7 | |0 (DE-588)4113937-9 |a Hochschulschrift |2 gnd-content | |
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Datensatz im Suchindex
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T
ABLE
OF
CONTENTS
TABLE
OF
CONTENTS
1.
INTRODUCTION
1
1.1
NEMATODES
1
1.2
FILARIASIS
1
1.3
ONCHOCERCA
VOLVULUS
2
1.3.1
CLASSIFICATION
2
1.3.2
LIFE
CYCLE
2
1.3.3
GENERAL
MORPHOLOGY
3
1.4
THE
DISEASE
ONCHOCERCIASIS
4
1.4.1
PREVALENCE
AND
GEOGRAPHIC
DISTRIBUTION
4
1.4.2
VECTOR
AND
TRANSMISSION
4
1.4.3
PATHOGENICITY
AND
SYMPTOMATOLOGY
4
1.4.4
DIAGNOSIS
AND
TREATMENT
4
1.5
CAENORHABDITIS
ELEGANS
5
1.5.1
CLASSIFICATION
5
1.5.2
LIFE
CYCLE
6
1.5.3
C.
ELEGANS
AS
AN
EXPERIMENTAL
MODEL
SYSTEM
7
1.6
OXIDATIVE
STRESS
7
1.6.1
THE
SUPEROXIDE
ANION
RADICAL
7
1.6.2
GENERATORS
OF
TOXIC
OXYGEN
SPECIES
9
1.6.3
OXIDATIVE
DAMAGE
IN
BIOLOGICAL
SYSTEMS
9
1.7
CELLULAR
ANTIOXIDANT
SYSTEMS
10
1.7.1
SCAVENGING
SYSTEMS
FOR
OXYGEN
RADICALS
AND
THEIR
DERIVATIVES;
THE
HOST
-
PARASITE
RELATIONSHIP
10
1.8
SUPEROXIDE
DISMUTASE
(SOD)
12
1.8.1
CLASSIFICATION,
OCCURRENCE
AND
DISTRIBUTION
12
1.8.2
SOD
AND
0.
VOLVULUS
14
1.9
AIMS
AND
OBJECTIVES
15
T
ABLE
OF
CONTENTS
2.
MATERIALS
AND
METHODS
16
2.1
CHEMICALS
AND
BIOREAGENTS
16
2.1.1
DNA
AND
RNA
MODIFYING
ENZYMES
16
2.1.2
IMMUNOREAGENTS
16
2.1.3
ELECTROPHORESIS
REAGENTS
16
2.1.4
STAINING
REAGENTS
16
2.1.5
POLYACRYLAMIDE
GEL
MARKER
PROTEINS
17
2.1.6
MEDIA
AND
PLATES
17
2.1.7
KITS
17
2.1.8
RADIOACTIVE
REAGENTS
17
2.2
NEMATODE
MATERIAL
17
2.2.1
ONCHOCERCA
VOLVULUS
17
2.2.2
CAENORHABDITIS
ELEGANS
18
2.3
VECTORS,
BACTERIA
AND
EUKARYOTIC
CELLS
18
2.3.1
BACTERIOPHAGE VECTORS
18
2.3.2
PLASMID
VECTORS
18
2.3.3
BACTERIAL
STRAINS
19
2.3.4
EUKARYOTIC
CELLS
19
2.4
DNA
PREPARATION
19
2.4.1
MINI-PREPARATION
OF
PLASMID
DNA
19
2.4.2
LARGE-SCALE
PLASMID
DNA
PREPARATION
19
2.4.3
GENOMIC
DNA
PREPARATION
19
2.4.4
ETHANOL
PRECIPITATION
20
2.4.5
PHENOL-CHLOROFORM
EXTRACTION
20
2.5
RNA
PREPARATION
20
2.5.1
PURIFICATION
OF
RNA
20
2.5.2
ISOLATION
OF
POLY(A)
+
RNA
21
2.6
RESTRICTION
ENZYME
DIGESTION
OF
DNA
1
21
2.7
AGAROSE
GEL
ELECTROPHORESIS
OF
DNA
21
2.8
OLIGONUCLEOTIDE
SYNTHESIS
22
2.9
POLYMERASE
CHAIN
REACTION
(PCR)
22
2.10
RECOVERY
OF
DNA
FROM
AGAROSE
GELS
22
T
ABLE
OF
CONTENTS
2.11
LIGATION
OF
DNA
23
2.11.1
SUB-CLONING
IN
PLASMID
VECTORS
23
2.11.2
SUB-CLONING
OF
PCR
FRAGMENTS
IN
THE
PCRYY
II
(TA-CLONING)
VECTOR
23
2.11.3
SUB-CLONING
OFPCR
FRAGMENTS
INTO
SMAL-CUT
PUC18
23
2.12
TRANSFORMATION
24
2.12.1
COMPETENT
BACTERIA
(DH5A,
BL21)
24
2.12.2
TRANSFORMATION
OF
COMPETENT
CELLS
24
2.13
RADIOACTIVE
LABELING
OF
DNA
24
2.14
SOUTHERN
BLOTTING
25
2.15
DNA-DNA
HYBRIDIZATION
25
2.16
GENOMIC
AND
CDNA
LIBRARIES
26
2.17
SCREENING
OF
GENOMIC
AND
CDNA
LIBRARIES
26
2.17.1
PREPARATION
OF
PLATING
BACTERIA
26
2.17.2
SCREENING
OF
THE
X
ZAP
II
BACTERIOPHAGE
CDNA
LIBRARY
26
2.17.3
IN
VIVO
EXCISION
OF
PLASMID
VECTOR
FROM
BACTERIOPHAGE
X
ZAP
II
ARMS
27
2.17.4
SCREENING
OF
THE
X
FIX
II
GENOMIC
LIBRARY
27
2.17.5
BACTERIOPHAGE
LYSIS
27
2.17.6
BACTERIOPHAGE
DNA
PREPARATION
27
2.18
DNA
SEQUENCING
AND
ANALYSIS
27
2.19
5'
RACE
28
2.20
ISOLATION
OF
THE
0.
VOLVULUS
MNSOD
CDNA
28
2.21
EXPRESSION
OF
THE
0.
VOLVULUS
MNSOD
29
2.21.1
SUBCLONING
OF
THE
OVMNSOD
IN
PLASMID
VECTORS
29
2.21.2
EXPRESSION
OF
RECOMBINANT
OVMNSOD
IN
E.
COLI
29
2.21.3
EXPRESSION
IN
BACULOVIRUS
30
2.21.3.1
INSECT
CELL
CULTURE
30
2.21.3.2
TRANSFECTION
OF
INSECT
CELLS
30
2.21.3.3
INFECTION
OF
SFL
CELLS
WITH
RECOMBINANT
VIRUS
31
2.22
PREPARATION
OF
CELL
EXTRACTS
31
2.23
PROTEIN
GEL
ELECTROPHORESIS
31
2.24
PROTEIN
STAINING
IN
GELS
32
2.24.1
COOMASSIE
BLUE
STAINING
32
2.24.2
SILVER
STAINING
32
T
ABLE
OF
CONTENTS
2.25
WESTERN
BLOTTING
33
2.26
IMMUNODETECTION
33
2.27
PURIFICATION
OF
RECOMBINANT
ENZYME
34
2.27.1
NICKEL
CHELATING
CHROMATOGRAPHY
(HIS-TAGGED
PROTEINS)
34
2.27.2
PREPCELL
(LARGE
SCALE
PURIFICATION
OF
RECOMBINANT
PROTEIN)
34
2.28
SOD
ACTIVITY
ASSAY
35
2.29
PROTEIN
DETERMINATION
35
2.30
PROMOTER
STUDIES
35
2.30.1
IN
VITRO
TRANSCRIPTION
35
2.30.1.1
PRIMER
LABELING
36
2.30.1.2
PRIMER EXTENSION
36
2.30.2
CHO
CELL
CULTURE
36
2.30.3
CLONING
INTO
THE
PGL3
VECTOR
36
2.30.4
TRANSFECTION
37
2.30.5
REPORTER
GENE
ANALYSIS
37
2.30.6
EMSA
(ELECTROPHORETIC
MOBILITY
SHIFT
ASSAY)
38
2.31
IN
VITRO
STRESS
EXPERIMENTS
39
2.31.1
ISOLATION
OF
POLYMORPHONUCLEAR
CELLS
(PMNCS)
39
2.31.2
O.
VOLVULUS
INDUCTION
39
2.31.3
C.
ELEGANS
INDUCTION
39
2.31.4
MRNA
DIFFERENTIAL
DISPLAY
40
2.31.5
NORTHERN
BLOTTING
41
3.
RESULTS
42
3.1
CHARACTERIZATION
OF
THE
OVMNSOD
CDNA
AND
RECOMBINANT
PROTEIN
42
3.1.1
ISOLATION
AND
SEQUENCE
OF
THE
CMVINSOD
CDNA
42
3.1.2
ANALYSIS
OF
THE
OVMNSOD
DEDUCED
AMINO
ACID
SEQUENCE
44
3.1.3
EXPRESSION
OF
RECOMBINANT
OVMNSOD
IN
PJC20,
PJC40,
PGEX-3X
ANDPBSV.8HIS
46
3.1.4
PROTEIN
REFOLDING
AND
SOD
ACTIVITY
ASSAY
50
3.2
CHARACTERIZATION
OF
THE
OVMNSOD
GENE
51
3.2.1
ISOLATION
AND
SEQUENCE
OF
THE
OVMNSOD
GENE
51
3.2.2
ANALYSIS
OF
THE
OVMNSOD
GENOMIC
CLONES
51
T
ABLE
OF
CONTENTS
3.2.3
THE
OVMNSOD
GENE
AND
DEDUCED
AMINO
ACID
SEQUENCE
53
3.3
CHARACTERIZATION
OF
THE
OVCUZNSOD
GENES
56
3.3.1
ISOLATION
OF
THE
OVCUZNSOD
GENES
56
3.3.2
ANALYSIS
OF
THE
OVCUZNSOD
GENES
56
3.3.3
THE
OVCUZNSOD
GENES:
NUCLEOTIDE
SEQUENCES
AND
ORGANIZATION
58
3.4
ANALYSIS
OF
THE
OVSOD
GENE
PROMOTERS
60
3.4.1
IN
VITRO
TRANSCRIPTION
TO
DETERMINE
TRANSCRIPTION
START
POINTS
(TSP)
60
3.4.2
REPORTER
GENE
ASSAY
-
LUCIFERASE
CONSTRUCTS
AND
ASSAYS
62
3.4.3
EMSA
-
ANALYSIS
OF
THE
SP-1
AND
AP-1
BINDING
TO
THE
OVSOD
PROMOTER
REGIONS
64
3.5
INDUCTION
STUDIES
66
3.5.1
C.
ELEGANS'.
INDUCTION
WITH
PARAQUAT
66
3.5.1.1
SUPEROXIDE
DISMUTASE
MRNA
LEVELS
IN
RESPONSE
TO
PARAQUAT
66
3.5.1.2
ISOLATION
OF
PARAQUAT-RESPONSIVE
GENES
BY
MRNA
DIFFERENTIAL
DISPLAY
67
3.5.1.3
NORTHERN
BLOT
ANALYSIS
USING
DIFFERENTIALLY
DISPLAYED
CDNA
FRAGMENTS
70
3.5.1.4
COMPARISONS
WITH
THE
C.
ELEGANS
DATABASE
72
3.5.1.5
ISOLATION
OF
COMPLETE
CDNAS
72
3.5.2
O.
VOLVULUS'.
INDUCTION
WITH
PMA-ACTIVATED
PMNCS
75
4.
DISCUSSION
78
4.1
ISOLATION
AND
SEQUENCE
ANALYSIS
OF
THE
OVMNSOD
CDNA
78
4.1.1
THE
OVMNSOD
CDNA
CONTAINS
A
5'
TRANS-SPLICED
LEADER
SEQUENCE
78
4.1.2
THE
OVMNSOD
CONTAINS
TYPICAL
FEATURES
OF
A
MITOCHONDRIAL
PROTEIN
79
4.1.3
THE
OVMNSOD
IS
QUITE
SIMILAR
TO
OTHER
MNSODS
80
4.1.4
RECOMBINANT
OVMNSOD
IS
A
HIGHLY
INSOLUBLE
AND
INACTIVE
ENZYME
81
4.1.5
RABBIT
ANTISERUM
RAISED
AGAINST
PURIFIED
ROVMNSOD
RECOGNIZES
NATIVE
O.
VOLVULUS
MNSOD
IN
WORM
EXTRACTS
83
4.2
ANALYSIS
OF
THE
OVSOD
1,2
AND
3
GENES
83
4.2.1
THE
O.
VOLVULUS
GENOME
ENCODES
THREE
DISTINCT
SODS
83
4.2.2
IN
VITRO
TRANSCRIPTION
IDENTIFIES
THE
TRANSCRIPTION
INITIATION
SITES
OF
SL-ACCEPTING
GENES
86
4.2.3
THE
OVSOD
PROMOTERS
ARE
CAPABLE
OF
DRIVING
TRANSCRIPTION
OF
THE
LUCIFERASE
REPORTER
GENE
IN
CHO
CELLS
87
T
ABLE
OF
CONTENTS
4.2.4
ANALYSIS
OF
THE
OVSOD
GENE
PROMOTER
SEQUENCES
89
4.2.5
GEL
MOBILITY
SHIFT
ASSAYS
IDENTIFY
SPECIFIC
TRANSCRIPTION
FACTOR
BINDING
SITES
IN
OVSOD
GENE
SEQUENCES
91
4.3
PARAQUAT-INDUCED
GENE
EXPRESSION
IN
C.
ELEGANS:
IDENTIFICATION
BY
MRNA
DIFFERENTIAL
DISPLAY
92
4.3.1
INDUCTION
WITH
PARAQUAT
92
4.3.2
MRNA
DIFFERENTIAL
DISPLAY
93
4.3.3
PARAQUAT
INDUCES
SUPEROXIDE
DISMUTASES
IN
C.
ELEGANS
95
4.3.4
PARAQUAT
INDUCES
A
GLUTATHIONE
S-TRANSFERASE
IN
C,
ELEGANS
95
4.3.5
PARAQUAT
INDUCES
A
C.
ELEGANS
ZINC
FINGER/LEUCINE
ZIPPER
TRANSCRIPTION
FACTOR
96
4.4
INDUCTION
IN
O.
VOLVULUS
97
5.
SUMMARY
98
6.
REFERENCES
102 |
| any_adam_object | 1 |
| author | Tawe, Wilson N. 1967- |
| author_GND | (DE-588)120068834 |
| author_facet | Tawe, Wilson N. 1967- |
| author_role | aut |
| author_sort | Tawe, Wilson N. 1967- |
| author_variant | w n t wn wnt |
| building | Verbundindex |
| bvnumber | BV012051601 |
| classification_tum | CHE 832d BIO 816d BIO 750d BIO 780d |
| ctrlnum | (OCoLC)51757959 (DE-599)BVBBV012051601 |
| discipline | Biologie Chemie |
| format | Thesis Book |
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| genre | (DE-588)4113937-9 Hochschulschrift gnd-content |
| genre_facet | Hochschulschrift |
| id | DE-604.BV012051601 |
| illustrated | Illustrated |
| indexdate | 2024-08-14T00:20:03Z |
| institution | BVB |
| language | German |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-008157098 |
| oclc_num | 51757959 |
| open_access_boolean | |
| owner | DE-29T DE-M49 DE-BY-TUM DE-12 DE-355 DE-BY-UBR DE-11 DE-188 |
| owner_facet | DE-29T DE-M49 DE-BY-TUM DE-12 DE-355 DE-BY-UBR DE-11 DE-188 |
| physical | 121 S. Ill., graph. Darst. |
| publishDate | 1997 |
| publishDateSearch | 1997 |
| publishDateSort | 1997 |
| record_format | marc |
| spelling | Tawe, Wilson N. 1967- Verfasser (DE-588)120068834 aut Molecules and mechanisms involved in the nematode oxidative stress response by Wilson N. Tawe 1997 121 S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Hamburg, Univ., Diss. Stressreaktion (DE-588)4138565-2 gnd rswk-swf Superoxiddismutasen (DE-588)4184118-9 gnd rswk-swf Gen (DE-588)4128987-0 gnd rswk-swf Genklonierung (DE-588)4123275-6 gnd rswk-swf Oxidativer Stress (DE-588)4353497-1 gnd rswk-swf Onchocerca volvulus (DE-588)4210380-0 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Onchocerca volvulus (DE-588)4210380-0 s Oxidativer Stress (DE-588)4353497-1 s Stressreaktion (DE-588)4138565-2 s Superoxiddismutasen (DE-588)4184118-9 s Genklonierung (DE-588)4123275-6 s DE-604 Gen (DE-588)4128987-0 s DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008157098&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Tawe, Wilson N. 1967- Molecules and mechanisms involved in the nematode oxidative stress response Stressreaktion (DE-588)4138565-2 gnd Superoxiddismutasen (DE-588)4184118-9 gnd Gen (DE-588)4128987-0 gnd Genklonierung (DE-588)4123275-6 gnd Oxidativer Stress (DE-588)4353497-1 gnd Onchocerca volvulus (DE-588)4210380-0 gnd |
| subject_GND | (DE-588)4138565-2 (DE-588)4184118-9 (DE-588)4128987-0 (DE-588)4123275-6 (DE-588)4353497-1 (DE-588)4210380-0 (DE-588)4113937-9 |
| title | Molecules and mechanisms involved in the nematode oxidative stress response |
| title_auth | Molecules and mechanisms involved in the nematode oxidative stress response |
| title_exact_search | Molecules and mechanisms involved in the nematode oxidative stress response |
| title_full | Molecules and mechanisms involved in the nematode oxidative stress response by Wilson N. Tawe |
| title_fullStr | Molecules and mechanisms involved in the nematode oxidative stress response by Wilson N. Tawe |
| title_full_unstemmed | Molecules and mechanisms involved in the nematode oxidative stress response by Wilson N. Tawe |
| title_short | Molecules and mechanisms involved in the nematode oxidative stress response |
| title_sort | molecules and mechanisms involved in the nematode oxidative stress response |
| topic | Stressreaktion (DE-588)4138565-2 gnd Superoxiddismutasen (DE-588)4184118-9 gnd Gen (DE-588)4128987-0 gnd Genklonierung (DE-588)4123275-6 gnd Oxidativer Stress (DE-588)4353497-1 gnd Onchocerca volvulus (DE-588)4210380-0 gnd |
| topic_facet | Stressreaktion Superoxiddismutasen Gen Genklonierung Oxidativer Stress Onchocerca volvulus Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=008157098&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT tawewilsonn moleculesandmechanismsinvolvedinthenematodeoxidativestressresponse |