Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii:
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| Format: | Thesis Book |
| Language: | English |
| Published: |
2012
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| Subjects: | |
| Online Access: | Inhaltsverzeichnis |
| Physical Description: | 155 S. Ill., graph. Darst. |
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| adam_text | Titel: Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strai
Autor: Li, Jingtao
Jahr: 2012
Table of Contents
1. Introduction....................................................................................................................................1
1.1 Parasitism...............................................................................................................................1
1.2 Toxoplasma gondii..................................................................................................................2
1.3 Mouse strain CIM is resistant to virulent T.gondii strains......................................................S
1.4 Mouse IRG proteins and T. gondii virulence factors..............................................................6
1.5 Host - parasite co-evolution and MHC.................................................................................12
1.6 The aim of this study............................................................................................................18
2. Materials and Methods................................................................................................................19
2.1 Materials...............................................................................................................................19
2.1.1 Chemicals and Solutions...............................................................................................19
2.1.2 Enzymes and Proteins...................................................................................................19
2.1.3 Reagent Kits..................................................................................................................19
2.1.4 Vectors..........................................................................................................................20
2.1.5 Stock solutions and buffers..........................................................................................20
2.1.6 Media............................................................................................................................23
2.1.7 Bacterial strains............................................................................................................24
2.1.8 Primary and secondary immunoreagents....................................................................25
2.1.9 Mouse genomic DNA samples......................................................................................26
2.1.10 Toxoplasma gondii strains............................................................................................27
2.1.11 Mammalian cells...........................................................................................................27
2.1.12 Oligonucleotides...........................................................................................................27
2.1.13 BAC library screening probes........................................................................................29
2.1.14 Laboratory equipment..................................................................................................29
2.2 Molecular biology.................................................................................................................30
2.2.1 Mouse sample collection and genomic DNA isolation.................................................30
2.2.2 Amplification of IRG genes from genomic DNA............................................................30
2.2.3 Cloning and sequencing of IRG genes...........................................................................30
2.2.4 Screening and shotgun sequencing of CAST/EI BAC Library.........................................31
2.2.5 Establishing the IFN-y induced CIM cDNA library.........................................................32
2.2.6 Screening and sequencing IRG genes from cDNA library.............................................32
2.2.7 Analysis of IRG expression in cells................................................................................34
2.2.8 Cell Bilology of IRG members from CIM.......................................................................35
2.2.9 In vivo test of mice resistance against T.gondii infection.............................................36
2.3 Bioinformatics.......................................................................................................................37
2.3.1 Collecting IRG sequence from Database......................................................................37
2.3.2 Assembly of IRG coding units from MGP and NIG........................................................37
2.3.3 Assembly the MSM/Ms BAC clones..............................................................................37
2.3.4 Data analysis.................................................................................................................38
3. Results I The haplotype polymorphism of IRG genes in mice......................................................39
3.1 IRG genes in the inbred mouse strain CAST/EU....................................................................39
3.1.1 Contigs containing IRG genes sequenced from BAC library CHORI-26.........................39
3.1.2 IRG genes are located in a recombination hotspot......................................................41
3.1.3 IRG alleles in CAST/Ei genome......................................................................................45
3.2 IRG genes in mouse strain CIM.............................................................................................46
3.2.1 Establish of a high quality cDNA library........................................................................46
3.2.2 Screening the cDNA library for IRG genes....................................................................48
3.3 IRG genes in mouse strain MSM/Ms....................................................................................51
3.3.1 lllumina sequencing of MSM/Ms BAC clones...............................................................51
3.3.2 The arrangement of IRG genes in MSM/Ms genome...................................................52
3.4 IRG genes in Wild Mice.........................................................................................................54
3.4.1 Wild mouse samples from China..................................................................................54
3.4.2 IRG genes in wild mouse samples.................................................................................57
3.5 Haplotype polymorphism of IRG genes................................................................................57
3.5.1 IRG genes in mouse strains...........................................................................................57
3.5.2 IRG genes in wild mouse populations...........................................................................61
3.5.3 IRG proteins rival the MHC in diversity and complexity...............................................63
3.5.4 Evolution and selection on Irgb6 and Irgb2..................................................................67
3.6 Expression levels of IRGs differ among mouse strains.........................................................71
3.6.1 IRGB tandem genes are strongly expressed in wild derived mouse strains.................71
3.6.2 Expression level of other IRG proteins varies among mouse strains...........................72
3.6.3 Promoter regions of Irgb6 and Irgb2-bl in different mouse strains............................73
4. Results II Mouse IRG haplotype mediates resistance against virulent T.gondii...........................77
4.1 Mouse strain CIM resists infection of virulent T.gondii strains in vivo.................................77
4.2 Transfected lrgb6_CIM behaves like lrgb6_BL/6..................................................................78
4.3 Irgb2-bl serves as a resistance factor in CIM against virulent T.gondii...............................78
4.3.1 ROP5 dependent loading of lrgb2-bl_CIM on the PV of virulent T.gondii..................78
4.3.2 lrgb2-bl_CIM but not lrgb2-bl_BL/6 assists the loading of Irgb6 onto RH-YFP..........80
4.3.3 Transfected lrgb2-bl_CIM inhibits the phosphorylation of Irga6................................83
5. Discussion.....................................................................................................................................87
5.1 Irgb2-bl is a defense factor against virulent T.gondii strains..............................................87
5.2 IRG proteins work as a highly complex and integrated system...........................................91
5.3 IRG system and MHC family.................................................................................................96
6. Summary.....................................................................................................................................101
7. Zusammenfassung......................................................................................................................102
8. Appendix.....................................................................................................................................103
Appendix Table 1. Morphology of mouse samples collected in China..........................................103
Appendix Table 2. IRG clones in IFN-induced CIM cDNA library...................................................104
Appendix Table 3. Mass spectrometry analysis of 100 Kda protein in CAST/Ei.............................107
Appendix Table 4. Polymorphism of Irgml in mouse strains and wild mice................................Ill
Appendix Table 6. Polymorphism of Irga6 in mouse strains and wild mice..................................115
Appendix Table 8. Polymorphism of Irgb2 in mouse strains and wild mice..................................124
Appendix Table 9 50 Random genes from 7 mouse strains.........................................................131
Appendix Table 10 Repeat Masker results of BAC clone CHORI-26 76B7......................................134
Appendix Data 1 Sequencing of CIM and C57BL/6 transcriptome by lllumina Technology...........139
Appendix Fig.l The distribution of wild mouse samples used in this research.............................140
9. References..................................................................................................................................141
10. Acknowledgement..................................................................................................................152
Abbreviations......................................................................................................................................153
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| author | Li, Jingtao 1980- |
| author_GND | (DE-588)1030376123 |
| author_facet | Li, Jingtao 1980- |
| author_role | aut |
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| dewey-tens | 570 - Biology |
| discipline | Biologie |
| format | Thesis Book |
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| spelling | Li, Jingtao 1980- Verfasser (DE-588)1030376123 aut Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii vorgelegt von Jingtao Li 2012 155 S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Köln, Univ., Diss., 2012 (DE-588)4113937-9 Hochschulschrift gnd-content HBZ Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=025963377&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Li, Jingtao 1980- Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii |
| subject_GND | (DE-588)4113937-9 |
| title | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii |
| title_auth | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii |
| title_exact_search | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii |
| title_full | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii vorgelegt von Jingtao Li |
| title_fullStr | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii vorgelegt von Jingtao Li |
| title_full_unstemmed | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii vorgelegt von Jingtao Li |
| title_short | Haplotypic polymorphism of the IRG protein family mediates resistance of mice against virulent strains of Toxoplasma gondii |
| title_sort | haplotypic polymorphism of the irg protein family mediates resistance of mice against virulent strains of toxoplasma gondii |
| topic_facet | Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=025963377&sequence=000002&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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