표제지
목차
Abstract 13
Ⅰ. 서론 14
Ⅱ. 재료 및 방법 17
1. Next Generation Sequencing (NGS) 분석 17
2. 분자적 특성 17
3. 유전자 발현 분석 18
1) 정상 어체에서의 조직별 발현분석 18
2) 병원체 인위감염 후 시간별 발현분석 21
4. Septin2, 3, 8 재조합단백질 제작 및 phagocytosis 측정 22
1) Septin 2, 3, 8 재조합단백질 제작 22
2) Septin 2, 3, 8 재조합단백질의 phagocytosis 측정 23
Ⅲ. 결과 25
1. 강도다리 Septin Gene family의 분자적 특성 25
1) Septin 2의 분자적 특성 25
2) Septin 3의 분자적 특성 29
3) Septin 5의 분자적 특성 33
4) Septin 8의 분자적 특성 37
5) Septin 9의 분자적 특성 41
6) Septin 10의 분자적 특성 45
2. 강도다리 Septin gene family의 mRNA 발현 특성 53
1) Septin 2 53
2) Septin 3 55
3) Septin 5 57
4) Septin 8 59
5) Septin 9 61
6) Septin 10 63
3. Septin 2, 3, 8 재조합단백질의 phagocytosis 측정 65
1) Septin 2, 3, 8 재조합단백질 제작 65
2) Septin 2, 3, 8 재조합댄박질 phagocytosis 측정 67
Ⅳ. 고찰 71
Ⅴ. 요약 76
Ⅵ. 참고문헌 77
Table 1. List of primer sets used for RT-qPCR 20
Fig. 1. Nucleotide and deduced amino acid sequences of septin 2 from starry flounder. The black boxes indicate a GTPases domain and coiled-coil domain, respectively. 26
Fig. 2. Multiple sequence alignment analysis of the septin 2 based on the deduced amino acid sequences of starry flounder with other species. Black boxes: preserved motif. 27
Fig. 3. A phylogenetic tree of GTPases domain in septin 2 and other known homologues based on the neighbour-joining (NJ) method. The numbers are... 28
Fig. 4. Nucleotide and deduced amino acid sequences of septin 3 from starry flounder. The black box indicates a GTPases domain, respectively. 30
Fig. 5. Multiple sequence alignment analysis of the septin 3 based on the deduced amino acid sequences of starry flounder with other species. Black boxes: preserved motif. 31
Fig. 6. A phylogenetic tree of GTPases domain in septin 3 and other known homologues based on the neighbour-joining (NJ) method. The numbers are... 32
Fig. 7. Nucleotide and deduced amino acid sequences of septin 5 from starry flounder. The black boxes indicate a GTPases domain and coiled-coil domain, respectively. 34
Fig. 8. Multiple sequence alignment analysis of the septin 5 based on the deduced amino acid sequences of starry flounder with other species. Black boxes: preserved motif. 35
Fig. 9. A phylogenetic tree of GTPases domain in septin 5 and other known homologues based on the neighbour-joining (NJ) method. The numbers are... 36
Fig. 10. Nucleotide and deduced amino acid sequences of septin 8 from starry flounder. The black boxes indicate a GTPases domain and coiled-coil domain, respectively. 38
Fig. 11. Multiple sequence alignment analysis of the septin 8 based on the deduced amino acid sequences of starry flounder with other species. Black boxes: preserved motif. 39
Fig. 12. A phylogenetic tree of GTPases domain in septin 8 and other known homologues based on the neighbour-joining (NJ) method. The... 40
Fig. 13. Nucleotide and deduced amino acid sequences of septin 9 from starry flounder. The black box indicates a GTPases domain, respectively. 42
Fig. 14. Multiple sequence alignment analysis of the septin 9 based on the deduced amino acid sequences of starry flounder with other species. Black boxes: preserved motif. 43
Fig. 15. A phylogenetic tree of GTPases domain in septin 9 and other known homologues based on the neighbour-joining (NJ) method. The... 44
Fig. 16. Nucleotide and deduced amino acid sequences of septin 10 from starry flounder. The black boxes indicate a GTPases domain and coiled-coil domain, respectively. 46
Fig. 17. Multiple sequence alignment analysis of the septin 10 based on the deduced amino acid sequences of starry flounder with other species. Black boxes: preserved motif. 47
Fig. 18. A phylogenetic tree of GTPases domain in septin 10 and other known homologues based on the neighbour-joining (NJ) method. The... 48
Fig. 19. A phylogenetic tree of GTPases domain in septin 2 subgroup and other known homologues based on the neighbour-joining (NJ) method. The... 49
Fig. 20. A phylogenetic tree of GTPases domain in septin 3 subgroup and other known homologues based on the neighbour-joining (NJ) method. The... 50
Fig. 21. A phylogenetic tree of GTPases domain in septin 6 subgroup and other known homologues based on the neighbour-joining (NJ) method. The... 51
Fig. 22. Overall phylogenetic analysis of 6 septin genes using sequences from Starry flounder, Atlantic halibut, Barramundi, Olive flounder, Large... 52
Fig. 23. Expression level of septin 2 mRNA in various tissues of healthy state starry flounder. The EF-1α gene was used to normalise the... 53
Fig. 24. Expression level of septin 2 mRNA in the kidney, spleen, gill, liver, brain, intestine and heart of starry flounder after infection with (A): viral... 54
Fig. 25. Expression level of septin 3 mRNA in various tissues of healthy state starry flounder. The EF-1α gene was used to normalise the... 55
Fig. 26. Expression level of septin 3 mRNA in the kidney, spleen, gill, liver, brain, intestine and heart of starry flounder after infection with (A): viral... 56
Fig. 27. Expression level of septin 5 mRNA in various tissues of healthy state starry flounder. The EF-1α gene was used to normalise the... 57
Fig. 28. Expression level of septin 5 mRNA in the kidney, spleen, gill, liver, brain, intestine and heart of starry flounder after infection with (A): viral... 58
Fig. 29. Expression level of septin 8 mRNA in various tissues of healthy state starry flounder. The EF-1α gene was used to normalise the... 59
Fig. 30. Expression level of septin 8 mRNA in the kidney, spleen, gill, liver, brain, intestine and heart of starry flounder after infection with (A): viral... 60
Fig. 31. Expression level of septin 9 mRNA in various tissues of healthy state starry flounder. The EF-1α gene was used to normalise the... 61
Fig. 32. Expression level of septin 9 mRNA in the kidney, spleen, gill, liver, brain, intestine and heart of starry flounder after infection with (A): viral... 62
Fig. 33. Expression level of septin 10 mRNA in various tissues of healthy state starry flounder. The EF-1α gene was used to normalise the... 63
Fig. 34. Expression level of septin 10 mRNA in the kidney, spleen, gill, liver, brain, intestine and heart of starry flounder after infection with (A):... 64
Fig. 35. 15% SDS-PAGE analysis of septin 2 recombinant protein. Lane M: Molecular weight marker; Lane 1: purified septin 2... 65
Fig. 36. 15% SDS-PAGE analysis of septin 3 recombinant protein. Lane M: Molecular weight marker; Lane 1: purified septin 3... 66
Fig. 37. 15% SDS-PAGE analysis of septin 8 recombinant protein. Lane M: Molecular weight marker; Lane 1: purified septin 8... 66
Fig. 38. Effects of starry flounder rSeptin 2 on the phagocytosis of macrophages. The histogram of flow... 68
Fig. 39. Effects of starry flounder rSeptin 3 on the phagocytosis of macrophages. The histogram of flow... 69
Fig. 40. Effects of starry flounder rSeptin 8 on the phagocytosis of macrophages. The histogram of flow... 70