Chan doan virut Rota trong phong thi nghiem 18 1.2.. Tinh hinh nghien cuu va phat trien vacxin Rota tren Th^ gioi va trong nuoc 26 1.5.1.. Tinh hinh nghien cuu va phat triln vacxin tai V
Trang 1DAI HOC QUOC GIA HA NQl
TRlTOfNG DAI HQC KHOA HOC TU^ NHIEN
DANG THI NGAN HA
NGHIEN ClTU TINH ANTOAN VA KHA NANG GAY DAP LTNCi MIEN D I C H T R E N K H I Macaca mulatta C U A 3 C H U N d VIRUT ROTA H E G I O N G G O C D E SAN XUAT VACXIN ROTA
Chuyen nganh: Vi sinh vat hoc
Ma so: 62 42 40 01
LUAN AN TIEN SI SINH H O C
NGLT^I Hl/^NG DAN KHOA HOC:
1 PGS.TS L E THI LUAN
2 PGS.TS NGUYEN DANG HIEN
D A I H O C Q U O C GtA HA N O l TRUNG TAM T H 6 N G TIN THU VIET
Ha Noi- 2010
Trang 2MUC LUC
• •
LOICAMON
LOI CAM DOAN
CAC KI HIEU VA C H C VIET T A T
MUCLUC
DANH MUC CAC BANG
DANH MUC CAC HINH
MCJDAU 1 CHUONG I: TONG QUAN 3
1.1 Virut Rota 3
1.1.1 Ljch siJ phat hien virut Rota 3
1.1.2 Phan loai virut Rota 3
1.1.3 Hinh thai va cau true 4
1.1.4 Cac thanh phan hoa hoc cua virut Rota va chuc nang 6
1.1.5 Phan nhom va typ huyet thanh 10
1.1.6 Tinh chat hoa ly 11
1.1.7 Nuoi cay 13
1.1.8 Su nhan len ciia virut Rota trong te bao 13
1.1.9 Kha nang gay benh 16
1.1.10 Cache gay benh 17
1.1.11 Cac trieu chung cua benh do virut Rota 18
1.1.12 Di^u tri 18
1.1.13 Chan doan virut Rota trong phong thi nghiem 18
1.2 Dich te hoc 20
1.2.1 Nguon benh 20
1.2.2 Lua tuoi mac benh 20
1.2.3 Mua benh 20
1.2.4 Duong lay truyen 21
1.2.5 Milndich 21
1.3 Tinh hinh nhiem virut Rota tren Th^ gioi 22
1.4 Tinh hinh nhiem virut Rota tai Viet Nam 23
1.5 Tinh hinh nghien cuu va phat trien vacxin Rota tren Th^ gioi va trong
nuoc 26 1.5.1 Tinh hinh nghien cuu phat trien vacxin tren Th^ gioi 26
1.5.2 Tinh hinh nghien cuu va phat triln vacxin tai Viet Nam 32
Trang 31.6 Tieu chuin ciia T6 chuc Y t^ Th^ gioi ve chiing san xuat vacxin Rota 34
1.6.1 Cac thuat ngu 34
1.6.2 Ki€m tra ngu6n nguyen lieu dau 35
1.6.3 Chiing virut 37
1.7 H6 so he thdng chiing gidng ung vien de san xuat vacxin Rota tai Viet
Nam 41
1.7.1 H6 so tom tk cac buoc cay truyen chiing giong goc G1P[8] de tao
chiing ung vien cho san xuat vacxin 42
1.7.2 H6 so tom tat cac buoc cay truyen chiing giong goc G1P[4] de tao
chiing ung vien cho san xuat vacxin 43
1.7.3 H6 so tom tat cac buoc cay truyen chiing giong goc G4P[6] de tao
chiing ung vien cho san xuat vacxin 44
CHLTONG II DOI TlJONG, VAT LIEU VA PHlTONG PHAP NGHIEN ClTU
45 2.1 Doi tugng nghien cuu 45
2.1.1 Chiing giong goc (Master seed) 45
2.1.2 Chiing san xuat vacxin (Working seed) 46
2.2 Nguyen vat lieu va phuong phap kiem tra he thong chiing 46
2.2.1 Phuang phap xet nghiem vi khuan va nam 46
2.2.2 Phuorng phap thu Mycoplasma bang phuang phap PCR 47
2.2.3 Phuang phap thu vi khuan lao 49
2.2.4 Phuong phap xac dinh virut gay hap phu hong cau 50
2.2.5 Thu nghiem tac nhan ngoai lai tren te bao than tho so sinh 51
2.2.6 Thu nghiem tac nhan ngoai lai tren te bao than khi tien phat 52
2.2.7 Thu nghiem tac nhan ngoai lai tren te bao Vero 53
2.2.8 Thu nghiem tac nhan ngoai lai tren te bao Hep2C 53
2.2.9 Xac dinh nhan dang virut Rota 54
2.2.10 Phuang phap xac dinh hinh the virut Rota bang kinh hien vi dien tu
62 2.2.11 Phuang phap xac dinh an toan tren dong vat thi nghiem 66
2.2.12 Phuang phap dap ung mien dich tren khi 68
2.2.13 Xac dinh trinh tu gen 4(VP4), 6(VP6), 9(VP7), 10(NSP4) cua virut
Rota 71 2.2.14 Phuang phap thong ke xii ly so lieu 77
CHITONG III: KET QUA VA BAN LUAN 78
3.1 Ket qua kiem dinh tinh an toan he thong chiing virut Rota 78
Trang 43.1.1 Kilm tra vi khuAn va nam va vi khuan lao 78
3.1.2 Kiem tra tac nhan ngoai lai trong hon dich chiing virut Rota bang
phuang phap PCR 79
3.1.3 Kiem tra tac nhan ngoai lai gay hiiy hoai te bao trong hon dich chiing
virut Rota tren nuoi cay te bao va hap phu hong cau 80
3.1.4 Kiem tra an toan tren dong vat thi nghiem 82
3.1.5 Quan sat hinh anh chiing virut Rota giong goc tren kinh hien vi dien
tu 93
r r r r
3.1.6 Ket qua nhan dang chiing virut Rota giong goc va chiing san xuat 94
3.1.7 Ket qua giai trinh tu gen he thong chiing virut Rota 101
3.1.8 Xac dinh 11 doan ARN ciia chiing giong goc bang SDS-Page 129
3.2 Ket qua dap ung mien dich he thong chiing tren khi thuc nghiem 130
3.2.1 Ket qua khang the trung hoa tren he thong 3 chiing 130
3.2.2 Ket qua khang the trung hoa tren he thong chiing doi chung 134
KET LUAN 137 KIENNGHI 138 CAC CONG TRINH DA CONG BO CO LIEN QUAN DEN LUAN AN 139
TAI LIEU THAM KHAO