Nghiên cứu làm sạch bệnh virus cho cây tỏi (allium sativum l) bằng kỹ thuật nuôi cấy meristem

NGUYN THI THANH PHNG * B GIO DC V O TO TRNG I HC BCH KHOA H NI - NGUYN THI THANH PHNG CHUYấN NGNH: CễNG NGH SINH HC NGHIấN CU LM SCH BNH VIRUS CHO CY TI (ALLIUM SATIVUM L) BNG K THUT NUễI CY MERISTEM LUN VN THC S KHOA HC CHUYấN NGNH: CễNG NGH SINH HC * KHO 2009 - 2011 H NI NM 2011 B GIO DC V O TO TRNG I HC BCH KHOA H NI NGUYN THI THANH PHNG NGHIấN CU LM SCH BNH VIRUS CHO CY TI (ALLIUM SATIVUM L) BNG K THUT NUễI CY MERISTEM Chuyờn ngnh : Cụng ngh Sinh hc LUN VN THC S KHOA HC CHUYấN NGNH: CễNG NGH SINH HC NGI HNG DN KHOA HC: PGS.TS NGUYN TH Lí ANH H Ni Nm 2011 i LI CAM OAN Tôi xin cam đoan công trình nghiên cứu riêng Kết nghiên cứu luận văn kết lao động tác giả Các số liệu kết trình bày luận văn trung thực ch-a đ-ợc công bố công trình khác Tôi xin cam đoan giúp đỡ cho việc thực luận văn đ-ợc cảm ơn thông tin trích dẫn luận văn đ-ợc rõ nguồn gốc Tác giả luận văn Nguyễn Thị Thanh Ph-ơng ii LI CM N Để hoàn thành luận văn tốt nghiệp này, cố gắng thân nhận đ-ợc nhiều quan tâm giúp đỡ thầy cô, bạn bè ng-ời thân Tr-ớc tiên, xin đ-ợc bày tỏ lòng biết ơn sâu sắc tới cô giáo, nhà giáo -u tú PGS.TS Nguyễn Thị Lý Anh Viện Tr-ởng Viện Sinh học Nông nghiệp Tr-ờng Đại học Nông nghiệp Hà Nội tận tình h-ớng dẫn, giúp đỡ trình thực đề tài hoàn thành luận văn Tôi xin đ-ợc gửi lời chân thành cảm ơn tới tập thể thầy cô giáo Viện Sinh học Nông nghiệp Tr-ờng Đại học Nông nghiệp Hà Nội tập thể thầy cô giáo Viện Công nghệ sinh học thực phẩm, Viện Đào tạo Sau đại học Tr-ờng Đại học Bách Khoa Hà Nội nhiệt tình giúp đỡ điều kiện thuận lợi cho thời gian thực đề tài Đặc biệt, xin gửi lời cảm ơn chân thành tới chồng tôi, anh Đinh Tr-ờng Sơn ng-ời bên động viên lúc khó khăn Đông thời, xin chân thành cảm ơn gia đình, bạn bè, đồng nghiệp ng-ời ủng hộ giúp đỡ trình học tập thực luận văn Hà Nội, ngày 26 tháng năm 2011 Tác giả luận văn iii Nguyễn Thị Thanh Ph-ơng MC LC Trang ph bỡa .i Li cam oan ii Li cm n iii Mc lc .iv Danh mc cỏc ch vit tt vii Danh mc cỏc bng viii Danh mc cỏc hỡnh v, th .ix CHNG I M U 1.1 t 1.2 Mc ớch, yờu cu 1.2.1 Mc ớch 1.2.2 Yờu cu CHNG II TNG QUAN TI LIU 2.1 Gii thiu v cõy ti (Allium sativum L) 2.6 Nhng thit hi virus gõy trờn cõy ti 2.2 Tỡnh hỡnh nghiờn cu v bnh virus trờn cõy ti Vit Nam v th gii 2.2.1 Tỡnh hỡnh nghiờn cu v bờnh virus ca cõy ti Vit Nam 2.2.2 Tỡnh hỡnh nghiờn cu v bnh virus ca cõy ti trờn th gii 2.3 Truyn lan ca virus h hnh ti 10 2.4 Phũng tr bnh virus hi hnh ti 11 2.4.1 Cỏc bin phỏp nhm ngn chn kh nng lõy lan ca vect truyn bnh 11 2.4.2 Hn ch s truyn lan ca cỏc virus qua vect truyn bnh 11 iv 2.4.3 Cỏc bin phỏp to ging sch bnh .12 2.5 Cỏc phng phỏp chun oỏn bnh virus 16 2.5.1 Phng phỏp lõy nhim c gii: 16 2.5.2 Phng phỏp ELISA (Enzym linked immunosorbent assay): 16 2.5.3 Phng phỏp RT PCR (Reverse Transcription Polymerase Chain): 17 2.6 Nhng nghiờn cu v nuụi cy nh sinh trng (meristem) trờn cõy ti (Allium sativum L.) 18 CHNG III I TNG, NI DUNG V PHNG PHP NGHIấN CU .21 3.1 i tng, vt liu, a im v thi gian nghiờn cu 21 3.1.1 i tng nghiờn cu 21 3.1.2 Vt liu thớ nghim 21 3.1.3 a im nghiờn cu 21 3.1.4 Thi gian nghiờn cu .21 3.2 Ni dung nghiờn cu 21 3.2.1 Cỏc nghiờn cu tin l c s cho nghiờn cu lm sch bnh virus trờn cõy ti (Allium sativum L) 21 3.2.2 Thu thp mu bnh v xỏc nh mu b nhim bnh virus t cỏc cõy ti thu thp ngoi ng rung 23 3.2.3 Nghiờn cu lm sch bnh virus cho cõy ti ta (Allium sativum L) 23 3.2.4 Nghiờn cu nhõn nhanh cõy ti sch virus 24 3.2.5 Nghiờn cu to cõy hon chnh .25 3.3 Phng phỏp nghiờn cu 26 3.3.1 Phng phỏp thu thp mu 26 3.3.2 Phng phỏp nuụi cy mụ t bo 26 3.3.3 Quy trỡnh tỏch nh sinh trng 27 3.3.4 Phng phỏp ỏnh giỏ sch virus ca cõy ti bng phng phỏp lõy nhim c gii .28 3.3.5 Phng phỏp ỏnh giỏ sch virus ca cõy ti bng phng phỏp RTPCR (Reverse Transcriptase-polymerase chain reaction) 28 3.3.6 Phng phỏp x lý s liu 30 v CHNG IV KT QU NGHIấN CU V THO LUN 31 4.1 Cỏc nghiờn cu tin l c s cho nghiờn cu lm sch bnh virus trờn cõy ti (Allium sativum L) 31 4.1.1 Nghiờn cu ch kh trựng to vt liu u .31 4.1.2 Nghiờn cu xỏc nh mụi trng nuụi cy tỏi sinh chi t meristem .33 4.2 Thu thp mu bnh v xỏc nh mu b nhim bnh virus t cỏc cõy ti thu thp ngoi ng rung 42 4.3 Nghiờn cu lm sch virus cho cõy ti ta (Allium sativum L) 46 4.4 Nghiờn cu nhõn nhanh cõy ti sch virus 52 4.3.1 Nhõn nhanh cõy ti sch virus bng chi n .53 4.3.2 Nhõn nhanh cõy ti sch virus bng tỏi sinh chi t callus 57 4.4 Nghiờn cu to cõy hon chnh 61 CHNG V KT LUN V NGH .65 5.1 Kt lun 65 5.2 ngh 67 TI LIU THAM KHO 68 PH LC 73 vi DANH MC CC CH VIT TT TST Cht iu tit sinh trng BA 6-Benzyl adenin Ki Kinetin NAA naphthuy acetic acid IBA - indolebutyric acid 2,4D Dichlorophenoxyacetic acid THT Than hot tớnh MS Murashige v Skoog, 1962 /C i chng CT Cụng thc CTAB Cetyl trimetyl amonium bromid PCR Polymerase Chain Reaction RT-PCR Reverse transcriptase-PCR Taq Taq Polymerase EtBr Ethyl bromid TN Thớ nghim CTPB Cõy ch th cú biu hin nhim virus vii DANH MC CC BNG Bng 4.1: nh hng ca HgCl2 0,1% n kh nng sng v sch ca mu cy meristem (sau tun) 31 Bng 4.2: nh hng ca Sodium Dichloroisocyanurate (NaDCC) n kh nng sng v sch ca mu cy meristem (sau tun) .32 Bng 4.4: nh hng ca kinetin n kh nng phỏt sinh hỡnh thỏi ca meristem (sau tun) 36 Bng 4.5: nh hng ca 2,4D n kh nng phỏt sinh hỡnh thỏi ca meristem (sau tun) 38 Bng 4.6: nh hng ca t hp BA + 2,4D n kh nng tỏi sinh ca meristem (sau tun) 40 Bng 4.7: nh hng ca t hp BA NAA n s phỏt sinh hỡnh thỏi ca meristem (sau tun) 41 Bng 4.8 Chun oỏn, phỏt hin cõy b nhim bnh virus ca cỏc mu ti thu thp ngoi ng rung bng phng phỏp s dng cõy ch th 43 Bng 4.9: nh hng ca kớch thc n kh nng tỏi sinh v lm sch virus ca meristem (sau tun) 46 Bng 4.10: Kt qu kim tra sch bnh ca cõy tỏi sinh t cỏc kớch thc meristem khỏc .47 Bng 4.11: nh hng ca vic kt hp tỏch meristem v x lý ribavinin n kh nng tỏi sinh v sch virus ca meristem (sau tun) 50 Bng 4.12: nh hng ca BA n s nhõn nhanh chi ti (sau tun) 53 Bng 4.13: nh hng ca NAA n s nhõn nhanh chi ti (sau tun) 55 Bng 4.14: nh hng ca t hp NAA v BA n s nhõn nhanh chi ti (sau tun) 56 Bng 4.15: Nhõn nhanh cõy ti sch virus bng tỏi sinh chi t callus 58 Bng 4.16 nh hng ca IBA n s r ca chi ti sch virus .61 Bng 4.17 nh hng ca than hot tớnh (THT) n s r ca cõy ti sch virus (sau tun nuụi cy) 62 Bng 4.18 nh hng ca t hp than hot tớnh (THT) vi IBA hoc NAA n s r ca cõy ti sch virus (sau tun nuụi cy) 63 viii DANH MC CC HèNH V, TH Biu 4.1: nh hng ca BA n s phỏt sinh hỡnh thỏi meristem 35 Biu 4.2: nh hng ca Ki n s phỏt sinh hnh thỏi meristem .37 Biu 4.3: nh hng ca kớch thc n kh nng tỏi sinh v lm sch virus 47 Biu 4.4: nh hng ca vic kt hp tỏch meristem v x lý húa cht ribavinin n kh nng tỏi sinh v sch virus 51 Biu 4.5: Nhõn nhanh cõy ti sch bnh virus bng tỏi sinh chi t callus 59 Hỡnh 4.1 nh hng ca BA kh nng phỏt sinh hỡnh thỏi meristem .36 Hỡnh 4.2 nh hng ca 2,4D v BA n kh nng phỏt sinh hỡnh thỏi meristem .39 Hỡnh 4.3: nh hng ca BA v 2,4D n kh nng phỏt sinh hỡnh thỏi ca meristem .41 Hỡnh 4.4: Mu ti biu hin triu chng bnh virus ngoi ng rung 43 Hỡnh 4.5: Biu hin bnh sc vng trờn cõy ch th rau mui 44 Hỡnh 4.6: Kt qu kim tra RT PCR 45 Hỡnh 4.7: Kt qu kim tra RT PCR sch bnh virus ca cỏc mu tỏi sinh t meristem cỏc kớch thc khỏc 49 Hỡnh 4.8: Kt qu kim tra RT PCR sch bnh virus ca cỏc mu tỏi sinh t meristem mụi trng cú b sung ribavirin .52 Hỡnh 4.9: nh hng ca BA n s nhõn nhanh chi ti .54 Hỡnh 4.10: nh hng ca NAA n s nhõn nhanh chi .56 Hỡnh 4.11: nh hng ca t hp BA v NAA n s nhõn nhanh chi .57 Hỡnh 4.12: Tỏi sinh to chi ti sch virus t callus 60 Hỡnh 4.13 : Cõy ti hon chnh 64 ix + Dựng dao mi mỏc tỏch b cỏc lỏ bao cũn li, nh sinh trng l l phn li, hỡnh cu trong, rt tn cựng phớa + Dựng dao phu thut ct phn hỡnh cu ny, tớnh t trờn xung di vi kớch thc 0,1 - 1,5mm v a vo mụi trng dinh dng nhõn to ó chun b sn QUY TRèNH LY NHIM NHN TO KIM TRA SCH VIRUS - Cõy ch th l cõy rau mui (Chenopodium album L) - Ngun lõy nhim: mu ti thu thp ngoi ng rung v mu ti tỏi sinh t nh sinh trng - Cỏch lõy nhim: Theo phng phỏp ca Hill (1984) v Kado (1972) Bc 1: Nghin cõy bnh bng ci, chy s sch khay ỏ lnh vi t l 1g lỏ 25ml m phosphate (KH2PO4, K2H PO4, hoc NaH2 PO4, Na2H PO4) 0,01M, pH = 7,0 Bc 2: Vt b phn x lỏ v bn, cho bt cacboradum bng ht u vo ci dch chit va nghin v khuy u Bc 3: Dựng bụng sch nhỳng dch cõy cú cha bt cacborandum ri ch lờn mt lỏ ca cõy ch th theo chiu t b lỏ ti u mỳt ton b thao tỏc c tin hnh 15 phỳt Bc 4: cõy iu kin cỏch ly v quan sỏt triu chng hỡnh thnh Lu ý: Cõy thớ nghim c búng ti ngy trc lõy nhim QUY TRèNH TCH CHIT RNA TNG S Trc chit m chit CTAB cha 2% CTAB, 2M NaCl, 0.025M EDTA, 0.1 Tris HCl pH = v 2% PVP ó b sung Betamecraptalthanol b nhit 60oC 30 phỳt (T l 1ml m CTAB:10 àl Beta-ME) 76 Bc 1: Ly 0.1g mụ lỏ bnh nghin vi 1àl m CTAB bng chy ci s Hỳt 600àl dch chit vo tube 1.5ml Bc 2: Ly tõm nhanh 10000v/1phỳt phỳt thu dch Bc 3: Thờm Chloroform: Isoamyl (24:1) vo ng vi th tớch tng ng, o u ng lnh -20oC 10 phỳt Bc 4: Ly tõm bng mỏy ly tõm bn 12000v/1phỳt phỳt, thu dch ni Bc 5: Chit li ln hai bng Chloroform:Isoamyl (24:1) lng tng ng nh bc v bc thu dch ni Bc 6: Kt ta RNA vi LiCl 10M (b xung LiCl vi th tớch bng ẳ th tớch dch ni), o u, ly tõm bng 10 phỳt loi b dch ni Bc 7: Thu ly cn RNA tng s Bc 8: Ra cn bng ethanol 70% ln Bc 9: cn RNA khụ t nhiờn nhit phũng thi gian 30 phỳt Bc 10: Hũa tan cn bng 50 ul m TE, bo qun -80oC 77 KT QU X Lí S LIU THNG Kấ BALANCED ANOVA FOR VARIATE CAO CAY FILE T3 16/ 9/** 14:59 PAGE THI NGHIEM 3: ANH HUONG CUA BA DEN TAI SINH MERISTEM VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 55.4775 9.24624 ****** 0.000 * RESIDUAL 14 296029E-01 211449E-02 * TOTAL (CORRECTED) 20 55.5071 2.77535 BALANCED ANOVA FOR VARIATE CHOI/MAU FILE T3 16/ 9/** 14:59 PAGE THI NGHIEM 3: ANH HUONG CUA BA DEN TAI SINH MERISTEM VARIATE V004 CHOI/MAU LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 3.57403 595671 243.84 0.000 * RESIDUAL 14 342001E-01 244286E-02 * TOTAL (CORRECTED) 20 3.60823 180411 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T3 16/ 9/** 14:59 PAGE THI NGHIEM 3: ANH HUONG CUA BA DEN TAI SINH MERISTEM MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 CT6 CT7 NOS 3 3 3 CAO CAY 4.52000 4.33000 4.33000 2.49000 2.31000 0.850000 0.200000 CHOI/MAU 0.820000 1.33000 1.88000 1.41000 1.00000 0.860000 0.560000 SE(N= 3) 0.265486E-01 0.285357E-01 5%LSD 14DF 0.805279E-01 0.865552E-01 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T3 16/ 9/** 14:59 PAGE THI NGHIEM 3: ANH HUONG CUA BA DEN TAI SINH MERISTEM F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY CHOI/MAU GRAND MEAN (N= 21) NO OBS 21 2.7186 21 1.1229 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.6659 0.45984E-01 1.7 0.0000 0.42475 0.49425E-01 4.4 0.0000 78 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T4 16/ 9/** 15:22 PAGE THI NGHIEM 4: ANH HUONG CUA Ki DEN TAI SINH MERISTEM VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 57.5985 9.59974 ****** 0.000 * RESIDUAL 14 368026E-01 262876E-02 * TOTAL (CORRECTED) 20 57.6353 2.88176 BALANCED ANOVA FOR VARIATE CHOI/MAU FILE T4 16/ 9/** 15:22 PAGE THI NGHIEM 4: ANH HUONG CUA Ki DEN TAI SINH MERISTEM VARIATE V004 CHOI/MAU LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 599829 999714E-01 114.72 0.000 * RESIDUAL 14 122000E-01 871429E-03 * TOTAL (CORRECTED) 20 612029 306014E-01 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T4 16/ 9/** 15:22 PAGE THI NGHIEM 4: ANH HUONG CUA Ki DEN TAI SINH MERISTEM MEANS FOR EFFECT CT$ CT$ NOS CAO CAY CHOI/MAU CT1 4.52000 0.820000 CT2 4.52000 1.00000 CT3 4.53000 1.15000 CT4 3.65000 1.27000 CT5 2.49000 1.17000 CT6 0.810000 1.00000 CT7 0.400000 0.780000 SE(N= 3) 0.296016E-01 0.170434E-01 5%LSD 14DF 0.897881E-01 0.516963E-01 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T4 16/ 9/** 15:22 PAGE THI NGHIEM 4: ANH HUONG CUA Ki DEN TAI SINH MERISTEM F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY CHOI/MAU GRAND MEAN (N= 21) NO OBS 21 2.9886 21 1.0271 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.6976 0.51271E-01 1.7 0.0000 0.17493 0.29520E-01 2.9 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T5 16/ 9/** 15:29 PAGE THI NGHIEM 5: ANH HUONG CUA 2,4D DEN TAI SINH MERISTEM 79 VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 43.7474 10.9369 ****** 0.000 * RESIDUAL 10 240043E-01 240043E-02 * TOTAL (CORRECTED) 14 43.7714 3.12653 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T5 16/ 9/** 15:29 PAGE THI NGHIEM 5: ANH HUONG CUA 2,4D DEN TAI SINH MERISTEM MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 NOS 3 3 CAO CAY 4.52000 0.470000 0.320000 0.280000 0.000000 SE(N= 3) 0.282868E-01 5%LSD 10DF 0.891327E-01 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T5 16/ 9/** 15:29 PAGE THI NGHIEM 5: ANH HUONG CUA 2,4D DEN TAI SINH MERISTEM F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY GRAND MEAN (N= 15) NO OBS 15 1.1180 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.7682 0.48994E-01 4.4 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T6 16/ 9/** 15:37 PAGE THI NGHIEM 6: ANH HUONG CUA TO HOP BA + 2,4D DEN TAI SINH MERISTEM VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 27.2060 9.06867 ****** 0.000 * RESIDUAL 164022E-01 205028E-02 * TOTAL (CORRECTED) 11 27.2224 2.47477 BALANCED ANOVA FOR VARIATE CHOI/MAU FILE T6 16/ 9/** 15:37 PAGE THI NGHIEM 6: ANH HUONG CUA TO HOP BA + 2,4D DEN TAI SINH MERISTEM VARIATE V004 CHOI/MAU 80 LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 1.84342 614475 155.56 0.000 * RESIDUAL 316003E-01 395003E-02 * TOTAL (CORRECTED) 11 1.87503 170457 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T6 16/ 9/** 15:37 PAGE THI NGHIEM 6: ANH HUONG CUA TO HOP BA + 2,4D DEN TAI SINH MERISTEM MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 NOS 3 3 CAO CAY 4.35000 1.27000 0.920000 0.570000 CHOI/MAU 1.85000 1.55000 1.31000 0.780000 SE(N= 3) 0.261424E-01 0.362860E-01 5%LSD 8DF 0.852478E-01 0.118325 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T6 16/ 9/** 15:37 PAGE THI NGHIEM 6: ANH HUONG CUA TO HOP BA + 2,4D DEN TAI SINH MERISTEM F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY CHOI/MAU GRAND MEAN (N= 12) NO OBS 12 1.7775 12 1.3725 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.5731 0.45280E-01 2.5 0.0000 0.41286 0.62849E-01 4.6 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T7 16/ 9/** 15:47 PAGE THI NGHIEM 7: ANH HUONG CUA TO HOP BA + IAA DEN TAI SINH MERISTEM VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 7.74982 2.58327 353.87 0.000 * RESIDUAL 584008E-01 730010E-02 * TOTAL (CORRECTED) 11 7.80823 709839 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T7 16/ 9/** 15:47 PAGE THI NGHIEM 7: ANH HUONG CUA TO HOP BA + IAA DEN TAI SINH MERISTEM MEANS FOR EFFECT CT$ - 81 CT$ CT1 CT2 CT3 CT4 NOS 3 3 CAO CAY 4.35000 4.90000 3.86000 2.72000 SE(N= 3) 0.493292E-01 5%LSD 8DF 0.160857 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T7 16/ 9/** 15:47 PAGE THI NGHIEM 7: ANH HUONG CUA TO HOP BA + IAA DEN TAI SINH MERISTEM F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY GRAND MEAN (N= 12) NO OBS 12 3.9575 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 0.84252 0.85441E-01 2.2 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T8 16/ 9/** 15:53 PAGE THI NGHIEM 8: ANH HUONG CUA TO HOP BA + NAA DEN TAI SINH MERISTEM VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 27.4106 9.13687 ****** 0.000 * RESIDUAL 422009E-01 527511E-02 * TOTAL (CORRECTED) 11 27.4528 2.49571 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T8 16/ 9/** 15:53 PAGE THI NGHIEM 8: ANH HUONG CUA TO HOP BA + NAA DEN TAI SINH MERISTEM MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 NOS 3 3 CAO CAY 4.35000 5.20000 3.00000 1.20000 SE(N= 3) 0.419329E-01 5%LSD 8DF 0.136739 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T8 16/ 9/** 15:53 PAGE THI NGHIEM 8: ANH HUONG CUA TO HOP BA + NAA DEN TAI SINH MERISTEM 82 F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY GRAND MEAN (N= 12) NO OBS 12 3.4375 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.5798 0.72630E-01 2.1 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CHOI FILE T10 16/ 9/** 15:55 PAGE THI NGHIEM 10: ANH HUONG CUA KICH THUOC TACH DEN KHA NANG LAM SACH BENH VIRUS VARIATE V003 CAO CHOI LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 41.6440 8.32880 568.51 0.000 * RESIDUAL 12 175803 146503E-01 * TOTAL (CORRECTED) 17 41.8198 2.45999 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T10 16/ 9/** 15:55 PAGE THI NGHIEM 10: ANH HUONG CUA KICH THUOC TACH DEN KHA NANG LAM SACH BENH VIRUS MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 CT6 NOS 3 3 3 CAO CHOI 1.24000 1.83000 3.06000 4.54000 4.97000 5.10000 SE(N= 3) 0.698815E-01 5%LSD 12DF 0.215329 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T10 16/ 9/** 15:55 PAGE THI NGHIEM 10: ANH HUONG CUA KICH THUOC TACH DEN KHA NANG LAM SACH BENH VIRUS F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CHOI GRAND MEAN (N= 18) NO OBS 18 3.4567 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.5684 0.12104 3.5 0.0000 BALANCED ANOVA FOR VARIATE CAO CHOI FILE T11 83 16/ 9/** 15:57 | | | | PAGE THI NGHIEM 11: ANH HUONG CUA RIBAVIRIN DEN KHA NANG LAM SACH BENH VIRUS VARIATE V003 CAO CHOI LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 27.2204 5.44409 ****** 0.000 * RESIDUAL 12 484018E-01 403348E-02 * TOTAL (CORRECTED) 17 27.2689 1.60405 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T11 16/ 9/** 15:57 PAGE THI NGHIEM 11: ANH HUONG CUA RIBAVIRIN DEN KHA NANG LAM SACH BENH VIRUS MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 CT6 NOS 3 3 3 CAO CHOI 4.22000 2.56000 2.05000 1.87000 0.750000 0.500000 SE(N= 3) 0.366674E-01 5%LSD 12DF 0.112985 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T11 16/ 9/** 15:57 PAGE THI NGHIEM 11: ANH HUONG CUA RIBAVIRIN DEN KHA NANG LAM SACH BENH VIRUS F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CHOI GRAND MEAN (N= 18) NO OBS 18 1.9917 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 1.2665 0.63510E-01 3.2 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T12 16/ 9/** 16:12 PAGE THI NGHIEM 12: ANH HUONG CUA BA DEN KHA NANG NHAN NHANH CHOI VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 18.4138 3.68277 ****** 0.000 * RESIDUAL 12 237995E-01 198329E-02 * TOTAL (CORRECTED) 17 18.4376 1.08457 BALANCED ANOVA FOR VARIATE CHOI/MAU FILE T12 16/ 9/** 16:12 84 PAGE THI NGHIEM 12: ANH HUONG CUA BA DEN KHA NANG NHAN NHANH CHOI VARIATE V004 CHOI/MAU LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 693850 138770 57.03 0.000 * RESIDUAL 12 292001E-01 243334E-02 * TOTAL (CORRECTED) 17 723050 425323E-01 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T12 16/ 9/** 16:12 PAGE THI NGHIEM 12: ANH HUONG CUA BA DEN KHA NANG NHAN NHANH CHOI MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 CT6 NOS 3 3 3 CAO CAY 6.50000 6.09000 5.50000 4.56000 4.00000 3.86000 CHOI/MAU 1.08000 1.52000 1.45000 1.26000 1.08000 1.00000 SE(N= 3) 0.257118E-01 0.284801E-01 5%LSD 12DF 0.792269E-01 0.877567E-01 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T12 16/ 9/** 16:12 PAGE THI NGHIEM 12: ANH HUONG CUA BA DEN KHA NANG NHAN NHANH CHOI F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE GRAND MEAN STANDARD DEVIATION C OF V |CT$ (N= 18) SD/MEAN | NO BASED ON BASED ON % | OBS TOTAL SS RESID SS | CAO CAY 18 5.0850 1.0414 0.44534E-01 0.9 0.0000 CHOI/MAU 18 1.2317 0.20623 0.49329E-01 4.0 0.0000 BALANCED ANOVA FOR VARIATE CAO CAY FILE T13 16/ 9/** 16:18 -THI NGHIEM 13: ANH HUONG CUA NAA DEN KHA NANG NHAN NHANH CHOI | | | | PAGE VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 4.28304 1.07076 754.03 0.000 * RESIDUAL 10 142006E-01 142006E-02 * TOTAL (CORRECTED) 14 4.29724 306946 BALANCED ANOVA FOR VARIATE CHOI/MAU FILE T13 16/ 9/** 16:18 PAGE THI NGHIEM 13: ANH HUONG CUA NAA DEN KHA NANG NHAN NHANH CHOI VARIATE V004 CHOI/MAU 85 LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 677760 169440 66.19 0.000 * RESIDUAL 10 256000E-01 256000E-02 * TOTAL (CORRECTED) 14 703360 502400E-01 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T13 16/ 9/** 16:18 PAGE THI NGHIEM 13: ANH HUONG CUA NAA DEN KHA NANG NHAN NHANH CHOI MEANS FOR EFFECT CT$ CT$ NOS CAO CAY CHOI/MAU CT1 6.50000 1.08000 CT2 7.86000 1.20000 CT3 7.84000 1.48000 CT4 7.84000 1.68000 CT5 7.80000 1.44000 SE(N= 3) 0.217567E-01 0.292119E-01 5%LSD 10DF 0.685560E-01 0.920477E-01 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T13 16/ 9/** 16:18 PAGE THI NGHIEM 13: ANH HUONG CUA NAA DEN KHA NANG NHAN NHANH CHOI F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY CHOI/MAU GRAND MEAN (N= 15) NO OBS 15 7.5680 15 1.3760 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 0.55403 0.37684E-01 0.5 0.0000 0.22414 0.50596E-01 3.7 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T14 16/ 9/** 16:25 PAGE THI NGHIEM 14: ANH HUONG CUA TO HOP BA + NAA DEN KHA NANG NHAN NHANH CHOI VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 6.98880 1.39776 ****** 0.000 * RESIDUAL 12 118003E-01 983356E-03 * TOTAL (CORRECTED) 17 7.00060 411800 BALANCED ANOVA FOR VARIATE CHOI/MAU FILE T14 16/ 9/** 16:25 PAGE THI NGHIEM 14: ANH HUONG CUA TO HOP BA + NAA DEN KHA NANG NHAN NHANH CHOI VARIATE V004 CHOI/MAU LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 20.7210 4.14420 522.37 0.000 * RESIDUAL 12 952015E-01 793346E-02 86 * TOTAL (CORRECTED) 17 20.8162 1.22448 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T14 16/ 9/** 16:25 PAGE THI NGHIEM 14: ANH HUONG CUA TO HOP BA + NAA DEN KHA NANG NHAN NHANH CHOI MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 CT6 NOS 3 3 3 CAO CAY 6.50000 6.89000 7.08000 6.55000 6.00000 5.20000 CHOI/MAU 1.08000 1.76000 3.02000 4.08000 3.88000 2.92000 SE(N= 3) 0.181048E-01 0.514246E-01 5%LSD 12DF 0.557872E-01 0.158457 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T14 16/ 9/** 16:25 PAGE THI NGHIEM 14: ANH HUONG CUA TO HOP BA + NAA DEN KHA NANG NHAN NHANH CHOI F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY CHOI/MAU GRAND MEAN (N= 18) NO OBS 18 6.3700 18 2.7900 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 0.64172 0.31359E-01 0.5 0.0000 1.1066 0.89070E-01 3.2 0.0000 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T15 16/ 9/** 16:29 PAGE THI NGHIEM 16: ANH HUONG CUA IBA DEN KHA NANG RA RE VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 3.20940 1.06980 198.11 0.000 * RESIDUAL 432002E-01 540002E-02 * TOTAL (CORRECTED) 11 3.25260 295691 BALANCED ANOVA FOR VARIATE SO LA FILE T15 16/ 9/** 16:29 PAGE THI NGHIEM 16: ANH HUONG CUA IBA DEN KHA NANG RA RE VARIATE V004 SO LA LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 309900 103300 13.50 0.002 * RESIDUAL 612000E-01 765000E-02 - 87 * TOTAL (CORRECTED) 11 371100 337364E-01 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T15 16/ 9/** 16:29 PAGE THI NGHIEM 16: ANH HUONG CUA IBA DEN KHA NANG RA RE MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 NOS 3 3 CAO CAY 4.70000 5.23000 5.65000 6.10000 SO LA 3.50000 3.62000 3.87000 3.87000 SE(N= 3) 0.424265E-01 0.504975E-01 5%LSD 8DF 0.138349 0.164667 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T15 16/ 9/** 16:29 PAGE THI NGHIEM 16: ANH HUONG CUA IBA DEN KHA NANG RA RE F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY SO LA GRAND MEAN (N= 12) NO OBS 12 5.4200 12 3.7150 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 0.54377 0.73485E-01 1.4 0.0000 0.18367 0.87464E-01 2.4 0.0019 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T16 16/ 9/** 16:32 PAGE THI NGHIEM 17: ANH HUONG CUA THAN HOAT TINH (THT) DEN KHA NANG RA RE VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 597800 298900 15.02 0.005 * RESIDUAL 119400 199000E-01 * TOTAL (CORRECTED) 717200 896500E-01 BALANCED ANOVA FOR VARIATE SO LA FILE T16 16/ 9/** 16:32 PAGE THI NGHIEM 17: ANH HUONG CUA THAN HOAT TINH (THT) DEN KHA NANG RA RE VARIATE V004 SO LA LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 974000E-01 487000E-01 60.87 0.000 * RESIDUAL 480001E-02 800001E-03 88 * TOTAL (CORRECTED) 102200 127750E-01 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T16 16/ 9/** 16:32 PAGE THI NGHIEM 17: ANH HUONG CUA THAN HOAT TINH (THT) DEN KHA NANG RA RE MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 NOS 3 CAO CAY 4.70000 4.23000 4.10000 SO LA 3.45000 3.68000 3.66000 SE(N= 3) 0.814453E-01 0.163299E-01 5%LSD 6DF 0.281732 0.564879E-01 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T16 16/ 9/** 16:32 PAGE THI NGHIEM 17: ANH HUONG CUA THAN HOAT TINH (THT) DEN KHA NANG RA RE F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY SO LA GRAND MEAN (N= 9) NO OBS 4.3433 3.5967 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 0.29942 0.14107 3.2 0.0052 0.11303 0.28284E-01 0.8 0.0002 | | | | BALANCED ANOVA FOR VARIATE CAO CAY FILE T17 16/ 9/** 16:34 PAGE THI NGHIEM 18: ANH HUONG CUA TO HOP THT + NAA, IBA DEN KHA NANG RA RE VARIATE V003 CAO CAY LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 303600 759001E-01 15.55 0.000 * RESIDUAL 10 487999E-01 487999E-02 * TOTAL (CORRECTED) 14 352400 251714E-01 BALANCED ANOVA FOR VARIATE SO LA FILE T17 16/ 9/** 16:34 PAGE THI NGHIEM 18: ANH HUONG CUA TO HOP THT + NAA, IBA DEN KHA NANG RA RE VARIATE V004 SO LA LN SOURCE OF VARIATION DF SUMS OF MEAN F RATIO PROB ER SQUARES SQUARES LN ============================================================================= CT$ 451560 112890 6.02 0.010 * RESIDUAL 10 187600 187600E-01 * TOTAL (CORRECTED) 14 639160 456543E-01 - 89 TABLE OF MEANS FOR FACTORIAL EFFECTS FILE T17 16/ 9/** 16:34 PAGE THI NGHIEM 18: ANH HUONG CUA TO HOP THT + NAA, IBA DEN KHA NANG RA RE MEANS FOR EFFECT CT$ CT$ CT1 CT2 CT3 CT4 CT5 NOS 3 3 CAO CAY 4.70000 5.00000 5.11000 4.90000 5.04000 SO LA 3.20000 3.60000 3.66000 3.61000 3.65000 SE(N= 3) 0.403319E-01 0.790780E-01 5%LSD 10DF 0.127087 0.249178 ANALYSIS OF VARIANCE SUMMARY TABLE FILE T17 16/ 9/** 16:34 PAGE THI NGHIEM 18: ANH HUONG CUA TO HOP THT + NAA, IBA DEN KHA NANG RA RE F-PROBABLIITY VALUES FOR EACH EFFECT IN THE MODEL SECTION - VARIATE CAO CAY SO LA GRAND MEAN (N= 15) NO OBS 15 4.9500 15 3.5440 STANDARD DEVIATION C OF V |CT$ SD/MEAN | BASED ON BASED ON % | TOTAL SS RESID SS | 0.15866 0.69857E-01 1.4 0.0003 0.21367 0.13697 3.9 0.0101 90 | | | | ... sch bnh virus cho cõy ti (Allium sativum L) bng k thut nuụi cy meristem 1.2 Mc ớch, yờu cu 1.2.1 Mc ớch S dng k thut tỏch v nuụi cy tỏi sinh nh sinh trng (meristem) nhm to cõy ti sch bnh virus. .. cu tin l c s cho nghiờn cu lm sch bnh virus trờn cõy ti (Allium sativum L) tt c cỏc thớ nghim nghiờn cu tin l c s cho nghiờn cu lm sch bnh virus trờn cõy ti (Allium sativum L) chỳng tụi s... thớch hp cho tỏi sinh nh sinh trng - Xỏc nh c kớch thc meristem thớch hp cho nuụi cy tỏi sinh cõy v lm sch bnh virus - Xỏc nh c nh hng ca ribavirin n kh nng tỏi sinh v lm sch bnh virus - Xỏc nh