i hc thi nguyn khoa khoa hc t nhin v x hi -------- --------

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Tch dng gen cp m ho protein v ca viruS xon l gy bnh trn mt s ging c chua a ph-ng trng ti thi nguyn
Lun vn tt nghip i hc
Gio vin h-ng dn: ThS. Nguyn Th Hi Yn Sinh vin: Nguyn Th Thu H-ng

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Ni dung bo co

1. 2. 3. 4.

M u Vt liu v ph-ng php nghin cu Kt qu v tho lun Kt lun v kin ngh

M U


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C chua l loi cy c gi tr kinh t cao v ang tr thnh mt hng quan trng ca nhiu quc gia trong c Vit Nam. Hin nay cc ging c chua trng trn th gii ang phi i mt vi rt nhiu tc nhn gy bnh. Trong cc loi bnh th-ng gp c chua th bnh xon l do Tomato leaf curl virus (TLCV) v bnh xon vng l do Tomato yellow leaf curl virus (TYLCV) gy nn lm thit hi hn c. Nguyn l RNAi -c tm hiu m ra mt h-ng mi trong phng tr cc bnh do virus gy ra. Xut pht t l do trn chng ti tin hnh ti Tch dng gen CP m ho protein v ca virus xon l gy bnh trn mt s ging c chua a ph-ng trng ti Thi Nguyn

Hnh 1: Cy c chua b xo n l do virus

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NI DUNG NGHIN CU 1. Tch chit DNA tng s t mu l c chua nghi b nhim bnh xon l do virus 2. Nhn gen CP bng cp mi c hiu 3. To dng vi khun mang gen CP

NGUYN LIU V PHNG PHP NGHIN CU 1. Nguyn liu

S dng mu l c chua bi (c chua thc hay c chua ta) nghi l nhim virus xon l.

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Hnh 2.1. Cy c chua b nhim bnh (a) v cy c chua lnh (b)

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2. Ph-ng php nghin cu

1. 2. 3. 4. 5. Ph-ng php thu mu l nhim bnh K thut in di DNA trn gel agarose Ph-ng php tch chit DNA tng s t mu l thc vt Nhn gen CP bng k thut PCR Cc k thut tch dng gen
    

Tinh sch sn phm PCR Ligation sn phm PCR vi vector tch dng pBT Bin np bng ph-ng php sc nhit Chn dng bng colony - PCR Tch chit DNA plasmid nhng dng d-ng tnh

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Kt qu v tho lun

1. Kho st v thu thp mu

Bng 1. Cc a im kho st v c im nhim bnh STT 1 2 a im kho st H. ng H H. V Nhai Ging c chua C chua lai C chua lai C chua ta C chua lai 3 4 H. Ph L-ng H. Ph Bnh C chua ta C chua lai c im nhim bnh +++ ++ 0 ++ + +++

C chua ta ++ 5 TP. Thi Nguyn Ch thch: (+++) nhim bnh nhiu; (++) nhim bnh trung bnh; (+) nhim bnh t; (0) khng nhim bnh

Ti li u chia s t i: wWw.SinhHoc.edu.vn 2. Tch chit v tinh sch DNA tng s t mu l c chua

1 2 3 4 5 6 7 8 9 10

Hnh 3.1. nh in di DNA tng s trn gel agarose 0,8% (1 -5) - DNA tng s ca mu QT1- QT5 (6 -10) - DNA tng s ca mu PL1- PL5

Ti li u chia s t i: wWw.SinhHoc.edu.vn 3. Nhn on CP ca virus bng k thut PCR

M 1 2

750 bp 500 bp

600 bp

Hnh 3.2. nh in di sn phm PCR nhn gen CP M -Thang DNA chun 1kb 1 - QT1 2 - PL1

Ti li u chia s t i: wWw.SinhHoc.edu.vn 4. Ligation DNA vi vector tch dng v bin np vo E. coli DH5

pBT 2705bp

Hnh 3.3. S vector tch dng pBT

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Hnh 3.4. Kt qu bin np vector ti t hp vo t bo E. coli DH5E

Ti li u chia s t i: wWw.SinhHoc.edu.vn 5. Kt qu colony - PCR nhng khun lc trng

M 1 2 3

750 bp

730 bp

Hnh 3.5. Kt qu in di sn phm colony - PCR vi cp mi PUC18 M - thang DNA chun 1 kb 1 - QT1; 2 - QT3; 3 - PL1

Ti li u chia s t i: wWw.SinhHoc.edu.vn 6. Kt qu tch plasmid 1 2 3

Hnh 3.6. Kt qu in di sn phm tch chit DNA plasmid 1 - 3: DNA plasmid nh ng dng khu n l c d-ng tnh

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Kt lun v kin ngh

I. Kt lun: 1. tch chit -c DNA tng s t l c chua nghi nhim bnh xon l do virus ca cc ging c chua thu thp t hai a im l huyn Ph L-ng v x Quyt Thng tnh Thi Nguyn. 2. nhn -c gen CP c kch th-c khong 600 bp t mt s mu l c chua nghi nhim bnh xon l do virus bng k thut PCR. 3. tch dng phn t gen CP ca virus xon l gy bnh trn c chua a ph-ng trng huyn Ph L-ng v x Quyt Thng tnh Thi Nguyn v l-u gi chng vi khun mang gen ti th- vin gen ca Vin Cng ngh Sinh hc, Vin Khoa hc v Cng ngh Vit Nam.

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II. Kin ngh:



Tip tc c trnh t gen CP ca nhng chng virus trn lm c s cho cc nghin cu tip theo. Tin hnh thit k vector chuyn gen v to cy c chua khng virus gy bnh xon l do TYLCV.

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