GIỚI THIỆU VỀ CÂY THUỐC LÁ

5/28/2018 GII THI U V CY THU C L

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1.1.GII THIU VCY THUC L

1.1.1.Phn loi, ngun gc

Thuc l thng mi hay N. tabacum l mt trong s 65 loi c xc nhthuc chi Nicotiana. Phn loi ca Wilson C.L. v Loomis W.E. (1967) c cc nh

khoa hc ng h v trch dn nhiu nht.

H (Family): Solanaceae(H c)

Chi (Genus) Nicotiana

Loi (Species) Nicotiana tabacum L.

Trong h c c trn 85 chi vi khong 1800 loi bao gm nhiu cy cy rau, cy

cng nghip, cy dc liu, cy cnh v cy trng c gi tr kinh t khc.

Cc nh khoa hc phn chiaNicotianathnh 3 chi ph, 14 phn chi v 65 loi. Trong

s 65 loi ny th c 45 loi l cy bn a Bc v Nam M, 15 loi bn a Australia.

Smith H.H. (1979) tng hp s phn loi i vi chiNicotiananh sau:

Chi phRusticac 3 phn chi vi 9 loi,

Chi ph Tabacum c 2 phn chi vi 6 loi,

Chi phPetunioides c 9 phn chi vi 50 loi,

Trong s 65 loi trn, ch c loi N. tabacum thuc phn chi Genuinae l c

gieo trng thng mi trn khp th gii. nc Nga v mt s nc chu ngi ta

trng loiN. rusticach yu cho tiu dng ti ch. Mt s loi hoang di nhN. alata, N.

sylvestrisc trng lm cy cnh.

Xut pht t chu M, cy thuc l lan ti chu u vo th k 15 v ti chu

khong gia th k 18 v ngy nay c trng ph bin ti nhiu quc gia trn th gii

(Harland, 1981; Poceh LA, 2002). Thuc l c tnh di truyn phong ph, tnh thch ng

rng ri v di s tc ng trc tip ca con ngi, ngy nay thuc l c nhiu ctrng phm cht, ngoi hnh khc nhau. C th k n loi thuc l vng sy c hng v

c o l Virginia (Hoa K, Zimbabwe...), thuc l Oriental - c sn ca vng a

Trung Hi, x g ni ting ca Cuba v Sumatra (Indonesia).


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Vit Nam, cy thuc l ang c trng ph bin cc tnh min ni v trung

du pha Bc, cc tnh Nam Trung B, Ty Nguyn v min ng Nam B v c nh

gi l mt trong nhng cy cng nghip ngn ngy, em li hiu qu kinh t cao cho

nng dn v thu nhp quc gia (www.vinataba.com.vn).

Ngoi ra, cy thuc l cn c coi l cy m hnh tin hnh nhiu nghin cu

sinh hc c bn v chc nng, vai tr cng nh c ch hot ng ca cc gen hay protein,

cc con ng trao i cht, tng hp cc cht s cp v th cp thc vt hay tng tc

gia thc vt vi cc sinh vt gy bnh,Bn cnh vi kh nng d ti sinh v chp

nhn cc gen ngoi lai, nn cy thuc l cng c s dng lm m hnh th nghim v

pht trin nhng nghin cu ng dng nh biu hin cc protein ti t hp c gi tr nh

cc vaccine hoc dc liu, lm ngun nhin liu sinh hc thay th,

1.1.2. Tnh hnh sn xut thuc l nguyn liu

Thuc l l mt cy cng nghip ngn ngy, l mt hng tiu dng v xut khu c

gi tr kinh t cao v l ngun thu ngn sch ng k ca nhiu nc trn th gii. Thuc

l nguyn liu tham gia thng mi trn th gii ch yu c sn xut cc vng t 45

v Bc ti 30 v Nam. Sn xut thuc l nguyn liu giai on 2001-2008 ton th gii

t tng din tch khong 2,5- 3,0 triu ha v tng sn lng 4,9 - 5,4 triu tn. Thuc l

vng sy chim t trng ln vi sn lng 3,5 - 4,1 triu tn; thuc l Burley 620 - 886

ngn tn; thuc l Oriental 237 - 438 ngn tn v cc loi thuc l khc (Bng 1). Snlng cc loi thuc l nguyn liu chnh ca th gii trong nhng nm qua tng i n

nh mc d ngnh cng nghip thuc l cc nc chu nhiu p lc v a phn khng

c khuyn khch sn xut v tiu th.

Cy thuc l thu ht nhiu lao ng nng nghip (trng trt, thu hi, s ch...),

lao ng cng nghip, gp phn to cng n vic lm, tn dng lao ng ph, s dng lao

ng hp l, tng thu nhp cho ngi lao ng, lm phong ph thm c cu lun canh,

gp phn ci to t trng trt, phn b li dn c (www.vinataba.com.vn). Theo s liu

ca Hi thuc l Zimbabwe, din tch thuc l ti y ch chim 3% tng din tch tnng nghip nhng ng gp 38% tng sn phm nng nghip xut khu v em li

30% tng s ngoi t xut khu cho t nc trong 98% thuc l c xut khu vi

gi tr trn 500 triu USD/nm. Trung Quc l quc gia c sn lng thuc l nguyn

liu v thuc l iu ng u th gii. Ch tnh ring tnh Vn Nam, doanh thu v thuc

l t ti trn 13 t USD/nm vi li nhun khong 1,5 t USD v np vo ngn sch nh


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nc khong 3 t USD tng ng 72% doanh thu thu ton tnh (theo bn tin Tng

cng ty thuc l Vit Nam,www.vinataba.com.vn).

Bng 1: Sn lng cc dng thuc l nguyn liu ch yu ca th gii

n v: ngn tnNm Vng sy Burley Oriental Nu Tng2004 3757,1 886,4 350,9 195,9 5190,3

2005 4035,6 771,8 353,1 181,9 5342,4

2006 3948,5 725,3 268,5 182,1 5124,4

2007 3872,6 620,6 237,0 185,5 4915,7

2008 4185,9 743,5 259,0 184,3 5372,7

(Ngun: Universal Leaf Tobacco Company 2008 Supply and Demand)

1.1.3.Tnh hnh bnh hi cy thuc l

Cy thuc l thuc h c, l vt ch trung gian ca nhiu loi virus. Do vy, bnh

virus thng hay gp trn cy thuc l, gy nh hng nghim trng ti nng sut, c

bit l cht lng phm cht l thuc l. Ngoi ra, mt s bnh vi khun hoc su hi

cng nh hng ti s pht trin ca cy thuc l cng nh vng trng thuc l nguyn

liu.

Bnh virus

- Bnh khm l thng do cc virus khm l thuc l (Tobacco mosaic virus: TMV),

virus khm da chut (Cucumber mosaic virus: CMV), virus khoai ty Y (Potato virus Y:

PVY), virus khm l c chua (Tomato mosaic virus: ToMV) trong TMV c nh

gi l nguyn nhn ch yu gy bnh ny

- Bnh xon t, ho ngn do virus ho m c chua (Tomato spotted wilt virus: TSWV)

- Bnh xon l do virus xon l thuc l (Tobacco leaf curl virus: TLCV) v virus xon lc chua (Tomato leaf curl virus: ToLCV)

Cc bnh virus cha c thuc phng tr, bin php tt nht l cch ly vng b nhim

bnh, nh v chn hoc t cy nhim bnh.

Bnh vi khun v nm
http://www.vinataba.com.vn/http://www.vinataba.com.vn/http://www.vinataba.com.vn/http://www.vinataba.com.vn/


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- Bnh en thn do nmPhythopthora parasitica var nicotianaeBreda de Haan Tucker

- Bnh m mt cua do nm Cercospora nicotianaeEll and Ev

- Bnh phn trng do nmErysiphe cichoracerum

- Bnh thn th do nmColletotrichum nicotianae Averna

- Bnh ho r do vi khunPseudomonas solanacearumSmith

- Bnh m l do vi khunPseudomanas tabaci

Bin php hu hiu phng tr cc bnh do nm v vi khun trn cy thuc l l

s dng cc ging khng bnh, lun canh trit , x l t v phn hu c trc khi gieo

trng, s dng cc phng php ti nc v bn phn hp l v phun thuc dit nm v

vi khun thch hp khi pht hin bnh.

Din tch trng thuc l ca nc ta hin nay mc 25.000 ha v sn lng

nguyn liu mc 40.000 45.000 tn mi nm. Nguyn liu thuc l c sn xut

trong nc mi p ng c khong 60% nhu cu nguyn liu cho sn xut thuc iu

(theo bn tin Tng cng ty thuc l Vit Nam, www.vinataba.com.vn). p ng nhu

cu nguyn liu cho tiu dng trong nc v xut khu th bn cnh vic m rng din

tch cn nng cao nng sut v n nh cht lng nguyn liu ti cc vng trng. Mc

tiu trn ang gp phi mt tr ngi l dch bnh. Ti mt s vng trng thuc l ln

nh Ty Ninh, Ninh Thun, bnh xon ngn do Virus ho m c chua (Tomato spottedwilt virus, TSWV) gy thit hi nng cho sn xut khi xut hin giai on sm v

khng cho thu hoch. Theo bo co ca phng u t -pht trin thuc Cng ty c phn

Ho Vit v Phn vin Kinh t K thut Thuc l, thit hi v kinh t ch ring vng Ty

Ninh trong 3 nin v t 2001-2004 c tnh mc 40 - 50 t ng. Ti cc vng trng

pha Bc, cc bnh khm l do virus khm thuc l (tobacco mosaic virus: TMV) v virus

khm da chut (cucumber mosaic virus: CMV) gy ra c t l nhim trung bnh t 15-

20%, c khi ln ti 100% nh ti x Lu Thng Thi Nguyn v gy thit hi cho

nng dn khong 1015 t ng/ nm (Nguyn Vn Chn v Nguyn Ngc Bch, 2008).

Trong sn xut thuc l nguyn liu hin nay, ging K326 c nhp ni t M

nm 1990 c cht lng rt tt nhng kh nng khng bnh rt km nn nng sut khng

n nh. Ging thuc l C9-1 do Vin Kinh t K thut Thuc l lai to v chn lc c

nng sut cao, cht lng tt ang c m rng din tch s dng nhng cng mn cm


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vi cc bnh do virus gy hi. Do vy, to ging thuc l khng bnh virus l yu cu cp

thit ph hp vi chin lc pht trin ca ngnh thuc l Vit Nam.

1.2. S LC VVIRUS GY BNH TRN CY THUC L

1.2.1.Virus khm thuc l (tobacco mosaic virus, TMV)

TMV thuc loi Tobamovirusl mt trong nhng virus gy hi trn thc vt c

m t sm nht bi Mayer (1886), Iwanowski (1892), Allard (1914) v Stanley (1935).

Virus khm thuc l c th gy bnh trn hn 120 loi cy trng v cy di khc nhau

nhng nhiu nht l trn cy thuc l (Plant virus online:

http://www.agls.uidaho.edu/ebi/vdie/descr803.htm). Khi cy b bnh cc l non u bin

mu thnh dng khm bao gm cc vng xanh nht v xanh m khng u xen ln nhau,

hnh loang l nh da ch, hay cn gi l dng khm. TMV c lan truyn bng ngc gii do tip xc virus thm nhp vo cc t bo qua vt thng v qua kh khng l,

thn hoc r (V Triu Mn, 2007).

Hnh 1: S cu trc genome ca TMVRdRp: replicase; CP: protein v; helicase: enzyme lin quan ti qu trnh dui xon;MP: protein vn chuyn. Chiu mi tn th hin chiu dch m

TMV rt bn k c dng kt ta cn v virus l kh vn cn hot tnh gy bnh

sau mt nm (Pfleger v Zeyen, 2008). Virus ny c genome l mt si RNA n

dng, c chiu di khong 6,4 Kb (Goelet et al., 1982) (hnh 1), m ho cho cc protein

gm:protein v virus (trng lng 17,6 kDa), protein vn chuyn (trng lng phn t

30kDa) gi vai tr trong s chuyn dch ca virus gia cc t bo v cc protein m ha

cho cc enzyme tham gia vo qu trnh phin m nh replicase (trng lng phn t

183kDa) v qu trnh tho xon bt u phin m nh helicase (126 kDa) ( Collmer et al.,

1983).

6,395 nt

MP

RdRp CP

Helicase


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1.2.2.Virus ho m c chua (Tomato spotted wilt virus, TSWV)

TSWV c pht hin l nguyn nhn gy bnh trn cy c chua vo nm 1919 ti

Australia. Cho ti nay, TSWV c nh gi l virus gy hi ph rng vi hn 170 loi

thc vt khc nhauti nhiu vng kh hu nhit i v cn nhit i trn th gii (Zitterv Daughtrey, 1989). Cy nhim virus thng xut hin nhng vt m vng nu hoc

vt hoi t trn l, cung hay gn l bin dng v cui cng ngn cy xon li v ho r

(V Triu Mn, 2007; Zitter v Daughtrey, 1989). Virus ny gy nn thit hi nghim

trng cho cy trng h c ti Bc M v Canada nh c chua, khoai ty, thuc l,. Ti

Vit Nam, virus ho m c chua c pht hin trn cy c chua ti Hocmn (V Triu

Mn, 1998) v thuc l (Nguyn Ngc Bch v Bi Cch Tuyn, 1998). Ti mt s vng

trng thuc l ln nh Ty Ninh, Ninh Thun, bnh xon ngn do virus ho m c chua

gy thit hi nng cho sn xut khi xut hin giai on sm v khng cho thu hoch.Virus ho m c chua thuc loi Tospovirus, h Bunyaviridae, c cu to dng

thin th kch thc 70-90nm. Virus c lan truyn ch yu thng qua b tr Thrips

tabaci, Thrips setosus, Thrips parmi, theo kiu truyn thng (V Triu Mn, 2007).

Genome ca TSWV gm ba si RNA n m (negative) hoc lng tnh (ambisense) l

si L (8,9 kb), si M (4,9 kb) v si S (2,9 kb) (hnh 2). Si L m ha cho cc replicase

lin quan ti qu trnh phin mca virus trong khi si M m ha cho protein vn chuyn

(NSm) v protein v (GNv GC) cn si S m ha cho nucleprotein (N) v protein c ch

qu trnh bt hot gen (NSs) (Kormelink et al., 1992; Takeda et al., 2002; Tsompana et

al., 2005; Sin et al., 2005).

Hnh 2: S cu trc genome ca TSWV

RdRp

NSm GN/GC

NSs N

8,9kb

4,9kb

2,9kb

L

M

S


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RdRp: replicase; Nsm: protein vn chuyn; GN/GC: protein v;NSs: nhn t c ch qu trnh bt hot gen; N: nulceoproteinChiu mi tn th hin chiu dch m

1.3. PHNG PHP PHNG CHNG BNH VIRUS HI THC VT

C khong hn 2000 loi virus thc vt c pht hin v nghin cu, trong s

khong mt na l nhng loi gy hi chnh cho cy trng. Mc thit hi do cc

bnh virus gy ra cho cy trng rt nghim trng c th ln ti 95 - 100%. Hn na, s

thit hi khng nhng ch dng mc suy gim v nng sut m cn nh hng c

n cht lng sn phm thu hoch. Cc v d v thit hi do bnh virus gy ra nc ta

c th k n cc dch virus vng ln xon l trn la ng bng sng Cu Long nhng

nm va qua v gn y l dch la ln li mi xut hin cc tnh pha Bc e danghim trng n an ninh lng thc. Cho n nay cha c mt loi thuc bo v thc

vt no c kh nng chng li bnh do virus gy ra, c nhiu bin php khc nhau ang

c s dng kim sot bnh virus trn cy trng tuy vy mi bin php u c nhng

hn ch nht nh chng hn nh:

(i) s dng ging cy trng c kh nng khng bnh virus. Bin php ny b gii hn bi

s lng rt hn ch cc ging cy trng khng virus cng nh cc ngun vt liu di

truyn khng virus phc v cho cng tc lito ging;

(ii) s dng cc ht ging, c, cy ging sch bnh c xem nh vy l bin php quan

trng nht trong nhiu loi cy trng. Tuy nhin, cc loi cy trng c th b nhim virus

trong qu trnh canh tc trn ng rung;

(iii) kim sot cn trng trung gian truyn bnh v loi b cc loi k ch trung gian nh

c di, Tuy nhin nhiu loi virus nh TMV ly truyn qua ng c gii, virus ly

lan trc tip qua kh khng hoc vt thng trn vt ch. Hn na, vic lm dng thuc

tr su c thgy nhim mi trng, gy nguy him cho nng dn v ngi tiu dng.

Bn cnh , virus c th c ly nhim vi mt s lng rt t mi gii truyn bnh v

d: ch cn 1 con ry nu/cy la c th truyn virus ln la c (RGSV). La chn

thi im canh tc trnh vi thi im pht trin ca trung gian gy bnh cng l mt

phng php c hiu qu trong vic phng chng bnh virus. Chng hn bin php n

ry gp phn hn ch s bng pht ca dch vng ln xon l la ti mt s tnh pha


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Nam ncta. Cc phng php canh tc khc nh lun canh, xen canh, gi v, cng

ang c nhiu ni trn th gii v Vit Nam p dng nhm gim bt thit hi do bnh

virus gy ra. Tuy nhin phng php canh tc ty thuc nhiu vo c im kh hu th

nhng catng a phng nn kh p dng c trn din rng;

(iv) cng ngh sinh hc: trong 15 nm qua mt s virus c th kim sot c thng

qua bin php to cy trng chuyn gen mang cc gen hoc on gen c ngun gc t

chnh cc virus gy bnh (Reddy el al., 2009). Cc cu trc gen c ngun gc t virus

gy bnh c s dng chuyn vo cy trng to tnh khng c th l cc loi khc

nhau nh: cc cu trc ca vius theo chiu xui (sense) hay chiu ngc (antisense)

(Boogaart et al., 1998), cc cu trc dng kp tc (inverted repeats/hairpin) (Yin et al.,

2005) v cc miRNA nhn to c ch l cc trnh t gen ca virus gy bnh (Niu et la.,

2006; Qu v Fang, 2008). Thnh cng trong vic ng dng cng ngh sinh hc trong toging cy trng khng bnhvirus c nh du bi cng b ca Beachy v cng s nm

1986 khi nhng cy thuc l chuyn gen m ha protein v ca TMV (cu trc dng

sense) biu hin tnh khng vi virus ny (Beachy, 1990; Register v Nelson, 1992).

Sau Beachy v cng s pht trin nhng th nghim ca mnh thnh phng php

CP to cy trng khng virus. Phng php ny c p dng to nhng dng cy

chuyn gen khng nhiu loi virus gy bnh trn thc vt hai l mm v sau l thc

vt mt l mm (xem tng quan ca Hull v Davies, 1992). Tuy vy, cc cy chuyn gen

ny ch khng c duy nht mt chng virus mang gen m ha protein v m n chuynvo. iu ny gy tr ngi trong vic pht trin cc ging cy trng khng virus da trn

phng php CP do a dng ca virus thng kh cao (Scholthof et al., 1993). Tng

t nh phng php CP, biu hin gen m ha cho enzyme lin quan ti qu trnh t sao

chp hoc sao m (cc replicase) cng c th to cc cy chuyn gen khng virus. Cng

nghin cu trn i tng TMV, Carr v cng s nhn thy nhng dng thuc l mang

gen m ha cho replicase c khi lng phn t l 183kDa hoc 54kDa c kh nng

khng TMV tt hn nhng dng cy chuyn gen CP. Biu hin replicase ca virus khoai

ty X (potato virus X: PVX) hay virus khm da chut (cucumber mosaic virus: CMV)chng minh hiu qu bo v thc vt ca phng php ny (Braun v Hemenway, 1992;

Anderson v cng s, 1992). Tuy vy, tng t nh phng php CP, phng php bo

v qua trung gian gen replicase cng ch c tc dng trn tng chng virus. Hn na, gen

m ha cho cc replicase thng rt ln, gy tr ngi khi biu hin trn thc vt. Nhm

khc phc nhng hn ch ca cu trc dng sense, cc cu trc dng antisense mang


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mt on DNA c trnh t b sung vi RNA ca virus c th c ch s phin m vi

nhng trnh t RNA tng ng c nghin cu v pht trin. Nhiu dng thuc l,

khoai ty, c chua, khng virus c to ra da trn k thut ny.Vic s dng

nhng on DNA thay v c on gen m ha lm nhng dng cy chuyn gen c kh

nng khng vi nhiu dng virus thay v ch mt vi dng nht nh (Boogart et al, 1998).

Cho ti nhng nm cui th k 20, cu trc dng kp tc (ihpRNA) hay k thut

RNAi c xem l mt k thut hin i v hu hiu nht trong vic chng li cc bnh

do virus gy ra thc vt. Nm 2004, Baulcombe cng b c ch hot ng ca

siRNA v coi l mt c ch quan trng trong vic khng li virus thc vt. Cc bc

chnh trong k thut ny bao gm: (1) thit k cc vector chuyn gen mang cu trc

RNAi, (2) bin np vector chuyn mang cu trc RNAi vo cy thng qua vi khun

Agrobacterium tumefaciens lm bt hot cc mRNA ca virus gy bnh, (3) sng lc cccy chuyn gen mang cu trc RNAi v kim tra tnh khng virus ca cc cy chuyn gen

(.Tenllado et al., 2004). Tnh hiu qu ca k thut RNAi trong vic to cy trng chuyn

gen m ho protein ca virus (gen m ha protein v CP, enzyme phin m RdRp,) c

kh nng khng li chnh virus c chng minh bng thc t trong nhng nghin

cu to cy trng khng cc loi virus khc nhau nh: Potato virus Y PVY (Waterhouse

et al., 1998;Smith et al.,2000), cucumber mosaic virus CMV (Kalantidis et al., 2002),

Plum pox potyvirus PPV (di Nicola-Negri et al., 2005), Tobacco mosaic virus TMV v

Potato virus Y PVY (Sun et al., 2005), , Tomato yellow leaf curl virus TYLCV (Zrachyaet al., 2007)Rice tungro bacilliform virus RTBV (Tyagi et al., 2008) African cassava

mosaic virus ACMV (Vanderschuren et al., 2009 ) Trong nhng nghin cu ny, cu

trc RNAi c cha trnh t gen ca virus lp li o chiu thng c s dng

chuyn vo cy v s c biu hin thnh RNA si i dng kp tc (hairpin RNA,

hpRNA) trong cy chuyn gen t kch thch c ch RNAi hot ng khi c s xm

nhp ca virus vo cy. Ngi ta nhn thy rng khi vng m ca hpRNA c lp

li vi mt trnh t intron (ihpRNA) th kt qu ihpRNA to ra s bt hot gen l cao

nht (Smith et al., 2000;Wesley et al., 2001). Nm 2007, Bonfim v cng s to rac mt dng cy u chuyn gen khng virus BGMV (Bean golden mosaic virus) vi

tnh khng ln n 93%. Cng nm ny, Shinichiro Kamachi v cng s cng b kt

qu to ra c mt s dng thuc l chuyn gen CP trong cu trc ihpRNA c kh nng

khng cao vi virus cucumber green mottle mosaic virus CGMMV n th h T2 (12/14

s cy kim tra) v nhng siRNA c pht hin trong nhng dng cy chuyn gen
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ny. Ni chung, hu ht cc cy chuyn gen lm chm s tch ly virus v lm chm hoc

gim nh cc triu chng bnh (Gottula et al, 2009)

Cho n nay c cc loi cy trng chuyn gen khng bnh virus c cng

nhn v trng thng mi nh: u chuyn gen khng bnh m vng (papaya

ringspot virus, PRSV) c cng nhn v trng M, Trung Quc, Philippine; B ao

chuyn gen khng ba loi vi rt Cucumber mosaic virus, Watermelon mosaic virus,

Zucchini yellow mosaic virus, c cng nhn v trng M; t v c chua chuyn

gen khng Cucumber mosaic virus, c cng nhn v trng Trung Quc Ngoi ra

rt nhiu cc loi cy trng chuyn gen khng bnh virus khc ang trong giai on kho

nghim c cng nhn l ging cy trng thng mi nh: Sn chuyn gen khng

African cassava mosaic virus (Begomovirus); Ng chuyn gen khng Maize steak virus

(Mastrevirus); Khoai ty chuyn gen khng ng thi 3 loi virus Potato virus X(Potexvirus), Potato virus Y (Potyvirus), Potato leafroll virus (Polerovirus); La chuyn

gen khng Rice Tungro viruses (Tungrovirus); Khoai lang chuyn gen khng Sweet

potato feathery mottle virus (Potyvirus).(Reddy et al, 2009).

1.4. GII THIU VRNAi ( RNA interference)

1.4.1.Lch spht hin

RNA interference (RNAi) l c ch t nhin ca t bo sng c th lm bt hot

s hot ng ca mt gen no . Hin tng ny c quan st ln u tin trn nhng

cy thuc l khng bnh do virus m vng thuc l (Tobacco ring spot virus, TRSV)

gy ra sau ly nhim ln hai (Wingard, 1928), trn cc cy chuyn gen c biu hin c

ch phin m nh RNA ''chiu i m''(antisense RNA) (Ecker v Davis, 1986) v nhiu

bo co ca nhng cuc th nghim gy bt ng c thc hin bi cc nh khoa hc

thcvt M v H Lan vo nhng nm u thp nin 1990 trn i tng cy d yn

tho (petunia) (Napoli et al., 1990; van der Krol et al., 1990). Trong nghin cu bin i

mu hoa ca petunia, cc nh khoa hc c lm m thm mu tm ca hoa bng cch

a vo mt on gen tng hp chalcone (Chalcone synthase, CHS - gen m ha cho

enzym quy nh mu sc hoa) di s iu khin ca promoter mnh (promoter 35S).

Nhng kt qu khng c nh mong mun, thay v lm m thm mu hoa th kt qu

li cho ra cy choa khm trng hoc trng tuyn (hnh 1.4). iu ny chng t gim

bt lng sc t ca hoa hay lng chalcone c tng hp t i hay s hot ng ca


11/17

gen tng hp chalcone b c ch. Hin tng ny cc nh khoa hc t thut ng l

"co-suppresion" ngha l "ng c ch" bi v s biu hin ca gen chuyn vo v gen

ni sinh trong hoa petunia u b c ch nh nhau. Mt iu th v na trong nghin cu

ny l mu trng ca cnh hoa petunia c th c quan st qua cc th h sau v dn dn

cc hoa chuyn sang mu tm.

Sau t lu, mt th nghim nhm tng mu cam ca nm Neurospora crassa thng

qua vic chuyn mt gen c chc nng to ra carotenoid. Tuy nhin thay v to ra mu

cam nh mong mun th nm hu nh khng th hin. Hin tng ny ccc nh khoa

hc t tn l "quelling" (Romano & Macino, 1992; Cogoni et al., 1994). Nghin cu su

hn v kiu hnh ca cy chng minh rng s mt iu khin l do s c ch sau phin

m ca biu hin gen thng qua vic tng t l phn hu mRNA (Van Blokland et al.,

1994).

A B C

Hnh 1.1. V d v cy hoa d yn tho trong cc gen quy nh s c thoa b lm bt hot bi RNAi (Napoli et al., 1990)

A, l cy wild-type(WT); B v C l cy c cha gen chuyn m gy ra skm hm biu hin c gen ni sinh v gen chuyn lm xut hin cc vngtrng khng mu trn cnh hoa

Nm 1995, hai nh khoa hc Guo v Kemphues th dng RNA i m lm

bt hot s biu hin ca gen par-1 ca Caenorhabditis elegans tm hiu chc nng

ca chng. ng nh d kin, s c mt ca RNA i m ph hu s biu hin ca

par-1, nhng tht ngc nhin khi ly nhim on RNA ''chiu dch m''(sense RNA) th

kt qu cng tng t. iu ny gy ra mt s lng tng bi v iu mong i nht ca

cc nh khoa hc l phn t RNA s bt cp vi mRNA chiu thun (sense mRNA) ca

gen par-1 nhm c ch kh nng m ha protein ca n.
http://en.wikipedia.org/wiki/Wild-typehttp://en.wikipedia.org/wiki/Wild-typehttp://en.wikipedia.org/wiki/Wild-type


12/17

Mt vi nm sau, cc nh nghin cu virus thc vt cng quan st c nhng

hin tng tung t trong cc th nghim nhm nng cao tnh khng virus ca thc vt.

H tin rng RNA virus c to ra bng chuyn gen cng c kh nng c ch s sao

chp virus (Covey et al., 1997). Th nghim ngc li, cc trnh t ngn ca gen thc vt

c a vo virus cng cho thy gen ch b bt hot trong cy bnh. Hin tng nyc gi tn l "virus-induced gene silencing" (VIGS) v tng hp cc hin tng ny

c gi chung l scm gen sau phin m (post transcriptional gene silencing,PTGS)

(Ratcliff et al.,1997).

Cc hin tng trn tht s khng gii thch c cho n nm 1998 khi Fire v

Mello ln u tin ly nhim si i RNA (double stranded RNA, dsRNA) gm cc on

dch m v i m vo C. elegans.H pht hin vic tim cc phn t RNA m ho mt

protein c bp khng lm thay i hnh vi ca giuntrn. M di truyn trong RNA c gi l''sense''. Tim RNA ''antisense'' cng khng vic g. Tuy nhin, khi Fire v Mello tim c RNA

''sense'' v ''antisense'' vo c th giun trn, h quan st thy giun c nhng chuyn ng l, c

th l co git. Cc chuyn ng tng t cng xy ra nhng con giun hon ton thiu mt gen

chu trch nhim to protein c bp ny. Hin tng c gii thch rng khi RNA ''sense'' v

''antisense'' gp nhau, chng kt hp vi nhau v hnh thnh RNA chui i v cho rng phn t

chui i c th lm cm lng gen mang cng mt m vi phn t RNA ny. Fire v Mello

kim tra gi thuyt bng cch tim cc phn t RNA chui i cha cc mt m di truyn sn

xut nhiu protein khc ca giun trn. mi th nghim, vic tim RNA chui kp mang mt mt

m di truyn xc nh khin gen cha mt m ny ngng hot ng. Protein c m ho bigen cng khng cn c to ra.

Sau mt lot cc th nghim tng t, Fire v Mello suy lun dsRNA c th lm cc gen

ngng hot ng v thut ng RNA interference (RNAi) c t tn cho c ch ny.

C ch RNAi hu hiu i vi gen m mt m ca n khp vi mt m ca phn t RNA c

tim. Ngoi ra, c ch RNAi c th lan gia cc t bo v thm ch c di truyn sang i sau.

Ch cn tim mt lng nh phn t RNA chui i l c th t c mt kt qu no . Do

vy, Fire v Mello cho rng c ch RNAi l mt tin trnh xc tc. Hai nh bc hc cng b

pht hin ca h trn tp ch Nature vo ngy 19/2/1998. Khm ph ca h lm sng t nhiuquan st th nghim mu thun v kh hiu, ng thi tit l mt c ch t nhin kim sot

dng thng tin di truyn. Khm ph cng bo hiu s khi u ca mt lnh vc nghin cu mi

(Fire et al., 1998).

Tng t, khi ly nhim mt vi on dsRNA vo trong mt t bo th dn n s

bt hot ca mt h gen. Hn na, khi ly nhim dsRNA vo b phn sinh dc (gut) ca
http://en.wikipedia.org/wiki/Post_transcriptional_gene_silencinghttp://en.wikipedia.org/wiki/Post_transcriptional_gene_silencinghttp://en.wikipedia.org/wiki/Post_transcriptional_gene_silencinghttp://en.wikipedia.org/wiki/Post_transcriptional_gene_silencing


13/17

C. elegansth s bt hot gen khng ch xy ra th h cha m m cn xy ra trong th

h con ci ca chng. Tim nng to ln ca RNAi c Fire v Timmons pht hin ra khi

th nui nhng con giun nh c thit k biu hin dsRNA tng ng vi gen unc-

22 ca C. elegans. Tht ngc nhin nhng con giun ny pht trin vi s v hiu gen

unc-22. Sau , khi th nghim dsRNA vi s lng ln giun t bin cho thy mt sln gen b loi b th c ch ca hin tng RNAi c tm ra. Sau mt thi gian

di khm ph ra c ch RNAi, Fire v Mello vinh d nhn gii thng Nobel v Y hc

v sinh l hc cho pht minh quan trng ny.

Bng vic khm ph vai tr ca dsRNA lin quan n vic phn hy cc mRNA

trong sut qu trnh tm hiu c ch ca RNAi v PTGS ngi ta nhn ra rng c ch

chung nh hng n biu hin ca gen i vi qu trnh quelling, co -supression, RNAi

hay PTGS u tng t nh nhau v trong c ch ca chng u c s hin din ca phnt siRNA (small intefering RNA) dn n vic hnh thnh cc phc cht siRNA -RISC

(RNA-induced silencing complex) v cui cng l s phn hy cc phn t RNA dn n

vic c ch s biu hin kiu hnh ca gen. Tu chung li ngi ta t mt thut ng mi

l RNA silencing (Baulcombe, 2004).

1.4.2. C chhot ng ca RNAi thc vt

RNAi l c ch c ch s biu hin vt cht di truyn giai on RNA. thc vt

cba con ng c ch RNA (hnh 1.5). Con ng u tin l s c ch gen sau phinm (post-transcriptional gen silencing, PTGS) qua trung gian l siRNA (short interfering

RNA). Con ng th hai gm c cc miRNA (micro-RNA). Con ng th ba l s

cm gen phin m (transcriptional gene silencing - TGS) bng cch lin kt vi qu trnh

bin i nhim sc th trc tip qua siRNA, n bao gm s methyl ha histone v DNA.

Vai tr sinh hc khc nhau ca cc con ng ny c chng minh, bao gm khng

virus, iu khin biu hin gen v s ngng t cht nhim sc thnh th d nhim sc

(Baulcombe, 2004).

Cc phn t quan trng tham gia vo c ch RNAi ny l hai enzym Dicer vArgonaute (AGO) trong phc h RISC (RNA-induced silencing complex) v nhng RNA

c ch nh (short interfering RNA, siRNA) hay miRNA.

S c ch RNA thng qua siRNA: xy ra tng bo v l con ng quan trng trong

t bo thc vt nhim virus ni m dsRNA c th sao chp gin tip hoc mt c im


14/17

cu trc th cp ca RNA virus si n. Trong trng hp l virus DNA thc vt,

dsRNA c th c to ra nh qu trnh phin m b sung gi nhau (Hamilton &

Baulcombe, 1999 ; Baulcombe, 2004). C ch cng xy ra tng t nh ng vt. Khi

c s xm nhp ca chui xon kp RNA vo tng bo, Dicer (mt thnh vin trong h

RNaseIII) - mt loi ribonucleases c hiu cho dsRNA vi s tiu hu mt phn t ATPlp tc ct nhng chui kp RNA ny ra nhng on ngn hn, khong 21 -25 nucleotid,

gi l siRNA (Hammond, 2000; Zamore, 2000; Bernstein, 2001). Khi lai nhng RNA

cho thy l nhng si i c cha nhm phosphat tn cng u 5. Sau khi b ct

ngn bi Dicer, chui kp siRNA c tch ra lm hai chui n, v ch mt chui n

RNA vi u 5' c lc bt cp base (base-pairing) nh nht c chn tip tc lin kt

vi Argonaute. Qu trnh la chn chui n RNA ny xy ra trong phc h RISC, trong

c cha Argounaute v Helicase (Schwarz et al., 2002). Phn t ATP phn tch

siRNA si i to thun li cho RISC hot ng, phc hRISC sau nhn bit cc

phn t phin m mRNA ca t bo c trnh t tng ng vi trnh t ca on chui

n siRNA lc ny ang c mt trong phc h RISC (Song et al., 2004). Sau khi nhn

dng mRNA qua vic bt cp cc base tng ng vi trnh t ca chui n siRNA,

mRNA b ct t khong gia ca chui kp siRNA-mRNA thnh nhng on nh

khong 12 nucleotid t u 3. Sau khi b ct t, mRNA nhanh chng b tiu hu bi

cc RNA nuclease (hnh 1.2).


15/17

Hnh 1.2. M phng cc con ng cm gen thng qua RNA (Matzke &Matzke, 2004)

AGO, ARGONAUTE; dsRNA, double-stranded RNA RNA si i; PTGS,posttranscriptional gene silencingS cm gen sau phin m; RDR, RNA

dependent RNA polymerase RNA polymerase ph thuc RNA; RISC,RNA-induced silencing complexPhc h kch thch cm gen RISC; RITS,RNA-induced initiation of transcriptional gene silencing Phc h khi scm gen phin m RITS; siRNA, short interfering RNA RNAi c ch nh;miRNA, miro-RNA; TGS, transcriptional gene silencing S cm gen sauphin m

S c ch RNA thng qua miRNA: Ngay sau khi pht hin ra c ch RNAi, ngi ta

nhanh chng nhn ra rng, c ch ny c s dng t lu trong t bo bi mt h

thng iu ha biu hin gen gi l micro-RNA (miRNA). miRNA l mt loi RNA nh

di 21-24 nucleotid khng m ha c chc nng ch yu l iu khin m sau phin m

v cp baz c trnh t b sung gn nh hon ton vi mRNA ch (Bartel, 2004). C ch

miRNA c nhiu im tng ng vi c ch siRNA. Trong t bo ng vt, khi nhng

phn t miRNA u tin (gi l pre-miRNAs) c to ra trong nhn qua qu trnh phin

RITS Phc hp kch thch RISC

Phc

hp

Methyl ha DNA/histone phn hy mRNA ngn cn dch m


16/17

m t gen, pre-miRNAs c ct gtbi mt enzym c mt trong nhn gi l Drosha

to thnh nhng si pre-miRNAs. Cc pre-miRNAs sau c di chuyn ra ngoi

tng bo v tng tc vi enzym Dicer ri k tip l phc h RISC nh trnh by trn.

Cn thc vt, miRNA c tin hnh nh Dicer v phn ln ch xy ra nhn, v c

mt protein lin kt dsRNA c hiu l HYL1. miRNA ng vt thng lin kt vivng 3 khng dch m (untranslated region, UTR) ca mRNA, trong khi miRNA thc

vt c gn kt vi trnh t m ha v thm ch vng5 UTR ca mRNA (Sunkar & Zhu,

2004). Mt iu ng ch l, thc vt miRNA c ch biu hin mRNA ch yu qua

s tiu hy mRNA, trong khi ng vt miRNA can thip ch yu bng cch c ch

qu trnh dch m ca mRNA (Rhoades et al., 2002 ; Jones-Rhoades & Bartel, 2004).

tng ng trong trnh t ca miRNA vi mRNA trong phc h RISC s quyt nh

mRNA s b ct v tiu hy lm bt hot qu trnh dch m ca mRNA. Nu trnh t ca

miRNA ging ht vi trnh t ca mRNA, mRNA s b tiu hy. Nu trnh t ca

miRNA tnh t u 5' ch cn ti thiu tng ng vi mRNA t nucleotid v tr s 2 n

s 8 th c ch miRNA s kch hot, tc l ch chn ng s dch m m khng lm tiu

hy mRNA (Denli et al., 2004).

Phn ln miRNA thc vt iu khin ch ca chng bng cch ct mRNA trc

tip vng m ho (Bartel & Bartel, 2003;Bartel, 2004; Dugas & Bartel, 2004). Mt vi

miRNA thc vt c chng minh l mu cht trong s pht trin l v ra hoa thng qua

gen ch l cc nhn t phin m lin quan

(Reinhart et al., 2002; Bartel & Bartel, 2003).Khc vi nhn t phin m, miRNA c th nhm ti mt gii hn phin m rng (Jones-

Rhoades & Bartel, 2004; Adaiet al., 2005). Biu hin ca chng chng t chng c

vai tr quy nh s pht trin theo hng c hiu m hoc phn ng vi mt gii hn p

lc mi trng (Reinhart et al., 2002; Jones-Rhoades & Bartel, 2004; Sunkar & Zhu,

2004; Adai et al., 2005) Cc kiu biu hin khc nhau v s phong ph tim nng gen

ch mRNA gi rng miRNA c th iu khin nhiu qu trnh l sinh v pht trin v

c th ng mt vai tr trc tip trong s di chuyn t t bo ny sang t bo khc thc

vt (Dugas & Bartel, 2004; He & Hannon, 2004; Kidner & Martienssen, 2005).

S khc bit quan trng gia siRNA v miRNA u tin l ngun gc ca chng,

miRNA bt ngun t cc locus h gen trong khi siRNA thng bt ngun t mRNA,

transposon, virus hoc DNA tp sc. Th hai, miRNA c ch bin t phin m c th

to cu trc kp tc RNA trong khi siRNA c ch bin t RNA si i di hoc dng


17/17

kp tc ko di. Th ba, si i miRNA:miRNA c to ra t tin phn t kp tc

miRNA, dn ti tch lu nhiu siRNA khc nhau t cc si dsRNA di ny. Th t, trnh

t miRNA thng gn bo th nhng sinh vt c quan h gn gi, trong khi siRNA ni

sinh him khi bo th (Bartel, 2004). Nhng s khc nhau ny l c s thc t phn

bit v ch gii nhng siRNA ni sinh v miRNA mi khm ph (Ambros et al., 2003a).

Con ng lm cm gen th ba thc vt c lin quan ti s methyl ho DNA v s

c ch phin m. Bng chng u tin cho dng cm gen ny l khm ph trn thc vt

m gen chuyn v RNA virus c xu hng methyl ho DNA to nn trnh t nucleotid

c hiu (Wassenegger et al., 1994; Mette et al., 2000; Jones et al., 2001). Gn y,

nhng pht hin ny c m rng bng vic quan st s methyl ho DNA thng qua

siRNA trn thc vt c lin kt vi bin i histone (Ziberman et al., 2003) v trong

s nhn i nm men, s hnh thnh si tp sc xung quanh vng tm ng c linkt vi siRNA (Volpe et al., 2002). Mt vai tr quan trng ca s cm gen mc

nhim sc th l bo v genome trnh nhng phn hu gy ra bi transposon

(Baulcombe, 2004).

Mt ngun gc chung ca c ba con ng ny l c th bi v c nhng v d ca

mi dng trn ng vt, nm v thc vt. Cy xanh th khc ch chng c kh nng

cha c ba dng cm gen, trong khi nhng sinh vt khc c th thiu mt mt hoc nhiu

hn cc con ng ny. V d, nm men ny chi dng nh thiu c ba con ng v

cho n nay tt c nhng v d cm gen t nhin tm thy ng vt u l miRNA(Bartel, 2004). Mc d RNA ngoi bo t bo ng vt c v khi u cho c ch

RNAi c in bao gm siRNA, n khng tht s r rng con ng siRNA c hiu c

dnh lu hay khng. C th l RNA ngoi bo c tuyn vo con ng miRNA

(Baulcombe, 2004).
http://hinari-gw.who.int/whalecomwww.sciencedirect.com/whalecom0/science?_ob=ArticleURL&_udi=B6WSN-4BJK582-F&_user=10&_coverDate=01%2F23%2F2004&_alid=843576362&_rdoc=1&_fmt=full&_orig=search&_cdi=7051&_sort=d&_docanchor=&view=c&_ct=1&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=41ba72121a9fc7ac9f5274dc62157b0f#bib3http://hinari-gw.who.int/whalecomwww.sciencedirect.com/whalecom0/science?_ob=ArticleURL&_udi=B6WSN-4BJK582-F&_user=10&_coverDate=01%2F23%2F2004&_alid=843576362&_rdoc=1&_fmt=full&_orig=search&_cdi=7051&_sort=d&_docanchor=&view=c&_ct=1&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=41ba72121a9fc7ac9f5274dc62157b0f#bib3http://hinari-gw.who.int/whalecomwww.sciencedirect.com/whalecom0/science?_ob=ArticleURL&_udi=B6WSN-4BJK582-F&_user=10&_coverDate=01%2F23%2F2004&_alid=843576362&_rdoc=1&_fmt=full&_orig=search&_cdi=7051&_sort=d&_docanchor=&view=c&_ct=1&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=41ba72121a9fc7ac9f5274dc62157b0f#bib3http://hinari-gw.who.int/whalecomwww.sciencedirect.com/whalecom0/science?_ob=ArticleURL&_udi=B6WSN-4BJK582-F&_user=10&_coverDate=01%2F23%2F2004&_alid=843576362&_rdoc=1&_fmt=full&_orig=search&_cdi=7051&_sort=d&_docanchor=&view=c&_ct=1&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=41ba72121a9fc7ac9f5274dc62157b0f#bib3http://hinari-gw.who.int/whalecomwww.sciencedirect.com/whalecom0/science?_ob=ArticleURL&_udi=B6WSN-4BJK582-F&_user=10&_coverDate=01%2F23%2F2004&_alid=843576362&_rdoc=1&_fmt=full&_orig=search&_cdi=7051&_sort=d&_docanchor=&view=c&_ct=1&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=41ba72121a9fc7ac9f5274dc62157b0f#bib3http://hinari-gw.who.int/whalecomwww.sciencedirect.com/whalecom0/science?_ob=ArticleURL&_udi=B6WSN-4BJK582-F&_user=10&_coverDate=01%2F23%2F2004&_alid=843576362&_rdoc=1&_fmt=full&_orig=search&_cdi=7051&_sort=d&_docanchor=&view=c&_ct=1&_acct=C000050221&_version=1&_urlVersion=0&_userid=10&md5=41ba72121a9fc7ac9f5274dc62157b0f#bib3

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