Tap chi Khoa hpc va Phat trien 2012: Tap 10, so 2: 263-271 TRUONG DAI HOC NONG NGHIEP MA NOI
NHAN GIONG LNULTROim LAN DENDROBIUMFIMBRIAWKim^^.
(HOANG THAO LONG NHAN)
In Vitro Micropropagation of Dendrobium Fimbriatum Hook.
Nguygn Thi So-n, Nguyen Thi Ly Anh, Vu Ngpc Lan, Tran The Mai
Vien Sinh hgc Ndng nghiep, Trmrng Dgi hgc Ndng nghiep Hd Ngi Email ciia tac gia lien lac: [email protected] Ngay gu'i bai: 29.11.2011 Ngay chap nhpn: 25.03.2012
T 6 M TAT
Lan Dendrobium fimbriatum Hoolt. (Hodng tliao Long nhan) ici ioii lan d^p du'p'c si> dpng \km canh va iam thu6c, dang de dpa tuyet ctiung. Ti^n lianh nghien ciru nhan in vitro vm mpc dich bao tdn va phat trien ngudn gen ioai lan quy. K^t qua nghien ci>u da chi ro: Nguyen ii^u su" dung ia qud ian 3 thang tu6i; ftfloi tru'd'ng thich hp'p cho nay mim va phat sinh protocorm cura hpt la moi tru'd'ng MS + 100ml ND + lOg saccharoza + 6,0g agar/Iit moi triro'ng; lUloi tru'd'ng nhan nhanh protocorm tdt nhit i i mdi tru'd'ng KC + 100mi ND -•• 10g sacaroza + 60g khoai tay + 6,0g agar/iit mdi tru'd'ng; IMdi tru'd'ng IMS + 100mi ND + 20g sacaroza + 60g chudi chin + 6,0g agar'iit mdi tru'd'ng ia thich hp'p nhat cho nhan nhanh choi in vitro; Moi tru'd'ng t^o cay hoan chinh la RE + lOg sacaroza + 1g THT + 6,0g agar/iit mdi tru'd'ng,
TCF khda: Dendrobium fimbriatum Hook., qua tan, nhan nhanh, protoconn.
SUMMARY
The orchid Dendrobium fimbriatum Hook (Hoang Thao Long Nhan), a beautiful orchid species, which is currently used for house decoration as well as medicine, is in danger of being extinction. In vrtro studies of orchid Dendrobium fimbriatum Hook. (Hoang Thao Long Nhan) were conducted in order to conserve and increase the genetic pool of this precious orchid species. The results showed that the suitable materials used for micropropagation were 3 months old fruits. Most suitable medium for gemiination and protocomn formation was MS+IOOmi ND+10g saccharose+S.Og agar/iiter. The optimum medium for propagation of protocomis was KC +100ml ND+10g saccharose+SOg potato+6.0g agar/liter. The most appropriate medium for fast in vitro propagation of shoots was MS+100ml ND+20g saccharose+SOg ripe banana+S.Og agar/iiter. The medium used for growing of complete plants was RE+10g sacharose +1g THT+6.0g agar/iiter.
Keywords: Dendrobium fimbriatum Hook., fruit, propagations, protocorms.
l . D A T V A N D E nhien, trong qua trinh phfit trien kinh te xa hpi, do nhiSng nguyen nhan khac nhau, Loai Ian Dendrobium fimbriatum nhieu loai Hoang thao da bi tuyet chung Hook. (Hofing thao Long nhan) dUdc dung hoac bi de doa tuyet chung. Mot sd loai lan lfim canh vi hoa to, cd hUdng thom, mau thudc chi lan Hoang thao da cd trong danh vfing cam vdi mdi xe tua, b hong cd mot luc Do cila "Sfich dd Viet Nam" Hofing thao ddm mfiu nau tim hoac tim dd gio'ng mfit Long nhan (Dendrobium fimbriatum rdng rfi't dep (Theo cud'n "Dupc dien" Trung Hook.) dUpc xip b mUc VU (sdp nguy c&'p) Qudc nfim 1997; D6 Huy Bich, 2004). Tuy (Nguyln Ti^n Ban, 2007).
Nhan gr6ng in vitro loai lan Dendrobium fimbriatum WooV.. (Hoang thjo long nhan) Doi vdi nhi^u lofii lan qu^ hid^m hi de
dpa tuyet chung trong ti^ nhidn thudng dUpc bao ton nhd phUdng thUc nay mim tii h^t (Kauth, 2005). Hi^n nay, vdi cdng ngh^ nhfin gid'ng in vitro, h^ sd nhfin gidng ti^ mpt qua lan lfi rfi't ldn, tU vfii ngfin d^n mdt tri^u cfiy con (Trfin Vfin Minh, 2001). Da cd cfic tac gia trong vfi ngofii nUdc nhfin gidng lan Dendrobium sp. bfing phUdng phfip gieo hat lan tren nfi'n mdi trUdng MS cd hS sung 0,5mg/l NAA (Luan vfi cs,, 2006). Nguyln Vfin Song (2011) ciing nhfin nhanh in vitro lofii lan rCtng cd nguy cd tuy^t chiing vdi ngudn nguyen li^u ban diu lfi gieo hat tren moi trUcfng MS + 15% dudng sacarose + 2,0mg/l BA.
Dendrobium fimbriatum Hook, lfi lofii lan qu^, vide nghien cilu vfi Ung dung ky thufit nudi cfi'y mo de nhan in vitro lofii lan nfiy lfi hofin tofin mdi d Viet Nam. De gdp phfin bao tdn va phat trifin ngudn gen lofii lan Dendrobium fimbriatum Hook, (mdt lofii bi danh gia lfi sap nguy cfi'p vfi phai ddi mat vdi nguy cd tuy^t chung cao trong tU nhien trong mdt tUdng lai khdng xa) thi viec nghien cUu nhfin gid'ng in vitro loai lan nfiy la viec lfim cfi'p thie't va cd y nghia.
2. VAT LIEU VA PHl/C^NG PHAP 2.1. Doi tifpTng, vat lieu, dia diem va thdi gian nghien cifu
Ddi tUdng: Hofing thao long nhan (Dendrobium Gmbriatum Hook.) tU Vi^n Sinh hpc Ndng nghigp - TrUdng DH Ndng Nghidp Hfi Ndi nghien cctu thu thfip tai Lai Chau.
Vfit lieu:
- Mfiu thi nghiem: Qua lan Dendrobium fimbriatum Hook.
Mdi trudng: Murashige & Shoog (1962), Vacin & Went (1949), Knudson C (1965), Robert Ernst (1979)
- Cfic djch nghien: Nudc dUa (ND), tao, chudi, khoai tfiy, cfi rdt...
Cfich lfim dich nghi&n: qua tao dd, cfi rdt di ca vd riia sach, chudi tiSu chfn bo vd xay nho min ridng tilng logi; khoai tfiy d^ ca vd rtta sach luOc chin dung ca nUdc lupc xay nhd min.
Dia di^m: Vi^n Sinh hoc Ndng nghidp - Trudng Dpi Hoc Ndng Nghidp Hfi N$i - Trau Qu^ - Gia Lfim - Hfi Npi.
Thdi gian nghidn cUu: tii thfing 6 nfim 2010 d^n thfing 6 nfim 2011.
2.2. PhUcfng p h a p nghiSn cihi - Cac phupng phfip nudi cfi^y md hiSn hfinh da dUdc sii dung trong nghien cUu.
- Bd trf thi nghiem vfi cfic chi tieu theo doi, diin tra deu dUdc tien hfinh theo phUdng phap nghi&n cUu ndng sinh hpc thdng dyng.
Cac so' lieu dUpc xilt 1^ thong ke bfing phfin mem IRRISTART 5.0 vfi Excel.
Hinh 1. Hoa lan Dendrobium Jimbriatum Hook.
3. KET QUA VA THAO LUAN 3.1. Khijf trung qua lan
Nhieu cdng trinh nghien ciiu trong nUdc vfi tren th^ gidi da cdng bd ket qua nghien
Nguyin Thj San, Nguyin Thfiy Anh, Vu Nggc Lan, Trin ThS Mai cUu khcf trung mfiu qua cua cfic gidng lan
(Hofing Thi Nga, 2008; Pham Thi Kim Hanh vfi cdng sU, 2009). Ke thtta cfic kg't qua d6, qua lan dUdc kbit trdng theo 3 cdng thilc vfi cfi'y trgn nen mdi trUdng MS + 100ml ND + lOg sacaroza + 6g agar/lxt. Cfic thi nghidm khcl trung qua lan deu cho kg't qua td'i Uu, ty le mfiu so'ng vo trung dat 100%.
Ban dau hat lan co mfiu vang nhat vfi sau 6 tuan nudi cay trUdng len vfi chuygn sang mfiu xanh tUdng tu loai lan rUng Kim Diep (Dendrobium cbrytosotoxum) theo md ta ciia Nguyln Van Song (2011).
Ca 3 cdng thdc deu cho ket qua tdi Uu nhU nhau nhUng xet hieu qua, cac cdng doan thao tac thi CTl (dd't bfing cdn 96" trong 1 phiit) lfi tdt nhfi't. Sau 6 tufin nudi cSTy tren lofii lan nghign cUu thi 100% hat dUdc gieo tren moi trudng MS + 100ml ND + lOg sacaroza + 6g agar/iit mdi trUdng phat sinh protocorm.
3.2. Nhan n h a n h protocorm
Hat lan nay mfim co the san xufi't ra mot khoi te' bfio chUa phan hofi ro rfing dUpc gpi lfi protocorm. Tfi't ca cac protocorm nfiy se phat trig'n trong nhieu tufin, nhigu thang thfim chi trong nhieu nfim phu thudc tUng lofii cho dg^n khi du ldn d^ tao choi (McKendrick, 2000).
3.2.1. Anh hudng cua nin moi trUdng nuoi cSiy den khi nSng nhan nhanh protocorm.
Ti^n hfinh thi nghiem tren cfic nen mdi trudng ddng trong nuoi cfi'y md dUpc cho lfi thfch hdp vdi cfic lofii phong lan ndi chung nhU: MS (Murashige & Shoog, 1962), VW (Vacin & Went, 1949), KC (Knudson, 1965), RE (Robert Ernst, 1979).
Cfic cum protocorm thu dUpc b thi nghigm trgn dupc chpn lUa vfi cfi'y vfio mdi binh 03 cum protocorm cd dudng kinh 5mm vfi mdi cum mang 50 protocorm. Sau 8 tufin nuoi cfiy, theo ddi thu dUpc ket qua d bang 2.
Bang 1. Anh hUdng ciia phUtfng phfip khtf t r u n g qua den ty le m a u song va h i n h thfii protocorm (sau 6 t u a n )
Cdng thijc Ty 1$ m l u s6ng cd phat sinh
protocomi {%)
Htnh thdi protocorm C T l (Dot bang con 96" trong 1 p h i i t )
CT2 (C6n 96" 1 phut + H j O i (2%) 5 phut) CT3 (C6n 96" 1 phCt + HgCI; (0,1%) 5 phiit)
100
100 Ghi chii: +++ Kich thUdc protocorm dong deu, mau sfic xanh cua protocorm
Bang 2. Anh hifdng cua n^n moi trifdng nuoi caiy d^n k h a n a n g n h a n n h a n h protocorm (sau 8 t u a n nuoi cay)
Cdng thipc Diri/ng kinh cgm S6 l u ^ n g
(mm) protocorm/ci^m MSN protocorm
(l^n) C T l : MS + lOOmlND + lOg sacaroza + 6g agar/iit mOi tru^Tig
CT2: VW + lOOmlND ••- lOg sacaroza -*- 6g agar/lll mdi ttu'dng CT3: KC +10OmIND -•• 10g sacaroza + 6g agar/Iit mfii tru'dng CT4: RE + lOOmlND + lOg sacaroza + 6g agar/llt mdi trubng
LSDofls C V %
19.33 17,80 20,50 16,61
189,00 178,56 205,33 169,44
3,78 3,57 4,11 3,39 0,34 4,8
Nhan gi6ng in vitro loai lan Dendrobium fimbriatum Hook. (Hoang th^o long nV Moi trudng KC (CT3) cho tfing trUdng vg"
dudng kinh cym protocorm ldn nhfi't 20,5mm; sd lupng protocorm ldn nhfi't d^t 205,33 protocorm/cym vfi h^ sd nhfin (HSN) cao nhfi't 4,11 lfin sau 8 tufin nudi cfiy vd so' h^u. Xgt v^
HSN thi CT3 cung cho h§ sd nhfin cao nhfi't.
Tuy dudng kfnh cym protocorm, sd lUpng protocorm vfi HSN protocorm thu dUpc b CT3 sai khac cd ^ nghla vdi cac CT2 vfi CT4 nhUng sai khac khong cd ;^ nghia vdi CTl nhung xdt vd m$t hi^u qua kinh t^ thi CT3 moi trUdng KC lfi mdi trUdng re ti^n vfi thdng di^g trong nudi cfi'y lan, chinh vi thfi' mdi trUdng KC lfi mdi trUdng dupc li^a chpn trong nudi cfiy nhfin nhanh protocorm lan Dendrobium fimbriatum Hook.
3.2.2. Anh liudng cua djch nghien cii, qua den khd ndng nhdn nhanh protocorm.
Dich khoai tfiy nghig'n, dich nghiin qua chudi, dich nghig^n cfi rdt, dich nghien qua tao, ._. dupc sit dyng d^ lfim tfing sU phat
tri^n cua md seo hay cd quan nuoi cfi'y trong nudi cfi'y md phong lan (Ld Vfin Hofing, 2007). D^ bi^t dupc loai dich nghign nfio Ik cfin thi^t cho giai dopn nhan nhanh protocorm cua gidng lan Dendrobium fimbriatum Hook., thf nghigm d U ^ tie'n hfinh cfiy vfio mSi binh 3 cym protocorm cd dudng kfnh 5mm vfi moi cym mang 50 protocorm dUpc dua vfio nudi cSiy trdn ngn mdi trudng KC cd b^ sung cfic dich nghifin (khoai tfiy, cfi rdt, tfio, chudi). KS^t qua theo doi trong 8 tufin dupc th^ hign 6 bang 3.
Khi b^ simg riSng tifng lo^i dich nghien khoai tay, cfi rdt, tao, chuoi vdi c ^ g lUdng vfio mdi trudng thi thfi'y dich nghiin khoai tfiy (CT2) lfi hieu qua doi vdi protocorm lan Dendrobium Gmbriatum Hook, (dudng kinh cum protocorm lfi 20,87mm; sd lupng protocorm/cum lfi 231,41 vfi HSN lfi 4,63 lfin).
Bang 3. Anh htrdng cua dich nghien cu, q u a den k h a n a n g n h a n n h a n h protocorm (Sau 8 t u a n nuoi cay)
cum (mm)
S6 lu'p'ng protocorm/cgm
HSN protocorm
(lin) CT1(DC)
CT2. DC + 60g khoai tSy/lit mfii tru'&ng cn. EK; + 60g cd r6t/lil mOJ tnr6ng
Crr4: OC + 60g tSo/lit mfli t r u i n g CT5: BC + 60g chuAiflit mfii tnrfing CT6: DC + 60g khoai tdy + 60g cd r6t/llt mfii trufing CT7: £)C + 60g khoai tiy + 60g tio/llt mfii trirtrng CT8- DC -•• 60g khoai tdy + 60g chuAt/llt mfii bvArig CT9, DC + 60g c a r6t + 60g t&j/lil mfii t r u i n g CTIC- OC + eOg cd rfit + 60g chuiMIt mfii tnKmg CT11: OC + 60g tdo + eOg chii6t/llt mfii tn/fing
20,43 20,87 12,33 12,17 11,83 12,61 11,35 11,11 10,98 10,06 10,96
200.03 231.41 159.78 157.33 118.44 115,70 115,56 115,33 113,89 113.44 112.56
4.00 4.63 3.20 3,15 2,39 2.31 2.31 2.31 2.28 2.27 2.26
*,C
"
«*
• • , C . R
• + • + •
**,c
•"
• • , c
• • . C . R LSDooi
CV%
0,24 5,0 Ghi chii: +++, ++, +; gfic x a n h d^ra, x a n h vfi xanh n h ^ t cua mfiu
C: xufi't hi^n choi R' xufi't hipn re
DC = Moi trUcing KC + 100ml ND + lOg sacaroza + 6g agar/iit mdi trUdng
Nguyen ThjSan. NguySn ThlLfAnh, VQNggcLan, Trin ThSMai Ddi vdi cac cdng thUc cd bo' sung ket hpp
2 loai dich nghien vfio mdi trUdng nudi cfi'y (tii CT6 dg'n CTll), khdng cho kit qua vUpt trdi theo tinh cpng hpp, th|tm chf cdn cho cac chi tieu dudng kinh cum, so' lUpng protocorm vfi HSN thfi'p hdn so vdi cac cdng thUc bo sung rigng tUng loai dich nghiin vfi thfi'p hdn ddi chUng.
Nhu vfiy, cdng thUc CT2 (Moi trUdng KC + 100ml ND + lOg sacaroza + 60g khoai tfiy/lit moi trUdng) la cdng thUc cho he so' nhfin protocorm dat cao nhfi't 4,63 Ifin/sau 8 tufin nudi c5y. Nghien cUu nhfin nhanh lan Kim Tuyln Anoectochilus roxburghii (Wall.) Lindl. ciing cho so' HSN protocorm lfi cao nhfi't 5,33 lfin sau 8 tufin nudi cfi'y khi bd' sung 100 ml ND va lOOg khoai tay vao moi trudng KC (Phiing Vfin Phg, 2010).
3.3. Nhan n h a n h cum choi
3.3.1. Anh huang cua nen moi trudng nudi cdy den khd ndng nhdn nhanh cum choi
Ddi vdi lofii lan Dendrobium fimbriatum Hook, trong giai doan nhan nhanh protocorm thi nen mdi trudng KC la toi Uu.
Vfiy b giai doan nhan nhanh cum chdi ddi vdi lofii nay thich hdp vdi nen mdi trUdng nfio?
Chiing toi tiln hanh tren 4 cdng thUc.
Moi cdng thUc cfi'y 3 blnh, moi binh thi nghiem 3 cum chdi co dudng kfnh 10 mm va
cd chieu cao 10mm, mdi cum c6 chUa 15 chdi dUdc dua vfio nudi cfiy tai cfic cdng thUc thi nghipm. Kit qua th^ hifin qua bang 4.
Thfinh phfin cfic nguygn td khoang cua mdi trudng MS (CTl) rfi't phCi hpp cho qufi trinh tfing nhanh v l so lupng chdi (45,07 chdi/cym) vfi HSN chdi (3,0 lfin) trong nudi cfi'y cum chdi cua gidng lan Dendrobium fimbriatum Hook. Mdi trUdng KC (CT3) phii hpp cho nhfin nhanh protocorm lofii lan nfiy nhUng trong nhfin nhanh cum chdi lai khdng phii hdp (sd lupng chdi chi dat 41,22 chli/cym vfi HSN chdi lfi 2,75 lfin).
Mdi trUdng VW vfi RE ciing khong phU hdp trong nudi cfi'y lan Dendrobium fimbriatum Hook. Moi trudng VW (CT2) cd chi tieu sd chdi/cym lfi 39,67 va HSN lfi 2,64 lfin. Mdi trudng RE (CT4) cho sd chdi/cym (38,37 chdi) vfi HSN (2,56 lfin) la thfi'p nhfit sau 8 tuan nuoi cfi'y.
0 mUc dd tin cfiy 95% sd chdi/cym vfi HSN chdi b CTl sai khac cd Jf nghia so vdi cfic cdng thdc khac. Nhu vfiy, mdi trUdng nudi cfi'y dUdc lUa chpn trong nhan nhanh cym chdi lan Dendrobium fimbriatum Hook, lfi nen mdi trUdng MS. Khi nhfin gid'ng dia lan Hdng Hofing (Cymbidium iridioides) tac gia ciing da diing n i n la mdi trudng MS bo' sung 15% ND (Hofing Thi Nga, 2008).
Bang 4. Anh hifdng cua n e n moi trUdng nudi cay d^n k h a n a n g n h a n n h a n h c u m choi (sau 8 t u a n nudi caiy)
Cfing thtfc S6 c h i i / c y m HSN ch6i Hinh thai
( i j n ) cym chfii CT1: ruiS + 10OmIND + 10g sacaroza + 6g agar/Iit mfii toffi-ng
CT2: V W + lOOmlND + lOg sacaroza + 6g agar/iit mfii tm-firg CT3 KC + lOOmlND + 10g sacaroza + 6g agar/llt mfii tru'd'ng GT4 RE + lOOmlND + lOg sacaroza + 6g agar/llt mfii tru'fi'ng
45.07 39,67 41,22 38,37
3,00 2,64 2,75 2,56
• ++, b ++, c ++, b ++. c LSDoos
C V %
3,6 4,6
0,2 3.9
Ghi chii: ++: Sfic xanh cua cym chdi; b: Than tnmg binh; c: Than manh
Nhan gi6ng in vitro loai lan Dendmbium fimbriatum WOQV.. (Hoang thao l o n o ^ ^ .
3.3.2. Anh hirang cua d\ch nghien cii, qud den khd ndng nhdn nhanh cum chdi
Cfi'y vfio mdi binh thf nghidm 3 cym chdi cd dudng kfnh 10mm vfi c6 chilu cao 10mm, mfii cym c6 chUa 15 ch6i dUpc dUa vfio nudi cfi'y tren nin mdi trUdng MS (Murashige vfi Skoog, 1960) c6 bd' sung cfic djch nghidn (khoai tfiy, cfi nVl, tfio, chudi) qua dd xac dinh dupc anh hudng cua cac chfi't bd' sung nfiy de'n kha nfing nhfin nhanh cua cym chdi. Kit qua dupc th4 hi^n cl bang 5.
Kg't qua cho thfi'y d cfic cdng thUc b^
sung rieng I^ cac dich nghiin vfio mdi trUdng nudi cfi'y thi CT5 (60 gam chudi chfn/Ift mdi trudng) cho s6' ch6i/cum nhilu nhfi't (51,56 choi/cym) vfi HSN chdi cao nhfi't (dpt 3,44 lfin). Cfic cdng thiic cd b^ sung kit hpp cfic
cho kit qua vupt trdi t h e ' h cr hpp mi d cfic cdng thdc tu CT6-C ' ^•^' " ° ''^'^
tigu sd chdi/cym vfi HSf • •' ' 'P *^^" ^'^^•
DICU nfiy cho thfi'y vige kca hdp cfic chtftilu nhi<;n vfio ciing m6i tru. -; nhan chdi kh6n«
mang Iai hd so nhfin chd, -i ng cao nhU monf mud'n. d mUc dp tin cfiy '%, sd chfii/cpi thu dupc vfi HSN fl'- 'tpt duu' sau 8 tuin nudi cfi'y b cong tit • cao hdn so vdi cftc cong thUc khfic. Mn, rong MS + lOOmlND + 20g saccaroza + 6g agar + 60g chufi chfn/lit mdi trudng la tdi Uu cho nhfin nhaidi cym chdi lofii lan nghifin cijtu. Kit qui nghign ciiu tr3n lan Hfii hfliig (Paphiopedilum bangianum Gurss.) khi nhfin nhanh choi thi cfin bo sung lupng chu$
cao hdn lfin d i n lOOg chudi/lit mdi trudi^
(Hofing Thi Giang, 2010).
dich nghien vfio mdi trUdng nudi cfi'y khdng
Bang 5. Anh hi/ctng ciia dich n g h i e n cii, q u a dl^n k h a nfing nhan nhanh cum choi (sau 8 t u a n nuoi cay)
Cfing thi>c S6 chii/cym HSN chAi (i^n) " ' m j H ^
o n (DC)
CT2: DC + 60g khoai lay/llt mfii trirfi'ng CT3 DC + 60g cd rit/llt mfii triri/ng CT4 OC + 60g tdo/lil mfii tririmg CT5. DC + 60g chuAl/llt mfii trirfitig CT6:00 + 60g khoai tay + 60g cd r6t/iit mfii tnrfrng CT7: DC + 60g khoai tfiy + 6O9 tSo/Ht mfii tn/fi^ig CT8 OC + 60g khoai tfly + 60g chuAi/lIt mfii tnibng CT9 DC + 60g cd rit + 60g tio/l(t mfii tn^&ng CT10; DC + 60g cd r6t + 60g chuii/llt mfii tn/frng CTl r OC + 60g tfio + 60g chuii/llt mfii truing
46.78 47,26 42.93 46.89 51,56 43,11 43.37 44.70 47,04 47^2.
47,56
3,12 3.15 2.86 3.13 3.44 2.87 2.89 2,98 3,14 3.1&
3,t7
*. c +. c
* * , c ++.b ++, a ++, b ++,b ++, b ++.b ++.b ++, b LSDow
CV%
3.43 4,8
0,24 4,6 Ghi chii: ++, +: Sfic xanh vfla, xanh nh^t ciia cym chdi
a: Than mfip; b: Than trung binh; c; Than manh
DC = Moi trUcing MS + 100ml ND+ 20g saccaroza + 6g agar/Kt mdi trUcing
Nguyin ThjSan, Nguyin Thi Lf Anh, VQNggcLan, Trin Thi Mai
j3.4. Tao cay h o a n c h i n h
3.4.1. Anh huang ciia nen mdi trudng nudi cdy iden sinh truang cua chdi
Day la giai doan cudi ciing cua qufi trinh [nhan nhanh in vitro. Vdi myc dich tao cay con cd sUc so'ng cao, dat tieu chufin ra cfiy.
cac mdi trUdng tren da dUdc thit nghigm cho sU sinh trudng phat trid'n cua cay lan in vitro
\ f i tiln hanh Ifiy choi thu dupc tii cac thf nghigm tren tach rigng re cfi'y tren 4 cong thiic, mdi cdng thUc cfiy 3 binh, mdi binh cfiy 3 chdi, moi chdi cd chieu cao 3cm, 3 la. Ke't qua dUdc the hign qua bang 6.
Kit qua bang 6 cho thfiy: 6 CT4 sii dung nin khoang RE cho kha nang phat triln ve chieu cao cfiy, so' la la cao nhfit (dat chieu cao 5,22 cm/cay, 5,41 Ifi/cfiy) vfi chat lupng cfiy tdt.
O dp sai khfic tin cay 95%, cong thUc 4 cho sinh trudng cfiy tdt nha't hdn h^n cac cdng thdc khac. Nin moi trudng nudi cfi'y phli hdp nhfi't cho sinh trudng cua cfiy lan
Dendrobium fimbriatum Hook, lfi nin mdi trudng RE.
3.4.2. Anh huang cua than hogt tinh (THT) din khd ndng ra re cua chdi
Bd' sung THT vao mdi trUdng nudi cfi'y, d giai dopn tao cay hofin chinh rfit cd hieu qua vdi cac lofii phong lan. THT c6 the lfim chfim qufi trinh hfi'p phy mpt sd chfi't dilu tig't sinh trUclng, dinh dudng cfin thilt khac vfi thiic dfiy tao cay hofin chinh. Ngofii ra THT khi dUdc bd' sung vfio moi trUdng cdn cd tac dung tao do xdp vfi tao dd toi cho moi trudng rfi't phii hpp cho qua trinh ra re ciia mfiu ch6i cfiy. De nghien cUu anh hudng cua THT din kha nfing ra r l cua chdi lan Dendrobium fimbriatum Hook. Thi nghiem dupc tien hfinh theo 5 cdng thUc b^ sung hfim lupng THT khfic nhau vfio mdi trUdng RE. Mdi cdng thUc cfi'y 3 binh, mSi binh cfi'y 3 chdi cay, mfli chdi cfiy cd chilu cao 3cm; 3 la dupc cfi'y vfio cac cong thiic thi nghiem.
Sau 30 ngfiy nudi cfi'y dfi thu dUdc ke't qua d bang 7.
Bang 6. Anh h u d n g ciia n e n moi trUcfng nuoi ca'y d e n sinh truding cua choi (Sau 8 t u a n nuoi ca'y)
Cfing thirc Chieu cao cdy (cm) S6 ia/cay C h i t l i i y n g cdy _ CT1 MS + lOg sacaroza + 6g agar/llt moi trifgng
CT2. V W + lOg sacaroza + 6g agar/Iit mfii tri/frng CT3: KC + 10g sacaroza + 6g agar/Iit mfii tru'd'ng CT4: RE + lOg sacaroza * 6g agar/Iit mfii tru'fi'ng
4,02 3,11 3,30 5,22
3,89 5.41 LSOow
C V %
Ghi chii: +++ Tot (xanh, map, c6 3-5 rl)
Bang 7. Anh hifdng cua THT den k h a n a n g r a re cua choi. (Sau 30 ngay nuoi ca'y)
Ty Ig tgo r§ (%)
Sau 10 ngdy Sau 20 ngdy Sau 30 "S6rfi Chi^u d^i r§
(cm) C T l (DC)
CT2: DC -t CT3. O C ••
CT4: OC + CT5 DC 4
0,5g THT/lit mfii tnr&ng I.OgTHT/IItmfiitnrfi-ng 1,5g THT/lit mfii tru'fi'ng 2,0g THT/llt mfii trirdng
7,40 18,51 48.14 55.56 62,96
81.48 100,00 100,00 100,00 100,00
1.15 2.37 5,67 3,33 3,48
2 , 8 7 3,32 3,66 LSD(,,Ds
C V %
Ghi chii: DC = Mdi trUctng RE + lOg sacaroza + 6g agar/iit mfii tmdng
Nhan gi6ng in vitro loai lan Dendrobium fimbriatum Hook. (Hoang thgc long m
Hinh 2. Anh hu^ng cuii THT Kg't qua bang 7 cho thfi'y tfi't ca cac cdng thUc cd bo' sung THT sau 30 ngfiy nu6i cfi'y cho ty le choi tao r l 100%. CT3 cd bd' sung lg THT/lft mdi trudng nudi cfi'y cho sd r l nhilu nhfi't la 5,67. Tuy nhien, tren mdi trUdng c6 bd sung THT nhilu hdn l,0g/lft mdi trUdng chieu cao cay thfi'p dfin, dilu nfiy c6 th^ do THT dfi hfi'p phy mpt so' chfi't dieu tiet sinh trudng vfi dinh dudng cfin thilit ngn cay sinh trudng chfim.
Vfiy bo sung vfio mdi trudng nuoi cfi'y choi lan Dendrobium fimbriatum Hook. l.Og THT/h't mdi trUdng nudi cfiy lfi toi Uu cho vige t^o re in vitro.
4. KET LUAN
- Ngudn vfit ligu ban dfiu lfi qua lan, c h l dp khli triing toi Uu lfi nhiing qua trong cdn 96° rdi hp trgn ngpn liia dgn cdn vfi cfi'y tren nen mdi trudng MS + 100ml ND + lOg sacaroza + 6g agar/lft mdi trudng
Mdi trudng tm Uu nhan nhanh protocorm lfi nin moi trUdng KC + 100ml ND + lOg sacaroza + 60g khoai tay/lit mdi trudng d^t hg sd nhfin 4,63 Ifin/sau Stufin nudi c^y.
d^n khfi nSng ra rl cua ch6i
- Mdi trUdng toi Uu nhan nhanh cum chdi lfi n i n mdi trUdng MS + 100ml ND + 20g sacaroza + 90g chuoi chfn/h't mdi trUdng dat he sdnhan 3,44 Ifin/sau 8 tufin nudi cfiy.
- Mdi trUdng nudi cfi^y toi Uu cho vi^c tai sinh tao cay hofin chinh ciia lofii lan Dendrobium fimbriatum Hook, la nen mfii trudng RE + lOg sacaroza + l,0g THT + 6g agar/Ht mdi trUdng.
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