KHOA HOC CdNG NGHE
IVGHIEIM CLUJ CAU TAQ GIAI P H A U R £ VA XAC DIIVH Vf TRt, DO DAI HOM R £ PHLJC VLJ IMHAIXi GIOIVG VO TilVH
DAT GIAC LIE[\I [Dysasma tonkinense ( G a g n e p . ) IVI. H i r o e ) ( D e r b e r i d a c e a e )
Phgm Ngpc KhinhS Ph^rn Tiianh Huy^S Ninh Tlii Phip^, Nghifem Tiki ChungS Trinh Tlii Nga^
T6MTAT
Bat giac lien {Dysosma tonkmense (Gagnep.) M. Hiroe) (Berbendaceae) la cay thuoc qu^, co phan bo chii yeu o Viet Nam va phia Nam cua Tmng Qudc. Duoc sit dung trong y hoc co truyen de chiia ran doc can, img nhot; nhieu nghien cuu gan day cho thay podophyllotoxin chiet xuat tir than rk va r^ cOia cac loai thu6c chi Dysosma Woodson, con duac sii dung lam thuoc chira ung thu. Nghien cmi vi phlu rh va su hinh thinh ch6i bat dinh tir rk Bat giac lien cho thay rh gom cac phan bieu bi, vo (rao mem v6), noi bi, tru bi va mo m^m mot; choi bat dinh duoc hinh thinh tir lop t^ bio tang phat sinh cua ri (tru bi). Khi sij dung r^ Bat giac Hen lira v&t lieu nhan giong, dimg hom dau rh cho ket qua nhan gidng tdt nhit voi thoi gian giam hom 310 ngiy, ^ 16 hom bit mam la 80,6%. Dong thoi, kich thitac hom rl tir 1 cm den 15 cm vi sir dung ca rl (dii > 30 cm) lim hom giam d^u co kha nang hinh thanh choi bat dinh, trong do hom rh dai > 30 cm co ty 16 hom xuat vuon cao nhat (94,7%).
Tirkhda: Bat giac li€n, vipbiu, choi bat dinh, nhan gidng vo tinh.
I.DATViWK
Chi Bat giac lien {Dysosma Woodson) la mot chi nho g6m 7 den 10 loai cay than thao song lau nam thuoc hp Hoang lien (Berbendaceae), co phan bo b Trung Quoc va Viet Nam; trong do co 6 loai d^c huu b Trung Quoc. Chi Bat giac Uen co quan h$ gin voi chi Diphylleia Michx phan bo a Tay A, Bac My; chi Podophyllum L phan bo a Tay Bic My va chi Sinopodophyllum Ying. phan bo a phia Tay day Himalaya.
Lx)ai Bat giac lien b Viet Nam co ten khoa hoc Dysosma tonkinense (Gagnep.) M. Hiroe [11, [2] voi cac ten dong danh Podophyllum tonkinense Gagnep. [3]; P. versipelle Hance subsp. versipelle, Dysosma difformis [4], Bat giac lien co phan bo tai mpt sd tinh thupc mien nui phia Bic Vi^t Nam nhu Hoa Binh, Son La, Lai Chau, Dien Bi&n, Lao Cai, Ha Giang, Tuyen Quang, Yen Bai b dp cao 470 den 1.720 m [3], [5], [6]; Kon Tum la dia diem phan bd men dupc ghi nhan va la diem phan bd xa nhat ve phia Nam cua loai nay [6]. Than rk va rk cua loai dupc su
dung trong y hpc co truydn voi ten vi thudc la "Bat giacli&n"[5).[6].m.
Do bl khai thac sir dung lam thuoc thi^u su ki^m soat trong nhieu n i m din den hien trang suy giam nguon gen nghidm trong trong tu nhien. Loai Bat giac lidn da dupc dua vao Sach Do Vi6t Nam (2007) voi miic phan hang "dang nguy cap" (EN Ala, c, d), cin ducfc nghien cuu nhan gidng va bao tdn.
Giam hom la mot trong nhung bien phap nhan giong vo tinh dupc ap dung de nhan giong cay trong.
Trong do, hom rk cua mot so loai cay cung duoc sii dung lam vat Ueu nhan gidng. O Viet Nam, nhieu loai cay dupc ap dung bien phap giam hom rk theo kinh nghidm truyen thong de nhan gidng nhu cac loai c^y an qua (man, hong) hay cay thuoc (d6 trpng, dan sam). Ngoai ra, nhi^u loai cay cung dupc nhan giong bing bien phap giam hom rk nhu Clusia flummensis [8], Detarium microcarpum [91, Vitexdoniana [10]
Cay Bat giac lien co ty le hinh thanh qua va qua
' Vien Duac lieu
Email; [email protected] - Hgc vien Nong nghiep Viet Nam
chin rat thap trong tu nhien cung nhu dieu kien trong trpt nen viec sii dung hat lam vat lieu nhan gidng rat kho khan. Cay co the nhan giong bing than rk nhu cac loai thupc chi Podophyllum L Z>Ksosma Woodson. [11]. Chua co nghien <^v„ i,-
• 1 •-*i. - I.- n. ,^.^'"^" cuu nhan giong cac loai thupc cm Dysosma \\ oodson hs = NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG 7/2020
KHOA HOC CONG N G H £
tuy nhi^n P h a m N g p c K h a n h v a c s d a g h i n h a n c a c truong h o p c a y B a t g i a c li6n m p c t u t h i n rk [12]. Voi myc tiku n g h i e n c i i u v a t l i e u va b i e n p h a p n h a n gidng phii h p p c h o c a y B a t g i a c lien, d a t i e n h a n h nghidn c i m c^u t a o g i a i p h i u rk, d a c d i ^ m h i n h t h a n h chdi bilt d i n h tir rk v a c a c b i e n p h a p k y t h u a t n h a n gidng v6 t i n h B a t g i a c lien b i n g h o m rk.
2. VAT UEU VA PHUDNG PHAP NBHei CUU 2.1. Vit li^u nghidn cmi
Ciy Bat U6n trdng tai Tram Nghifin cuu Trong cay thudc Sa Pa, Vien Dupc lieu. Cay 3-4 tuoi, trong trong nha mang, co ludi den che giam anh sang. Rk cay duoc thu thap vao thing 10 khi phin than tren mat bit dau lui.
2.2. Phuong phip nghidn ohi
2.2.1. Phuongphap cdt miu va nhudm kep va soi tiiu ban
Vi phiu rk duoc thuc hien tai Khoa Hoa phan tich Ti^u chuan, Vien Dupc U^u.
Miu rk sau khi thu ve dupc rua sach bing nuoc sau do dupc co dinh va bao quan trong dung dich FAC gdm ethyl alcohol 96°, acetic acid 40%, formalin va nuoc cat (Nguyin Khoa Lan, 1997). Miu rk dupc cit mong bang dao lam, nhupm kep va lam tieu ban tam thdi. Phuong phap nhupm kep nhim phan biet duoc te bio co mang bang cellulose (bit mau do cua thudc nhupm carmine va te bao cd mang tham lignin (bit miu xanh cua thudc nhupm methylene blue) (Hoang Thi San va Nguyen Te Chinh, 1982). Tir do xac dinh diroc su co mat va phan bo cac loai md cua r^. Miu rk dirpc quan sat va chup anh tieu ban dudi kinh hi^n vi Kruss MBL 2000 - T cua Due.
2.2.2. Phuongphap nhin gidng vo tihh Thi nghiem nhan gidng vo tinh Bat giac Uen bing hom r^ dupc thuc hien tai Tram Nghien cuu Trdng cay thuoc Sa Pa, Vien Dupc lieu.
Thi nghiem 1: Anh hudng cua vi tii hom ri din kha n^g nhan gidng Bat giac lien.
Thi nghiem gdm 3 vi tri hom rd khac nhau: hom dau vk (Vri), hom giua rk (Vr2) va hom cudi rd (VT3). Cic hom rd dong deu, dai 10 cm, dudng kinh
~ 0,2 cm.
Thi nghidm 2: Anh hudng cua kich thudc hom ri den kha nang nhan gidng Bat giac hen.
Thi nghiem gom 5 kich thudc hom khac nhau:
hom dai 1 cm (KTl), hom dai 3 cm (KT2). hom dai 5
cm (KT3), hom dai 10 cm (KT4), hom dai 15 cm (KTS) va hom la ca mdt doan rk dii > 30 cm Q^IQ).
Cac hom rk dupc lay tir vi tri giua rd, dong d6u \k dudng kinh (~ 0,2 cm)
Cach thuc hidn: Rd sau khi thu tir cay me dupc nia sach, sau do cit thanh cac doan hom va xii ly theo cac cdng thuc thi nghidm. Cac hom rk dupc giam trong gia the l i mun nui; khi giam dit hom nim ngang cich nhau 3 4 cm sau dd phu kin bing \bp miin day 2-3 cm; tudi giu dm cho den khi hom bat mim va cay dat tieu chuan xuat vud^.
Xii ly sd Ueu tren chuong trinh Excel.
3. KET QUA NGHDU CUU VA THAU LUAN
3.1. D$c di^m hmh thii, vi phSu rk Bit giic Udn
Hinh 1. Die di^m hinh thai v i vi phAu rfi Bit giic uen
a, b- than ri mang cac ri; c- ri Bat giac Uen. cl:
chop ri. c2: vimg ri trudng thanh mang long ri; d- mat cdt ngang tai vimg ri trudng thanh. dl. bieu bi, d2-m6 mim vo, d3-md cdng, d4~m6 mim ruot, dS-gd, d&hbe.
Bat giic lien la cay hai la mam nhung cd kieu rd chum gom nhieu rk nho mpc tir thin r^. Rd hinh spi, dai 30 - 70 cm, dudng kinh 1,5 - 2,5 mm; mat ngoii cd nhidu Idng rk, mau vang chanh. khi gia co miu niu nhat Mai cit ngang rd hinh trdn. Bieu bi gdm mot lap te bao da giac hay hinh gin trdn, xep tuong ddi ddu dan. Dudi lop bieu bi la md mdm gdm cac te bao trdn CO thanh mdng. Rai rac trong md mdm vd cd te bao cung kich thudc to hon te bio md mem. Gd va 125
KHOA HOC CONG NGHE
libe tao thanh vong tron ngay lop tru bi. Phia trong la mo mem nipt co kich thuoc nho. va co thanh day (Hinhl).
3.2. Sir hinh thanh ch6i bit dinh trta rJ Bat giic U6n
Hinh 2. Vi phiu rk Bit giac lifin dang binh thanh m ^ b^t dinh
a, c- vi phiu va vi tri ri dang trong qua tiinh phan hoa; b, d. hinh thai va viphiu mam bat dinh.
Phan phan hda cua te bao la qui trinh ma te bao da phin hda trd' lai thanh te hao goc (callus), cd tinh toin nang [13]. Vi phiu vi tri rd dang trong qua trinh hinh thanh mam bat dinh cho thay: tai vi tri rd hinh thinh chdi bat dinh, Idp td bio cua tang phit sinh rk
phan phin hda, phan chia manh tao thinh mdt khdi md gdm nhidu te bao nhd, cac te bao tiep tuc phin chia va tang kich thuoc (theo ca chieu dai va chieu rpng) lam cho khdi md ldn len, rich lop bieu bi, vet rich dai tir 6,0 -12,0 mm (Hinh 2a, 2c). Sau khi lop bieu bi rich ra, mim bat dinh mdi hinh thanh; mim bat dinh gdm 3-4 l i bao chdi hinh tam giac, mau ving nhat hay miu tring (hinh 2b, 2d); kich thudc cua chdi bat dinh phu thudc vao dp siu cua rd trong dat, neu rk mpc sau trong dat, chdi cd the hinh thanh va vuon dii den hon 5 cm. Khi mam vuon ldn khoi mit dit thi la that mdi xuat hien.
3.3. Kha ning nhin gidng Bit giic USn bing r^
3.3.1. Anh hudng cua w tri hom ri den kha nang nhan gidng Bat giac Uen
Ddi vdi cac loai thuc vit khac nhau, viec su dung vi tri hom khic nhau (tuong ung vdi tudi hom) trdn than hoac rd cd anh hudng ddn ket qua nhan gidng. Trong khi mdt sd loai thuc vit cd the su dung hom ngpn (hom non) lam vit lieu nhin gidng nhu cac loai thudc chi Tu ciu {Hydrangea L), thi da sd cic loii thuc vit khac nen sir dung hom giiia (hom binh te) hoic hom gdc (hom gia) lam vat Ueu nhin gidng. Khi nhan gidng loai Vitex donianna bing rd, hom cd dudng kinh 1,1-2,5 cm se cho ket qua nhan gidng tdt hon so vdi hom cd dudng kinh 0,5-1,0 cm [10].
^ \ ^ Chi tieu Cong thirc ^ \ ^
VTl V K
vn
Thoi gian tir khi giam hom dfin.... (ngay) Bat mam
275 265 260
Xuat vudn 325 310 310
Ty le hom bat mam (%)
67.7 74,8 80.6
Ty le cay xuat vuon (%)
62.6 72.3 78.9 Ghichu: VTl - Hom cudi ri: VT2 - Horn giua ri: VT3 - Horn diu ri
Bang 2. Bgc di^m cua cay gidng Bit giic lita tli cic vi tri hom re Chieu cao cay
(cm) 8.0 + 0.9
Sd la fla) 1.2+0.5
Sd rh (r§) So nhanh (nhanh) 1,0 1.0 Ghi chu: VTl - Hom cudirS: VT2 - Hom giua ri: VT3 - Hom dau ri
KHOA HOC CONG NGHt Khi nhan gidng Bat giac lita bang hom rS cho
thay hom d vi tri dau ri - "VT3 (hay hom gia. giap than r4) cd kha nang nhan gidng tdt hon. Hom bat mam sau 260 ngay va dat tieu chuan xuat vudn sau 310 ngay voi ty le hom xuat vudn la 78,9%. Ddi vdi hom cudi r i - W l (hom ri non) cd thdi gian bit mam va dat Heu chuan xuat vudn lau hon (sau 275 va 325 ngay), ddng thdi, hom cd ty le bat mam va ty le dat tiSu chuan xuat vuwn (67,7% va 62.6%) thap hon hom dau rd va hom giua r§ (Bang 1).
Ciy gidng Bat giic lien tu hom tai vi tri dau rd cd chieu cao cay cao hon. sd la va sd rd nhidu hon so vdi cay gidng tir hom giua rd va cudi rd. Tuy nhien. sd
nhinh (sd mam) h-ta mdi hom d vj hn cudi rd lai nhieu hon hom giua rd va cudi rd (Bang 2).
3.3.2. Anh hudng cua kich thudc hom rd den kha nang nhan gidng Bat giac hen
Chidu dai hay kich thudc cua doan hom dem giam cd kita quan tdi chat dinh duong du tru trong hom de cung cap cho qua hinh bat mam. Khi nhan gidng ciy Detanum microcarpum bing phuong phip giim hom rd. doan hom cd kich thudc dii (20 cm) cd kha ning nhin gidng tdt hon so vdi doan hom ngin (10 cm) [131.
Bing 3. Anh hudng cda vi tri hom rf ddn kha nantr nhan ^^«
^ \ Chi tita
Cong thuc ^ ^ KTl KT2 KT3 KT4 KTS KT6
Thdi gian hi khi giam hom den.... (ngiy) Bit mam
290 285 270 265 258 250
Xuat vudn 350 340 320 305 295 295
Ty le hom bit mam (%)
51.1 55.6 63.3 75.7 96.7 97.8
Ty le cay xuat vudn (96)
45.5 50,8 61.7 73.1 93.7 94.7
Ghi chu: KTl - Hom ri dai 1 cm; KT2 - Hom ri kich thudc 3 cm; KT3 - Hom ri dai 5 cm; KT4 - Hom ri dai 10 cm; KTS - Hom ri dai 15 cm. KT6 - Dung ca ri lam hom giam (ri dai >30 cm)
Bang 4. Bac didm cua ciy gidng Bit giic Udn tii cic vi tri hom rd
^~"~~--,^^^ Chi tita Cong thuc ^""~--^^^
KTl KT2 KTS KT4 KT5 KT6
Chidu cao cay (cm) 6.0+1.3 7.5+ 1.5 8.5 + 1.5 10.1 + 1.4 14.7 + 1.2 16.5 + 1.5
Sd la Oi) 1.1 ±0.5 1.5 + 0.3 1.5 ±0.4 1.7 ±0.5 2.1+0.5 2.5 ± 0.7
Sd rd (rd) 2.5 ± 0.6 2.5 + 0.4 3.5 + 0.5 3.7 ±0.3 5.5 + 0.6 6.3 ± 0.3
Sd nhinh (nhinh) 1.0 1.0 1.0 1.0 1.0-2.0 1.0-2.0 Ghi chu: KTl - Hom ri dii 1 cm: KT2 - Hom rd kich tiiudc 3 cm: KT3 - Hom ri dai 5 cm: KTl - Hom ri dai 10 cm: KTS-Hom ri dai 15 cm. KT6 - Dting ca ri lamhomgiam (ridai>30cm)
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG 7/2020 127
KHOA HOC C O N G N G H t
Dd dii cua hom rd cd inh hudng ro ret ddn k h i ning nhan gidng Bit giic Uta. Kdt qua theo doi su b i t m i m ciia hom rd cho thay ring: cic hom cd kich thudc hi 1 cm (KTl) den 16 cm (KTS) hay su dung c i r l lam hom giim (KT6) ddu cd kha nang b i t mam sau thdi gian giim hom til 250 den 290 ngiy v i hinh thanh cay con hoan chinh. dat tita chuan xuat vudn sau 295 ddn 350 ngiy; vdi ty le bit mam 51.1% den 97.8% v i ty le cay d^t tita chuan xuat vudn 45,5% ddn 94.7%. Hom rd cing dii thi thdi gian b i t mam cing sdm va ty 16 ciy xuat vudn cing cao; trong dd khi su dung ca rd lam vit heu giam hom (rd dii >30 cm) cd thdi gian bat mam sdm nhat (sau 250 ngiy) v i ty le ciy xuit vudn cao nhit (94.7%) (Bang 3).
Hom rd Bit giic Uta cd kich thudc dii kh6ng nhiing k h i nang bat mam tdt hon ma dac diem hinh thii ciing vuot trgi so vdi hom cd kich thudc ngin.
Theo doi die diem ciy gidng Bit giic hta tir hom rd cd kich thudc khic nhau cho thay: cay cd chieu cao tir 6.0 ± 1.3 cm den 16.5 ± 1,5 cm; ciy con cd 1.1 ± 0.5 la den 2.5 + 0,7 la v i 2.5 ± 0.6 rd ddn 6.3 ± 0.3 rd. Cac hom cd kich thudc 1 cm den 10 cm cd 1 nhinh; trong khi hom dai 15 cm v i hom l i rd dii >30 cm cd tir 1 - 2 nhinh (Bing 4).
4 . KET LUAN
Rd Bit giic Udn hinh soi. dai 30 - 70 cm, dudng kinh 1,5 - 2,5 mm; mat ngoai cd nhidu long rd, miu ving chanh, khi gii cd miu niu nhat. Mat cit ngang rd hinh trdn gdm cic phin bieu bi. vd (mo mdm vd).
ndi bi. tru bi v i md mdm m d t Mam bat dmh duoc hinh thanh tir Idp te bio cua ting phit smh rd (tru bi). Hom duoc cit d cic vi tri rd khic nhau cd inh hudng ddn kha nang bit mam. frong dd hom duoc c i t d vi tri dau rd cd ty le b i t mim tdt nhat. Ddng thdi, dd dai hora rd khic nhau cd inh hudng ro ret den kha nang bat mam. hom la rd dii > 30 cm cd ty le b i t m i m tdt nhat Tuy nhita do vat lita nhan gidng l i rd Cling duoc sir diuig lim duoc Ueu cho nSn sir dung hom rd dai 15 cm de lim vit Uta nhan gidng la phu hop nhat.
TAJ UHJ THAM KHAO
1. Hiroe M., 1973. The plants of Basho's and Buson's Hokku Utera-ture: The Plants of Buson's Collected Haiku. - Ariake 8 (3): 328.
2 Pham Ngoc Khanh. Pham Thanh Huyen, Nguyen Quynh Nga, Aaron Polden, Ninh Thi Phip, Phan Ke Long and Tran Thi Viet TTianh. 2020.
Characterisation of Dysosma tonkmense (Gagnep.) M. Hiroe based on morphological characteristics and ITS sequence. Nordic Joumal Botany Vol 38(1). PP 1- 9.
3. Gagnepain. F., 1938. PodophyUumL hi:
Lecomte. P. H. (ed.). Flore Generale de ITndo-Chme.
Museum National d'Histoire Nahirelle, 1. pp. 146- 147.
4. Ying. J. etal., 2011. DysosmaWoodson. • In:
Wu. Z. Y. etal. (eds). Flora of Chma. Science Press.
Beijmg and Missouri Botanical Garden I>ress. 19. pp.
783-784.
5. Ho. P. H.. 1999. An iUushated flora of Vietnam. - Tre PubUshmg House 1. p. 327. m Vietnamese.
6. Hien. N. H.. 2005. PodophyUumL. - hi: Ban. N.
T. et al. (eds), CheckUst of plants in Vietnam.
Agriculture PubUshing House 2, pp. 162.
7. Loi, D. T , 2004. Medicinal plants and materials medicinal of Viemam. Medical PubUshing House, p. 544.
8. Nayagam J. R and T. K Simon.. 2017. Rooting of root cuttings: a breakthrough in vegetatative propagahon. Int.!. Adv. Res. Vol 5 (11). pp 911-916.
9. Dembele K. C . M. Tygabu. J. Bayala. P.
Savadogo. I. J. Buossim and P. C Oden. 2010. Clona propagation of Detarium microcarpum from root cutting. Silva Fennica Vol 44(5). pp 775-786
10. Mapongmetsem P. M.. G. Fawa. J. B.
Noubissie-Tchiagam. B. A. Nkongmeneck. S. S. H.
Biaou and Ronald BeUefontaine. 2016. Vegetative propagation of Vitex doniana Sweet from root segments cuttings. Bois et forets des U-opiques. Vol 2016 (327). 30-37.
11. Nadeem M.. L M. S. PaUii. A. N, Purohit H.
Pandey and S. K. Nandi. 2000. Propagation and conservation of PodophyUum hexandnim Royle- an important medicinal herb. Biological Conservation Vol 92 (2000). pp. 121-129.
12. Pham Ngoc Khinh. 2017. Danh gia dac diem ndng sinh hpc vi kha nang smh trudng. phit tridn cua ciy Bit giic Uta tai Sa Pa. Bao cao nhiem vu thudng xuyen. Vien Duoc Ueu.
13. Wu. J.. 2011. Many facets of
Moi. CeU Biol.3.75. stem ceUs. J.
NONG NGHIEP VA PHAT TRIEN NONG THON - KY 2 - THANG 7/2020
KHOA HOC CdNG N G H ^
STUDY ON ROOT ANATOMY AND DETERMINE THE PAKTS AND LENGTHS OF ROOT SEGMENTS FOR VEGETATIVE PROPAGATION OF Dysosma tonkinense (Gagnep.) M. Hiroe (Berbendaceae)
Pham Ngoc KhanhS Pham Thanh Huyen^
Ninh Thi Phip^, Nghiem Tien Chung', Trinh Thi Nga'
^NationalInstitute olMedicinal Materials,
^Vietnam National University ofAgricultiiral Summary
Dysosma tonkmense (Gagnep.) M. Hiroe (Berbendaceae) is a precious medicinal plant witii most of its disUibution occumng witlim Vietnam and Soutliem area of China. The dried roots and rhizomes of D.tonkinense have been used m forit medicine for the treatment poisonous snake bite, ulcer; recently studies reported that podophyllotoxin content in roots and rhizomes of all species of [>ysosma Woodson genus have been used for treated cancer. Study on root anatomy and arising adventitious shoot of root showed that the root of D. tonkinense include epidermis, cortex (parenchyma), endodermis, pericycle/cambium and pith (parenchyma); adventiUous shoots onginated to layer of cells of root pencycle/cambium. In vegetative propagation of D. tonkinense, old parts of root has sprouUng rate higher than middle parts and young parts of root (with sproutmg is 80.6 percents). Both root segments cutUng (1 cm to 15 cm length) and whole root (> 30 cm length) have abilities to sprouted with highest sproutmg in whole root (94.7 percents).
Keywords: Dysosma tonkinensis, anatomy, adventitious shoot, vegetable propagation.
Ngudi phan bi^n: PGS.TS. La T u ^ Nghia Ngiy nhan bai: 28/02/2020
Ngiy Ihdng qua phan bito: 01/4/2020 Ngiy duyet dang: 8/4/2020
