全 文 ：第 25卷　第 2期　　　　　　　　　　　　　植　　　物　　　研　　　究 2005年　4月
Vo .l 25　No. 2　　　　　　　　　　　　BULLETIN OF BOTAN ICAL RESEARCH April, 　2005
Foundat ion item:Supported by the S tate Key Basic Research and Developm ent P lan of Ch ina (G19990160).
*　Au thor in troduction:Zu Yuangang (1954— ), Ph. D. , Professor, m ajor in p lan t science.
Received date:2004 - 03 - 07
酸性培养基对喜树叶片细胞壁降解的影响
祖元刚*　于景华　唐中华　张宇亮　李艳华　孟庆焕
(东北林业大学森林植物生态学教育部重点实验室 , 哈尔滨　150040)
摘　要　在组织培养过程中对喜树叶片外植体进行解剖学观察。发现在组织培养条件下 ,喜树叶
片在培养基的酸性环境中细胞壁呈现模糊微弱降解 、明显降解和完全降解直至消失的解剖学特
征 。在同样的组培条件和相同的时间内 ,同一喜树叶片不同部位出现细胞壁程度不同的降解和消
失现象 ,可能是喜树叶片因上表皮凸凹不平进而导致其不同部位与酸性培养基接触的程度不同 ,
因而使培养基中的酸性物质对喜树叶片上表皮的不同部位影响出现异质化。本文对培养基中的
酸性成分对喜树叶片细胞壁降解的影响有了进一步的认识和理解 。
关键词　组织培养;细胞壁;酸性降解
Influence of ac id culture m ed ium on ce llwa ll degradation of
Camptotheca acum ina ta leaves
ZU Yuan-G ang*　YU Jing-Hua　TANG Zhong-Hua　ZHANG Yu-Liang　LI Yan-Hua　MENG Q ing-Huan
(Key Labo ra to ry o f Forest P lan t Eco lo ry, M inistry o f Educa tion, Northeast Fore stry University, H arbin　150040)
Abstract　During the course of plant tissue culture, Camptotheca acum ina ta leaf exp lants w ere ob-
served on anatomy charac te ristics. W e found that ce llw a ll of C. acum inata leaves presented ana tomy
charac te ristic of w eakly deg rading, obv iously degrading, to tally degrading and ce ll wa ll disappea rance
unde r tissue cu lture conditions. Ce llw a ll of identica lC. acum inata leaf exh ib ited distinct properties in
their degree of degradation and even disappearing at the same condition of tissue culture. Itmay be the
upper epiderm is protruding to influence diffe ren t contac ts position w ith cu lturemedium andmake acidma-
teria l o f culturemedium to cause different position ofC. acum inata leaf exp lants to appear he te rogeneity.
Key words　 tissue culture;cellw all;acid deg rada tion
1　 Introduction
Plant tissue cultu re is the coursew hich a cultu re
medium m ixed by artificia l induces plant to ce lldedif-
ferentiation and red ifferen tiation
[ 1]
. But it isn’ t so
distinct that the mechanism of cu lture medium indu-
cing the plan t ce ll to dediffe rentiation
[ 2 ～ 4]
. W e have
proposed the hypothesis that, ac idm aterial of the cul-
ture medium pre scription make the leaf in the acid
environment and lead to the acid deg radation o f the
no rmal plant cellw all at first, then present pro toplast
break ing aw ay from the ce llw a ll, furthe r ce ll organs
re form ing o r rebuild ing
[ 5]
. Th is paper used the leaf of
the pecu lia r species of our country-C. acum inata ,
through the course of ana tom ical observation o f tissue
cu lture to understand the in fluence of the acid culture
medium on the ce llw all degrading o f the leaves ofC.
acum inata.
2　Materials and methods
The young leafw as chosen and cu t to the slice o f
1 cm ×1 cm , and then the upper epiderm is w as put
inWP cu lturemedium w ith pH 5. 8wh ich containsα-
naph thy l aceta te (NAA) 0. 2mg /L, 6-Benzy lam ino
purine (6-BA) 1. 0 mg /L, KH2 PO 3(mg /L). The re
w as ca llus in explants after 30 day culture in the con-
dition 28℃[ 6, 7] . Exp lants callus w ere taken for free-
hand slicew ith the sec tions smaller than 10 μm. The
selected sections w as pu t in to 1% to lu id ine b lue to
dye about ha lf am inute and then took ou t for w ashing
and moun ting w ith the distilled w ater. The opticsm i-
croscope of O lympus (BH— 2 )was used to observe
and take pictu res under 1 000× oilm irro rs.
3　Results and Analysing
3. 1　A natomy cha racteristic of w eak ly deg rading o f
C. acum inata leaves cellw a ll
Epide rm is ce ll, palisade and sponge tissue cell
in a part o f epide rm is on the slice of C. acum inata
leaves, whose ce llw all demarcation line is still c lear
and in tac t, but some cell w alls such epiderm is and
sponge tissue cell had a lready presen ted the w eak ly
deg rada tion phenomenon 30 day after the cultivation
(P lateⅠ :1).
3. 2　Anatomy cha racteristic of obv iously deg radation
ofC. acum inata leaves ce llw a ll
Anothe r pa rt o f epiderm is on the slice o fC. acu-
m inata leaves behaved that the ce ll w all dem arcation
line of palisade and sponge tissue cell had been al-
ready fuzzy, incomple te. Its typical mo rpholog ical
charac te ristics had already disappeared, most cell
w a lls w e re obviously degraded and the ce ll began to
rediffe rentiate. (P la teⅠ:2).
3. 3　Ana tomy characteristic of to tally degrad ing o f
C. acum inata leaves cellw a ll
Parenchyma ce ll be tw een upper epiderm is and
ve in vascula r bun lde in C. acum ina ta leaves, whose
localized cellw all had degraded to ta lly, had the cha r-
acteristics o f p lasma membrane broken, cell nuc leus
escaped from cy toplasm and ente r to in te rstitial
space. A pa rt of cy top lasm flew in to interstitial space
too, and in the p lace of broken p lasm a membrane
some o rgane lles began to assemble (Pla teⅠ:3).
3. 4　Anatomy characteristic of disappea rance o f C.
acum inata leaves cellw a ll
Some phenomena appear in sponge tissue ce ll
be tw een uppe r epiderm is and ve in vascu lar bunlde of
C. acum ina ta leaves 30 day after the cultiva tion in
the cu lturemed ium , such thatwhose cellw all is com-
plete ly deg raded, p ro toplast w ithout ce llw a ll p resent
uniform distribution, a ll kinds of o rganelles and ce ll
nuclears assembled (P la teⅠ:4).
4　Ddiscussion
Ce llw all o fC. acum inata leaves p resented ana t-
omy characteristic o fw eakly degrading, obviously de-
grad ing, to tally deg rading and ce ll w all disappearing
under tissue cu lture condition. W e th ink that it may
be the acid materials in the culture medium w ith pH
5. 8 enab le the leaf in acid env ironmen t and lead to
the acid deg rada tion of the ce ll w a l.l A fter the ce ll
w all appea r the acid degradation, plasm a membrane
because of expand ing and p ressing m ight make ce ll
nucleus escaped, some protoplast and o rganelles ove r-
flow. And then it makes these o rganelles concentra te
on distributing to form ce ll recomb ination w ith es-
caped ce ll nucleus
[ 8, 9]
.
Ce llw all of identical C. acum ina ta leaf appea r
vary g reatly in the ir degree of degradation and even
disappearing at the same the condition of tissue cu l-
ture and cultiva tion time. Itm ay be the uppe r epide r-
m is pro truding to in fluence diffe rent contacts po sition
w ith cu lture m edium and make acid m aterial of cu l-
ture med ium to cause diffe rent po sition o fC. acum i-
nata leaf exp lants to show hete rogeneity
[ 1, 10 ～ 12]
.
Reference
1. DeP ierre JW , E rna ster L. Enzym e hopology of intace llula
m em branes. Ann Rev B iochem , 1977, 46:201 ～ 262
2. B row nlee C, Be rger F. Ex trace llu lar m atrix and pa ttern in
p lant embryo s:on the lookout for developm en ta l inform a-
tion. T rends Gene t, 1995, 11:344 ～ 348
3. Ray le, C le land. The Acid G row th Theo ry o f aux in-induced
cell e longation is a live and we l.l P lan t Phy sio l, 1992, 4:
1271 ～ 1274
4. Dan ie l J. Cosg rove. Expansive g row th of plan t cell wa lls.
P lant Physiol Biochem, 2000, 38:109 ～ 124
130 　　　　　　植　　物　　研　　究　　　　　　　　　　　　　　　　　　25卷
5. Zu Yuangang, Yu Jinghua, Tang Zhonghua, et a l. P ossible
m echan ism s invo lved w ith the form a tion of leaf callus. Bu l-
letin of Bo tanical Resea rch, 2005, 1(25):1 ～ 2
6. Le Van T H , Takam ura T, Tanaka M. Ca llus fo rm a tion and
p lant reg eneration from ca llus through som a tic emb ryo struc-
tu res in Cym bidium orchid. P lant Science, 2004, 166:1443
～ 1449
7. Nay lo r J, Sande rG , Skoog F. M itosis and Cell enlargem ent
w ithout cell d iv ision in excised tobacco pith tissue. Physio.l
P lan t, 1954, 7:25 ～ 28
8. S tan ley H , Bo tas J, M alho tra V. The m echanism o f Go lg i
seg rega tion du ring m itosis is ce ll type-spec ific. P roc Natl
Acad Sci, 1997, 94:14467 ～ 14470
9. M alinow ski R, F ilipeck iM. The ro le o f ce llw a ll in plan t em-
bryogene sis. C ellu la r and m o le cula r bio logy le tte rs, 2002, 7:
1137 ～ 1151
10. Tob ias R, C onwayW , Sam s C. Po lyga lactu rona se isozymes
from Bo try tis c inerea g rown on app le pectin. B iochem M o l
Bio l In t, 1993, 5:829 ～ 837
11. M cQueen-M ason S J, F ry S C, Du rachko D M. The re la-
tionsh ip be tw een xy log lucan endotransg lycosy la se and in-
v itro ce ll w a ll ex tension in cucumber hypoco ty ls. P lan ta,
1993, 3:327 ～ 331
12. Biro R T, Sun D Y, Se rlin B S. Cha rac te riza tion o f oat
calm odu lin and radioimm unoassay o f its subce llular d istri-
bution. P lant Phy sio logy, 1984, 15:382 ～ 386
Explanation of plate
P t:pa lisade tissue;Eu:upper ep iderm is;Sp: sponge tissue;
E l:low er ep iderm is.
P la teⅠ 　1. Under tissue culture conditions, w eak ly deg rad-
ing o fC. acum ina ta leaves ce ll wa l.l A rrow m a rks w eak ly de-
g rading ce ll w al.l × 1 000;2. Unde r tissue cu lture condi-
tions, strong ly degrad ing o fC. acum inata leaves ce llw a l.l A r-
row m a rks pa lisade tissue cell w all disappeared and obv iou sly
degraded. ×1 000;3. Unde r tissue culture conditions, com-
p le te deg rada tion o fC. acum ina ta leave s ce ll wa l.l B lack ar-
row m a rks escaped ce ll nucleus;W hite a rrow m a rks a ll kinds
of o rgane lles and cell nuc lea rs concentra ting and distribu ting in
the place of broken p la sm a m em brane. × 1 000;4. Unde r
tissue cu lture condition, disapea ring o f C. acum inata leaves
cellw a l.l A rrow ma rks all kinds of o rgane lles and ce ll nuclears
assemb led. × 1 000
ZU Yuan-Gang et a l:Influence o f acid cu ltu re medium on ce ll w all deg radation ofCamptotheca acum inata leave s P lateⅠ
See explanation at the end of text
1312期 祖元刚等:酸性培养基对喜树叶片细胞壁降解的影响