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Growth A na l ysis and Plant Hormone Stud i e s
i n A p p l e ( Malus sylve s t r i s Mi l l . )
A the s i s submi t t e d i n par t ia l f u l filme n t of the
r e quireme n t s for the d e g r e e o f D o c t or o f Phi l o s ophy
at
De par t me n t o f H o r t icu l t ur e
Mas s e y Univ e r s i t y
Palme rs t on
N o r th
N e w Zeala nd
Heung Sub Park
1 975
To
My pare n t s , S o on Ja , J i n S o o , S i Young, and J i n Hyoung .
S in c e we a r e a s su r e d t ha t t h e a l l - w i s e Creat o r has
o b se rve d the m o s t e xa c t propor t i ons , o f number , we igh t and
m e asur e , i n the
make of all things ; t he most like l y wa y
t he r e fore , t o ge t any i n s i ght i n t o t h e nature o f t hose part s
o f t h e c r e a ture , whi c h c ome w i t h i n our ob s e rvat i on , mus t i n
a l l r e a s o n b e t o numb e r , we i gh a n d me a sure .
"
G o d ha s n o t only c omprehende d t h e du s t of the e ar t h i n
a mea sure, and w e i ghe d the mount a ins i n s c ales , a n d t he h i l l s
"
i n a b a lanc e , I s a i . X l . 1 2 .
( Hale s , 1727 ) .
i
A c knowle d geme n t s
S i n c e r e t hankG t o my supervi s o r Pr ofe s s o r J . A .
V e a l e ; o the r me mb e rs o f t he De part m e n t o f Hort i cu l ture
par t i cularly Dr . D . J. W o o l l e y for his a dvi c e on e xp e r i me n t a l
p r o c e dure s and a s s i stan c e wi th t h e manus c r ipt ; Pr o f e s s or
R . E . Munford for a s s i s t an c e w i t h the gro wt � analys i s data ;
Dr . E . D . P e nn y f o r advi c e o n t e chnique s in the e a r l y s ta ge s ;
Dr . J . P . K e r r Pla nt Phys i o l ogy Divis i o n , D . S . I . R . f o r
as s i s tanc e wi t h t h e pre para t i on o f t h e t he s i s ; several who
a s s i s t e d wi th p r o o freadi ng ; t o le ran c e o f
the
B o t any/Zo o l o g y
D e par tment i n t h e u s e o f e quipment and fa c i l i t i e s ; Mr . K .
J . F ourneau who e nc ourage d f i e l d e xpe r i me nta t i on i n h i s
o r chard and Mr . K .
W.
Ki ddle ( A pple and Pear B oard ) wh o
o rgani z e d for the expe r i me nt a l f i e ld i n Hasti ngs .
3
Fur th e r ext e nd e d thanks f o r g i f t s of H- z e a t i n ,
G i bb e r e llins , a n d Tan-ginb o zu dwa r f r i c e s e e d s ; t o Dr . D . S .
Le tham ( R e s e a r c h S c h o o l o f Bio l o g i c a l S c i enc e , Aus tralian
N a t i onal Unive r s i t y Canb e rra , Aus t ra l ia ) , Dr . D. B r oadb e n t
(I. C.
I.
Pharma c eu t i cals Divi s i on , A ld e r l e y Park , Mac c le s ­
f i e ld , Che s h i r e , England ) , and Dr , Y . Murakami ( Na t i o na l
I ns t i tute o f A g r i cultur a l S c i e n c e , T okyo , Japan ) r e spe c t iv e l y .
Spe c i al thanks t o my wi fe f o r support , e n c ourage­
men t , and t y p i n g .
Gra t i tude expr e s s e d t o the f ollowing o r gani z a t i on
whi c h gave f i na n c ial supp o r t and e n c ourageme nt ; K o r e an
G ove rnme n t , UNESCO, C o l omb o Plan an d Mas s e y Unive r s i t y .
ii
A b s t ra c t
Part I
Growt h ana l y s i s s t ud i e s
Pre v i o u s d a t a o n gravi t a t ional e f fe c t s on s h o o t growt h and
f l o we ring have b e e n inc o ns i s t e nt .
A t t empt s h a v e been mad e to i nve s t i ga t e
s h o o t grow t h and flowe ring on sho o t s w i t h a 3 /8 phyll o t a x i s in 9 0 c m and
1 50 cm l a t eral s .
The s e w e re be n t at d i f fe re nt t ime s t o t h e h ori z on t al
or t o a p e nd ul ous p o si t i on in ap ple varie t ie s Re d Deli c i ou s and Granny
MM
Smi t h on
1 06 ro o t s t o cks grown unde r a semi -int ensive s ys t e m on a
c omme rc ial orchard in t h e ma j o r app l e gro w ing area o f Ha s t ings .
t r e a t ment s c ompri se d :
T h e four
h orizontal or p e nd u l o u s b e nding d uring t he d o rmant
p e r i o d , at p e t al fall, s e c o nd c o ve r s t a ge and w i t h normal vert i c a l
l a t era l s a s c ontrol s .
Hori z o� t al bend ing inc re a s e d t o t a l s ho o t growt h and fl owe ring
re l�tive to the vert i c a l c ontro l s in t he 90 c m treatment in oa t h vari e ­
t ie s .
There se e me d t o b e a t e nd e nc y t o d e c re a se t o tal sho o t gro w t h w h e n
t h e t ime o f b e nd ing w a s l a t e r in t h e s e a s on and
no
d i f f e re nc e s in fl ower­
ing o c c urre d among the h o ri zont a l b e n d i ng t re a t me nt .
O n the o t h e r hand
s h o o t growt h wa s re latlve l y c o n s t ant in a l l t re a t ment s in the 1 5 0 c m
A very si�ni f icant inc rease i n f l owering , h ow e v e r, w a s f ound
t re a t me nt .
in t h e p e t al fall pe ndul o u s b end i�g .
In t h e d ormant p e r i o d p e nd u l ou s
b end ing t he re was a sl i ght e ffe c t o n t h e fl owe r promo t i on re l a t i v e t o
the v e rt i c a l s .
The prod u c t i o n o f l a t e rals and f l o w e r bud s wa s always more
pronou nc e d on the uppe r s id e o f the b e nt s ho o t s, with a n i nt erm e d i a t e
o n t h e flanks, and gre a t ly inhib i t e d e f fe c t s o n t h e lower s i d e , i nd i ­
c a t i ng a s t e e p l inear re l a t i onship from t h e l ower t o t h e u p p e r d u ring
t h e d o rmant p e ri o d t r e a t me nt in all e xp e rime nt s .
G e ne ral ly , t h e p e r­
c e n t a ge s o f sh o o t growt h and flower prod u c t i o n were incre a s e d fro m t he
d ormant period b e nd ing t o the p e t al fal l , s e c ond c ov e r and t o t h e
ver t i c al c ontrol .
The gre a t e s t inc re a s e in s h o o t l e ngth and incre a s e d p e r c ent age
of fl o w e ring in all e xp e rime nt s were fou nd in the ap i c a l whorl zone�
and t h e s e furt h e r d e c r e a s e d from t h e 1 s t t o the 2nd , and t o the 3rd
w h o rl;
th i s was t he c a s e for sho o t grow t h
w h orl wa s
n
flowering i n t h e f i r s t
the inhe re n t prope rt i e s in Red D e l i c i ou s .
The s h o o t gro w t h a nd flowering at the d i f fe re nt wh orl s in 1 50 c m l e n g t h
l a t era l s b e n t p e nd ul o u sly i n R e d D e l i c i o u s s h owed a quadra t i c r e l a t i on­
sh i p d u e to the long e r sho o t s in t h e ap i c a l and t h e arc h p o s i t i o n on t he
s h o o t when b e nt a t t he 5t h whorl in a l l t rea tment s .
Bu t at t he 5 t h w horl
fl owe ring was r e d u c e d c onsid e rabl y , b e c au s e o f sub s t an t i a l l a t e ra l grow t h .
iii
In ord e r t o ae scribe t h e grow t h r e lat i o n ship b e t we e n s h o o t
v o lume a nd t o t al lea f a r e a a n i n d e x b a s e d o n t he r a t i o o f v e ge t a t i v e
a n d r e pr o d uct ive r e sp o n s e s w a s e st abli s h e d e .g . v e ge t a t ive 10.83
a n d r e pr o duct i ve 19.R0-24.4o.
The re l a t io n s h ip of s h o o t grow t h and fl ow e r ing a r e di scus s e d
i n t e rms of a h or m o n e b a l ance t h e o r y .
iv
Ab s t ract
Part I I
Plan t h o rmone
I n ord er t o e s t ab lish a
�nderst�nding
for an
studies
ratio
o f physiolo gi c a�
of
different
phenomena,
p l a n t hormone s
app r o p r ia t e e x t ra c t i o n
pro c e d u r e s a r e re q u i r e d f o r espec i a l l y app l e l e a ve s whic h are ric h in
p he n o l i c c ompo u nd s and ot� e r inhibi t ors . T h e re fore e xtrac t i on pr o c ed 3
1
f
.
.
+..,
H - z e a t 1n.
.
u r e s a n d purl 1c a '-,.�on
w e r e exa m1ne
L- I AA an d
.
d us1ng
14
Loss of
c-I A A d uring ext rac tion pro c e dure s w a s due t o a
.
.
high p H in t h e aqueous phase during sol v e n t
14
r e c o v e ry o f
G - I A A wa s 3 . 8%
o:l; vent
:g�rt.it{!i>� "and] c o lumn
partitioning.
The final
c hr orna tography.
o n a silic a ge l - c e l i t e c olumn ani a S e p hadex LH..:.2o c o lu mn , g i v i n g 80%
r e c o ve ry in 30 ml
i "l 2 0
ml
elution
volume
aro u nd t h e maL1. peak and 9 Cf'/o r e c ove r y
e l uti on vo lume a round t h e main p e a k r e spe c t iv e ly .
Nearly
1 00%
r e c o very from a Se pha d e � S - 1 0 c o lumn wa s ob t ained .
50-57% re c ov e ry o f
14
c- IAA was ob t ained in c e l l ulose thin l a y e r c hroma t o graphy a t t h e H f o f
14
I AA , an d; J n o l o s s o f
c - I A A o c c u rred during 3 days s t o r a ge in a d a rk
[
c ab ine t .
The par t i tion c oe f f i c ie n t o f
3
H - z e a t ift a t p H 8 . 3
w i t h e t hyl a c e t at e and 0 . 48R w i t h n -bu t a no l ;
was
13 . 1 2
a t pH 2 . 5 , 1 08 - 8 9 w i t h
e t h y l ace t a�e and 16 . 73 w i t h n -bu t a n o l . Bac kw a s h ing c a n r e c over
3
H - z e a t in from ethyl a ce t ate pha se wh i c h was partit ion e d a t p H 2 . 5 .
3
80% r e c o very o f H - z e a t i n in the first 1 , 000 m l wa s o b t a i n e d fr o m
S e p h a d e x G - 1 0 and Dowex 5 0
o b t aine d in a 20 m l
c on t a inin g
p eak
0 . 001 M H C l .
W
x
8.
88 . 6% r e c o v e ry o f
3H-zeatin
c ou l d b e
S e p h a d e x LH- 2 0 eluted w i t h 9 5% E t O H
3
The b e havio u r o f H - z e a t in w a s s t u d i e d in
using
pape r c h r o mato graphy and c e l lu l o se , DEAE c e l lulose and s i l i c a ge l ,
3
3
t h in lay e r c h r omat o graphy , ab out 82 -6Cf'/o o f H - z e a t i n t h e R f o f H-ze a t in
b e i n g r e c ove re d .
F our se r ies o f plan t hormo �es w e r e d e t e rm i n e d f r o m app l e
l e a v e s b y e t hyl a c e t A t e p a r t i t i o nin g , Se phad�x G- 1 0 c o lumn , s i l i c a ge l ­
c e l i t e c o lumn
and
c e l lul o se t h i� laye r c h r omat o graphy o f a cid i c f ract ­
i o n s c on t ai n in g auxi n - , g ibbe re l l in- , ani ABA-l ike s u b s t anc e s , a n d by
b u t ano l p a r t i t ionin g , S e phad ex G - 1 0 , S e p had e x LH-20 , an d"DEAE c e l lu l o se
t hi n l ay e r c h r oma t o graphy for c yt okini n - l ike sub s t an c e s f r o m b a s ic
f ra c t io n s .
Po s s ib l y t w o kind s o f aux i n - l i ke s u b s t an c e s we re found a n d
p o s s i b ly GA , GA , G A , GA , or GA and G A -like sub s t an c e s w e re e lu t e d
1
4
5
3
9
8
from a s i l i c a ge l - c e l i t e ad s o rpt ion c o lumn .
S e v e r al grou p s o � c yt okinin-
V
like s u b st a n c e s w e r e ob t a ine d from S e p ha d e x LH-20 c olumn c hr omat o gr aphy ,
p o s s ibly z e a t in , z e a t in -r i b o s i d e and o t h e r cyt okin i n s we re found in
apple leave s .
Base d on t he e s t lmat io� o f e ach plant h o rmo n e f r o m t hi n
l aye r c hr oma t o graphy , a rel�tive
plan t hormone ind e x w a s e s t abli s he d ,
i . e . , Relative Aux in Ac t i v i t y I n d e x , Relativ� I n h ib i t o r A c t ivi t y I nde x ,
Relative G i b b e rellin A c t ivity I n d e x , and Rela t i v e Cyt o ki n i n A c t iv it y
Ind ex ,
re spe c t i v e ly , t he h o r mone
giv ing t he h i gh e st rat i o
be i n g c on s ije r e d the d ominan t h o r m o n al f act o r
a t t ha t st a ge o f d e velopment .
GLC t e c hn i q ues w e r e also s t ud i e d for plant hormo n e analys i s ,
u si n g 7% OV- 1 a n d NAA , I A A , I PA , GA , GA , GA , GA , GA , GA , GA
and
4
3
13
5
9
7
1
A B A marke r s t o e s tabli s h reten t i on t imes a�d d e t e c t o r r e s p o n s e a t t h e
2 . 5 n g level .
N . O . -bis ( trime t hyls ilyl ) t riflu o roa c e tamide ( BS TFA )
t oge t h e r w i t h Tri�e t hylc hloros ilane ( TMC S ) silyl r e a ge n t s p r o d u c e d t h e
b e s t peak h e i gh t s f o r IAA , I PA , GA
hal f and t he GA
9
peak by 20% .
3
and GA
1
bu t r e d u c e d t he A BA p e a k by
vi
Table o f C on t e n t s
C ha p t e r
A c knowle dgements
Abs trac t Par t I : G r owth· analy s i s s t u di e s
Par t II : P lant hormone s t u d i e s
T a b l e o f C ont e nt s
L i s t o f Tab l e s
L i s t o f Figur e s
Abb r e v i a t i ons
Part I
ii
iv
vi
xv i
xviii
xxi
Growth ana lysis s tudi e s
1
1
Int r o duc tion
2
R e v i e w o f L i t e rature
2.1 .
H i s t ori cal out l i n e o f gravimorphi c s tudi e s
2
2.2.
B e nd ing t e c h n i qu e s in ge ne ral orc ha r d prac t i c e
4
2.3.
The e ffe c t o f grav i ty o n f l owe r i n g
4
2 .3.1 .
Pre c o c i ous flowe r i n g
4
2 .3 . 2 .
App le t re e s
5
2.3.3.
S i l v i culture
7
2 .4 .
The e ff e c t of grav i ty o n v e ge ta t i v e gr owth
7
2 .4 . 1 .
Ext e n s i on o f s h o o t gr owth
7
2.4.2.
Out gr owth o f buds
8
2 .4.3.
D i s t r i bu t i on of shoots on the la t e ral
10
2 . 4 . L� .
Sho o t po s i t i on o n outgrowth o f late ral buds
11
2.4.5.
The i nt e ra c t i on b e t we e n api c a l dominanc e
and e f fe c t o f gravity
2.5.
2.6.
3
12
Grav i t a t i onal s t i mulus
12
2.5. 1 .
R o t a t i on e ffe c t
12
2.5.2.
D i s t r i b u t i on o f nut r i e nt , as s i m i lat e s , and
e nzyme s
13
2 .5.3.
Gr o w t h sub stan c e s
14
2 .5.4.
Ana t omy o f wood and grav i t a t i ona l e f f e c t
17
2 .S. 4 . 1 .
C ompr e ss i on wood
17
2.5.4.2.
Te ns i on wood
18
2.5 . 4 . 3 .
Phloem
19
T h e o u t growth o f sho o t a n d sho o t gr owth ( Ap i c a l
d om i nan c e )
19
2.6.1 .
Ava i la b i l i ty o f c y t okinins
27
2 .6 . 2 .
IAA c on j u ga t ing syst e m w i t h a sparta t e
29
2.6.3.
A s i �n i f i c ant r o l e o f aspar t a t e
31
Mat erials a n d Me t hods
33
3. 1 .
Organ i sa t i o n of f i e ld e xpe r ime nt s
33
3.2.
App l i c a t i on o f t r e atments
33
3.3 .
).4.
4
4. 1 .
vii
3.2.1 .
G e ne ral
3 .2.2.
Expe r i me nt 1
3 .2 .3.
Exp e r i m e nt 2
35
3 . 2 . �- .
Expe r i m e nt 3
35
33
33
M e a s ur e s o f s h o ot gr ov1th
35
3.3. 1 .
S h o ot diamet e r s
35
3 . 3 .2.
Leaf a r e a s
35
3 .3.3.
Rat i o o f t ota l leaf area to total i n c r e me nt
o f s ho ot v o lu m e
36
S h o ot le n gth
37
Stati st i c a l �nalyG e s
37
3 .4. 1 .
C las s i f i c ati on of bud pos iti one ( S e e Fi g . 4 )
37
3 .4.2.
Analys i s o f varian c e m o d e l s
37
3 .4. 3 .
Ortho gonal sub d i v i s i on o f ma i n e f f e cts and
3 .3.4.
i ntera ctions
39
Expe rimental r e sults
41
S h o ot l e n gth s e c t i o n
41
4. 1 . 1 .
Exp e r i m e nt 1
( Re d D e l i c i ou s with 2 9 p o s iti ons
of late rals , h o r i z ontal b e nd i n g )
41
4.1 . 1 . 1 .
Effe c t o f tre atm e nts
47
4. 1 . 1 .2 .
E f f e c t o f p o s i t i on
47
4. 1 . 2 .
Exp e r i me nt 2 ( Re d De l i c i ous w i th 45 p o s i t i o n s
o f late r a ls , p e ndul ous b e nd ing )
50
4. 1 .2 . 1 .
E f f e c t o f treatments
50
4. 1 . 2 .2.
E f f e c t of p o s i t i ons
50
4. 1 . 3 .
Expecime nt 3 ( Granny Smith , h o r i z ontal b e nd i n g )
55
4.2.
L� . 1 . 3 . 1 .
E f f e c t o f tr eatme nts
56
4 . 1 . 3.2 .
E f f e c t o f p o s i t i ons
56
Flowe r i ng sec t i o n
Lf . 2 . 1 .
( Re d D e l i c i o u s 90 c m late ral )
56
'+ . 2 . 1 . 1 .
E f f e c t o f treatments
56
4. 2 . 1 . 2 .
Eff e c t o f pos iti ons
60
4.2 . 2 .
Exp e r i me nt 2
61
4 . 2 .2. 1 .
E f f e c t o f t r e atme nts
61
4. 2 . 2 .2.
E f fect of p o s i ti ons
61
4.2.3.
4.3.
Exp e r i m e nt 1
56
Expe r ime nt 3
63
4 . 2 . 3.1.
E f f e c t o f tr eatments
63
4.2.3.2.
E f fe ct o f pos iti ons
63
Re lat i v e growth r e lati onship ( b e twe e n le a f a r e a and
Gho ot growth )
64
4.3. 1 .
64
Le a f area
viii
4 . 3 . 2.
S tudi e s o f ra t i o s o f t otal l e a f are a t o t o ta l
i n c r e ment o f s h o ot v olume (R)
5
5.1 .
5.2.
5.3.
D i s c u s s i on
73
S h o o t growth
73
5.1 .1 .
Total sho o t l e n gth
73
5.1 .2.
Or i e nta t i on e ff e c t
73
5.1 .3.
Whorl e ffe c t
78
Fl owe r i ng
80
5.2.1 .
Total flowe r i ng
80
5.2.2.
Or i e nta t i on e f f e c t
81
5 .2.3.
Wh orl e f f e c t
84
Relat i v e gr owth r e la t i onship
86
5 .3 . 1 .
Leaf area
86
5.3.2.
Rat i o of t o t a l leaf area to t o tal i n c r e me nt
of s h o o t v o lume
5.4.
66
Summa r y of d i s cu s s i on
Par t I I
Plant hormone s tu d i e s
86
89
97
6
I n t r o ducti on
97
7
Rev i e w of l i t e ra tu r e
99
Auxi n - like sub s t an c e s
99
7.1 .
7.1 . 1 .
The h i s t o r i c a l d e v e l opme n t o f e x t ra c t i on
pr o c e dur e s fer a c i d i c , neutra l , a n d b a s i c
fra c t i on i n t he de t e rmina t i on o f aux i n -l i ke
sub s tan c e s
7.1 .2.
The ac i d i c aux i n - like sub stanc e s a t p H 2 . 5 3 . 0 and pH 8 . 0 - 8 . 5
1 00
7. 1 . 3 .
A b ound or c o njuga t e -aux i n - l i ke s ub s tance
1 01
7.1 .4.
C o lumn c hr oma t ography
1 02
7. 1 .4.1 .
7.2.
99
Ads o rp t i o n c o lumn
1 02
7.1 .4. 1 . 1 .
Alumi na c olumn
1 02
7. 1 .4. 1 .2.
S i l i c a ge l - c e l i t e c o lumn
1 02
7 . 1 .4.2.
I on e x c ha nge c o lumn
1 03
7.1 .4.3.
Par t i t i o n c olumn
1 03
7.1 .4.3.1 .
S e l i c a gel c olumn
1 03
7.1 .4.3.2.
G e l .chroma t ography
1 04
Abs c i si c a cid and o t he r i nhib i t o r s
1 05
7.2 . 1 .
Hi s t o r i c a l pr ogre s s i n the i s ola t i on of AEA
1 05
7.2.2.
Solve n t par t i t i on a t d i f fe r e nt p H ' s w i t h
d i ffe r e n t o r ganic s o lv e nts
1 06
ix
7.2. 3 .
A n e utral i nh i b i t or i n t h e ne utral frac t i on
1 08
7 . 2 . 4.
A c on j ugat e o f a b s c i s i c a c i d
1 08
7.2 . 5 .
C olumn c hromat ography f o r pur i f i c a t i on o f
_
AEA
7.3.
G i b b e r e ll i n - l ike sub s tanc e s
7 . 3. 1 .
H i sto r i c a l p r o g r e s s o f i s olat i on f o r
g i b b e rellins
7 . 3 . 2.
G i b b e r e llin- l ike s ubstan c e s i n d i f f e r e nt
f ra c t i ons
7 . 3.2 . 1 .
An a c i d i c frac t i o n
7 . 3.2 . 1 .1 .
1 1 t1
1 1 rJ
Extrac t i on o f g i b b e re llins a t pH
2 . 5 - 3.0
7 . 3 . 2.1 . 2 .
Extra c t i on of g i bbe r e llins a t pH
8.0-8.5
7 . 3.2.2 .
A neutral frac tion
112
7.3.2.3.
B ound o r c on j u gate g i b b e relli n s
1 13
7 . 3 . 3.
7.4.
11 2
Par t i t i on o f g i b b e re l l i n w i t h d i f fe r e n t
s olve n t
113
7 . 3.4.
C ou n t e r c ur r e n t d i s t r i b u t i on
1 1 :4
7 . 3 . 5.
C o lumn chr o ma t o graphy
1 15
7.3.5.1 .
Par t i t i on c olumn
115
7.3.5.3.
G e l c hr o ma t o graphy
116
7 . 3.5.4 .
I on exc hange c olumn
116
C y t o k i n i n - like sub s tanc e s
1 16
7 .4 . 1.
D i s c overy o f kine t i n
116
7 . 4 . 2.
H i s t o r i cal p r o gr e s s o f i s o la t i o n f or z e a t i n
117
7 . 4 . 3.
An o u t l i ne o f e x t rac t i o n p r o c e dur e s and
c o lumn c hr omat o graphy
7.4.3. 1 .
119
The 6-sub s t i t u t e purinecand t h e i r
rib osides
1 20
7 . 4 . 3 . 1.1 .
D o w e x 5 0 c olumn
1 ·2 0
7 . 4 . 3.1 . 2.
Ze o -Karb 225 c olumn
1 20
7 . 4 . 3. 1 . 3.
An a n i o n exchange r e sin c olumn
as D ow e x 1
1 '20
7 . 4 . 3 . 1 . 4.
Oth e r c olumns
1 20
7 . 4 . 3 . 1 .5 .
Pre c ip i ta t i on
121
7 . 4 . 3.1 �6.
Pap e r and t h i n - layer chr oma t o graphy
121
7 . 4 . 3 . 2.
Z e a t i n r ib ot i de and o t h e r b ound f orms
of c yt ok i n i n s
7 . 4 . 4.
1 21
The o c c ur r e n c e o f c yt ok i n i ns in d i f fe re n t
p lants
12 2
X
7-5·
Dete r m i nati on o f plant h o r m o n e s by gas - l i q u i d
c hr omatography ( GLC )
1 22
7.5. 1 .
A u xins by GLC
123
7.5.2.
A b s c i s i c a c i d b y GLC
1 24
7 . 5 .2 . 1 .
Dete rminati o n o f ABA by a f lame i on i zat i o n
dete ctor
7.5.2.2.
7.5.2.3.
124
Determ i nat i o n o f ABA b y an e le ctr on
c a ptur e dete c t o r
1 26
I d e nt i f i c at i o n o f ABA
1 26
7.5.2.3.1 .
GC-MS f o r ABA
7.5.2.3.2.
I d e nti f i c a t i on of ABA b y rac e m i sati on
1 2.6
with U V i r ra d i ati on
7 .6.
8
8.1 .
7.5.3.
G ib b e r e l l i ns b y GLC a n d GC - MS
7.5.4.
C ytoki n i ns by GC - MS
L o s s o f authe nti c plant h o r m o n e s dur ing extrac t i o n
pr o c e du r e
129
7 .6. 1 .
L o s s o f IAA
129
7 .6 . 2 .
Loss o f other plant h o rmone s
1 31
Mate r ia l s and Methods
14
Loss o f
C - IAA dur ing e xtra c t i o n and puri f i c at i on
1 3'2
pr o c e dur e s
1 32
8.1 .1.
The plant mate r i a l s
132
8a1 . 2 .
S olve nt partiti o n c hr omato graphy
1 32
8.1 .3 .
C o lumn c hr omat ography
133
8. 1 .3 . 1 .
S e phadex ge l c h r o mato graphy
1 33
8. 1 .3.2.
A d s orpti on c o lumn chr omat ography
1 33
8.1 .4.
C e llulose thin laye r c hr omatography
1 35
8.1 .5.
8.2o
Deter m i nati on o f r a d i oactiv ity
3
B e ha v i our of H- ze atin in c h r omat o graphy
8.2.1 .
S o lv e nt partit i o n c hr omatography
8 . 2 . 1 . 1 . Pre parat i on and d e te r m i nation o f
1 35
1 36
3
1 36
H-
z eati n
8 .2 . 1 .2.
B a c k wa shing f r o� ethyl a c etate pha s e
8 . 2 . 1 . 3 . B a c k wa shing f r o m BuOH phase
3
8.2 .2.
H- z e atin i n c o lumn c hr omato graphy
1 37
1 37
8 . 2 .2 . 1 .
S e pha dex G - 1 0
1 37
8 . 2.2 . 2 .
Sephadex LH - 2 0
1 38
8 . 2 .2 .3.
8.2.2.4.
D owex 50 W x 8
3
B e havi our o f H - zeatin i n pap e r and
thin laye r c hr o mato graphy
8 .3.
1 36
B i oassay t e c h n i que s for IAA , ABA and GA
1 38
1 39
1 39
xi
8.3.1 .
The c ombina ti on of 5 mm oat and 1 0 mm whea t
c ole op t ile s e c tions f o r IAA and AEA
8.3.2.
T a n- ginb ozu dwa r f ric e s e e dlings and le t tu c e
h yp o c o t yl bioas s a y for gib b e rellins
8 .4.
141
8.3.2 . 1 .
Dwa r f ri c e bioassay wi t h gla s s t ub e s
1 41
8.3.2.2.
Mo difie d dwar f ric e bioassay
1 42
8 . 3 . 2-3 .
Le t tu c e h yp o c o t yl b i oa s say
1 42
Endogenous pla nt hormon e s in apple leave s
8.4. 1 .
E x t r a c tion p r o c e dures o f IAA, AEA , and GA s
1 44
1 44
8.4. 1 . 1 .
Le a f sample s and e xtrac tion p r o c e du r e s
1 44
8.4. 1 .2 .
E t h yl a c e ta t e phas e
1 44
8.4. 1 .2. 1 .
The s ys t e m A sili c a gel- c e li t e c olu mn1 4 4
8.4. 1 .2. 1 . 1 .
8.4. 1 .2. 1 .2.
8.4. 1 . 2 . 2 .
De t e rmina tion o f IAA - and ABAlike sub s t a nce s
1 45
De t e rmina tion o f gib b e r e llins
1 45
The s ys t e m B sili c a gel- c eli t e
c olumn c h r omat o graphy
1 45
8.4. 1 .3.
Bioassay for IAA and AEA
1 47
8.4.1 .4.
Bioassay f o r gib b e r e llin s
1 47
8.4. 1 .4. 1 .
8 . 4. 2 .
8 .5.
1 39
Barle y endosperm t e s t
Extrac tion pr o c e dure f o r c yt okinins
1 48
1 48
8 .4.2 . 1 .
A qu e ou s phase
1 48
8. 4.2 .2 .
Bi oassay for c yt okinin-like sub s tanc e s
1 49
A u t h e n tic pla nt hormone s and pla n t mate rials in gas
liquid chroma t o graphy
1 51
8 .5. 1 .
151
G e n e ral pr o c e dure of GLC
8 . 5. 1 . 1 .
Pac king c o lumn
151
8.5. 1 .2.
The GLC c onditions
1 51
8 .5 . 1 .3 .
P r e paration o f d e rivativ e s
151
8.5.2.
Gas chroma t o graphic s e para tion o f hormones
f r om apple leave s and c omparison wit h bioa s s a y
r e sult s
1 52
8.5.2.1 .
Plant e x t ra c t
1 52
8 .5.2 .2 .
GLC o f plant e x t ra c t s
1 52
8.5.2 .3.
The IAA - and ABA-lik e substanc e s in t h e
aliquo t o f e ther acidi c frac tion b y
·bi oassay
8.5.3.
GLC c alibra tion o f nat ural plant hormone s
and r e la t e d c ompounds
8 . 5 .4 .
1 53
GLC o f nat ural pla nt hormon e s and rela t e d
1 53
xii
synth e t i c p la nt h ormo n e s a fter c e l l u l o s e thi n
laye r c hr omat o gra phy
8.5.5.
1 53
E f f e c t of d i f fe r e nt s i lylat i on meth o d s o n
r e te nt i o n t i m e a n d p e ak he i ght o f natural
plant hormone s
8.5.5 . 1 .
8.5.5.2.
9
G i b b e r e l l i n A3
S e v e r a l authe nti c p la nt hormone s
1 54
1 54
1 54
Exp e r i me nta l r e sults
14
Loss o f
C - IAA dur i n g e xtracti o n a n d pur i f i c at i o n
1 57
pro c e dure s
1 57
9.1 .1.
E f fe c t o f p H i n the aqu e o u s phase s o n the
14
d e gradat i o n of
c - IAA in the extrac t i o n
pr o c e dure
1 57
9.1 .1 . 1 .
At pH 8 . 0 i n the aque o u s phase
1 57
9.1 .1 .2.
At pH 2 . 5 in the a qu e ou s pha s e
1 57
C o lumn c hr omato graphy
14
C - I AA f r o m Sepha d e x G - 1 0
9o 1 . 2 . 1 . R e c overy o f
9.1 .2.
9.1 .2.2.
c o lumn c hr omato graphy
14
c- I AA in the gra d i e nt
B e havi our o f
1 57
1 57
e lution o f the s i li c a ge l -c e l ite c o lu mn
c hromat o graphy
14
Re c ov e ry o f
C- I AA f r o m c e llulose t h i n
9.2 .
1 59
laye r chromato graphy
3
B e hav i our o f H - z e a t i n i n c hr omato graphy
162
9.2. 1 .
162
S o lv e nt part i t i on c hr omat o graphy
9.2. 1 .1 .
E f f e ct o f pH i n the aque ous pha s e o n
3
partiti o n o f H- z e at i n b y ethyl a c e tate
and butan o l
162
9. 2. 1 .2 .
B a c k washing f r o m e thyl a c etate pha se
162
9 . 2 . 1 .3.
Ba c k wa shing f r o m b utan o l pha s e b y
9.2 .2 .
3
0 . 5 M p h o s phate b u f f e r
H - z e atin i n c o lumn c hr o mato graphy
165
165
9.2 .2 . 1 .
Se pha d e x G - 1 0
165
9 . 2 .2. 2 .
D o wex 5 0 W x 8
1 65
9 . 2 .2.3.
S e phade x LH-20
3
9.2.3.
H - z e at i n i n pape r and thi n layer
c hromatography
9.3.
1 59
B i oassay te c hn i que s for IAA , ABA , and GAs
9.3. 1 .
165
168
168
The c omb inati on o f 5 mm oat and 1 0 mm wheat
c ol e o pt i le s e c t i o n s
168
x iii
9.3.2.
The d wa r f r i c e s e e d l i ngs and l e t tu c e h yp o c o t yl
b i oa s s a y f o r g i b b e r e llins
9.3.2. 1 .
1 71
Tan - g i nb o zu dwa r f r i c e s e e d l i n g i n t he
gla s s tub e s a n d a gar c u l ture m e d i u m
9.3.2.2.
1 71
C o mpar i s on o f d i f fe r e n t l e t t u c e var i e t i e s
f o r use i n t he l e t tuc e h yp o c o t yl b i oa s say
9.4.
for g i b b e r e l l i n s
1 73
Endogenous p la n t hormone s in apple l e ave s
1 73
9 .4. 1 .
IAA- a n d ABA - l i ke sub stanc e s i n apple
leave s
1 73
9.4. 1 . 1 .
IAA - l i ke sub s tanc e s
1 73
9.4. 1 .2.
ABA - l i ke substanc e s
1 78
9.4.2.
Gibb e r e l l i n-like s ub s tanc e s ln e t hyl a c e ta t e
1 78
pha s e
9.4.2 . 1 .
G ib b e r e ll i n- l ike sub s tan c e s i n the
f ra c t i on from t he s ys t e m A s i l i c a g e l ­
c e l i t e c olumn
4
9 . . 2 . 1 . 1 . Fra c t i ons from c o lumn
9.4.2. 1 .2.
1 79
C e llul o s e thin laye r chroma t ography
from the p o o l e d frRc t i ons o f c o lumn
9. 4 . 2 . 2 .
1 79
1 82
G i bb e r e l l i n- l i ke sub s tanc e s from the
s ys t e m B s i li c a g e l - c e l i t e c olumn e lu t e d
b y h e xane , d i e thyl e t he r , e thyl a c e ta t e ,
and me t hanol
9.4.3 .
1 82
Cyt oki n i n- like sub s tanc e s of apple leave s
f r om S e phadex LH- 2 0 a n d DEAE c e l lu l o s e t h i n
laye r c hr oma t o graphy
9.4.3. 1 .
On S e p hade x LH-20 c o lumn c hr oma t o graphy
9.4.3.2.
On DEAE c e l l u l o s e t h i n layer c h r o ma tegraphy
9.5.
G LC o f authe n t i c p lant hormo n e s and plant amt e r ials
9.5. 1 .
1 83
1 83
1 86
1 90
Gas c hr o ma t ograp h i c s e pa ra t i on from the
apple l e av e s and c ompar i s o n w i th b i oa s sa y
r e su l t s
9.5 . 1 . 1 .
GLC o f p lant e x t ra c t s
9.5. 1 .2.
T h e IAA and ABA-like sub s tan c e s i n the
1 90
1 90
a l i qu o t o f a c i di c e ther frac t i on b y
"b i oa s say
9.5.2.
1 90
S tandard c a l i b ra t i on o f s e v e ral authe nt i c
plant h ormone s b y GLC
9.5.3.
GLC o f a u the n t i c p lant h o r m o ne s and real t e d
syn th e t i c pla n t hormone s a f t e r c e llu l o s e TLC 1 94
xiv
9. 5.4.
E f fe c t o f d i f fe r e nt r e a gen t s on f orma t i on o f
ME-
and ME - T MS d e r i va t i v e s o f s e v e r a l authe nt i c
p lan t hormone s and o n t he r e t e nt i o n t ime and
peak he i gh t s
9 . 5 .4. 1 .
9. 5 .4 . 2.
10
Gibberellin A
3
Several aut hen t i c p lan t h o r m o n e s
D i s c u s s i on
14
1 0. 1 . L oss o f
C - IAA dur i n g extra c t i on and puri f i c at i on
p r o c e dur e s
10. 1 . 1 .
10 . 1 . 2 .
10 . 1 .3 .
1 95
195
201
201
E f fe c t o f p H i n the aque ous pha s e s on the
14
C -IAA
loss o f
14
c�IAA
C olumn c h r o ma t ography o f
.
14
Re c ov e r y o f
C-IAA fr om c e llul o s e t h i n
layer c hr o ma t o graphy
3
10.2 . Behav i our of H- z e a t i n in c h r o ma t o graphy
10 . 2 . 1 .
195
S olvent par t i t i on c hr oma t ography
201
204
205
205
205
10 . 2 . 1 . 1o E f fe c t of pH in aque ous phase o n par t i ­
3
t i on o f H - z e a t i n with e thyl a c e t a t e
and b u t a n o l
10 . 2 . 1 . 2 . Bac k was h i ng
3
H - zeat i n i n c olumn c hr omat o graphy
10.2 . 2 .
3
H-zeat i n i n pap e r and t h i n laye r
10 . 2 . 3 .
205
206
207
c hromatography
208
10 . 3 . Bi oas say t e c hni que s for IAA, ABA and GAs
208
10 . 3 . 1 .
The c omb i na t i on o f 5 mm and 1 0 mm whe a t
c ol e op t i le s e c t i on
1 0.3 . 2.
208
The dwa r f r i c e s e e d l i ngs and le t t u c e
hypoc o t yl b i oassay f o r g i b b e re l l i ns
10 . 3 . 2 . 1 .
Tan - gi nbo z u dwar f r i c e se e dl i n g in the
glass t�b e s and agar culture me d ium
1 0.3 . 2 . 2 .
209
209
C ompar i s on o f d i f f e r e nt var i e t i e s f o r
use i n t he l e t tuc e hyp o c o t yl b i oas says
for g i b b e r e l lins
1 0.4 .
End o g e n ous plant h o r m one s i n apple leav e s
10 . 4 . 1.
10.4 . 2.
210
210
IAA- a n d A BA -l i ke sub s t anc e s i n apple
leave s
210
G ibbe r e ll i n - like sub s tanc e s
215
XV
1 0.4 . 3 .
C ytoki n i n - l ike sub s tanc e s
10.5.
G LC o f authe n t i c p la n t hormone s and plant ma t e rials
1 0.6.
G e ne ra l d i s c u s s i o n i n c on junc t i o n w i t h fur th e r work
21 9
223
and c on c lu s i on
224
Lit e ra t ur e c i t e d
234
xvi
List of Tables
page
Tab le 1 .
E f f e c t o f hori zont a l b e nd ing a t vari ous t i m e s o n
s h o o t l e n g t h , t o t a l l e a f area and numb e r o f l e a ve s :
Expe r i m e nt 1 Red De l i c i ou s - All 29 shoot p os i t i ons
Tab le 2 .
40
E f f e c t o f hori zontal b e n d i ng a t vari ous t i m e s o n
s h o o t l e n gth
a n d fl o w e r i n g
at the e nd o f t h e
gr o wing s e a s on a n d a t b l o s som: Expe r i me n t 1
R e d Deli c i ou s - Analys i s o f var ian c e for a l l who r l s
o t h e r t h a n t h e api c a l w h o r l
Table 3 .
46
E f fe c t o f p e n dulous b e nd ing a t var i ous t im e s o n
s h o ot l e ngth
a n d fl owe r i ng
measur e d at the e nd
o f the gr o w i n g s e a s o n a n d at b l o s s o m : Exp e r i m e n t 2
Re d De lic i ou s - A nal ys i s o f vari ous fo� a l l wh o r l s
o t h e r t h a n t h e api c a l wh orl
Tab le
L�.
E f fe c t o f h o r i zontal b e n d i n g a t vari ous t ime s on
shoot l e ngth
and f l o w e r i ng
measur e d at t h e e nd of
t h e gr owing s eason and at b l o s s o m : Expe r i m e n t 3
Granny S m i t h- Analys i s o f vari ous for all whor l s
62
o t her t ha n t h e api c a l w h or l
Tab le 5 .
S t ud i e s o n shoot gro wth i n r e la t i on t o l e a f a r e a i n
65
t h e v e r t i c a l and h o r i z o ntal t r e a tme nts
Tab l e 6 .
A flow c ha r t of e x t ra c t i on p r o c e dure for I AA-, ABA-,
GA - , and C k-Jike sub s t a n c e s in apple leave s
1 43
14
Tab le 7 .
E f fec t o f pH i n t h e a qu e ous pha s e on loss o f
Tab le 8 .
1 56
IAA i n the extra c t i on p r o c e dure
14
c - IAA fro m a s i l i c a g e l- c e l i t e s olumn
R e c ov e r y o f
C-
b y the gra d i e n t e lu t i on me t h o d
1 60
Tab l e 9.
E f fe c t of s olve n t o n t h e par t i t i o n of
3
H- z e a t i n a t
pH 2 . 5 and 8 . 3 an d par t i t i on c o - e f f i c i e n t s w i th
161
e thyl a c e ta t e and b u tanol
Tab le 1 0 A a n d B.
Back wa shing fr om e thyl a c etate a n d b u tanol
phase
Tab l e 1 1 .
Par t i t i on of
3
1 63
H - z e a t i n from S e phadex G- 1 0 c olumn
c hr oma t ography
Tab le 1 2 .
Pa t t er n i n the fra c t i on at i on o f
D owex 50
Tab l e 1 3 .
W
3
1 64
H-zeat i n from
x 8
1 64
B e ha v i our of d i f f e r e n t c yt okinins i n Se pha d e x LH-20
c o lumn chr oma t o graphy
1 66
xvii
T a b l e 1 4.
S tandard
l i b ra t i on of the oa t and wh e a t c o l e op t ile
b i o a s s yas e i th e r s i ngly or c o mb i n e d in the t e s t tube
f or the r e s p o n s e to IAA and AEA
Tab le 1 5 .
1 70
C o mpar i so n of dwarf r i c e s e e d l i ngs a t d i f f e r e n t
c on c e nt ra t i on o f GA
3
i n t h e gla s s t ub e s and a gar
1 71 a
c ul tu r e m e d i um
Table 1 6.
C o mpar i s o n o f dwarf r i c e s e e d l i ng s f o r the i r r e s ­
p on s e t o d i fferent c o nc en t r a t i ons o f GA
3
under
di f f e re nt gr o wt h c on d i t i on s
Tab le 1 7.
C ompar i s o n o f d i f fe r e n t le t tu c e var i e t i e s f o r u s e
i n t he le t tuc e hyp o c o ty l b i oassay f o r gib b e re l l i n s
Tab le 1 8.
1 73 a
E s t i ma t i o n o f IAA - a n d ABA - like sub stanc e s f r o m
s i l i c a ge l - c e li t e c olumn a s u g e qu i va l: e nt s
Tab le 1 9 .
1 72
1 74
The ug e quivale nt t o GA
per gram dry we i gh t o f
3
apple leave s from the e lu t i on o f s i l i c a g e l-c e li te
c olumns b y grad i e n t e lu t i o n w i t h d i f f e rent s o lv e n t s
used
Tab le 20.
1 80
De t e rmian t i on o f c yt ok i nin -like a c t iv i t y i n apple
l e av e s u s i ng the radi s h c o t yl e d o n b i oassay and
puri f i c a t i on b y Se phadex LH-20 and DEAE c e l lu l o s e
c hr o ma t ograp hy
Tab le 2 1 .
1 84
G LC r e t e n t i on t i me s and he i ght of peaks of authen­
t i c plan t hor m o n e s w i th the d i f f e r e n t me t hyl e s t e rs
and me thyl e s t e r s t r i me t hy l s i lyl e ther de r i va t iv e s
Table 2 2.
1 99
A n e xample o f the u s e o f Relat iv e Plant Hormone
I n d i c e s ( RPHI ) t o r e pr e s e n t the b a la n c e of plant
hormone s ; i n this c a s e apple le ave s t aken o n the
1 2 t h of N ov e mb e r
228
xviii
Li s t o f F i gure s
Page
Fig. 1 .
S p e c ulat iv e f l o w chart r e pr e s e n t i ng t h& i n t e gra t i on
b e t w e e n b i o c he mi cal and
Fi g. 2.
y s i o l o g i c a l phenom e na i n the
who l e plant.
20
S p e c ula t iv e s c h e me o f j uv e n i l i t y a n d f l owe r i n g.
22
F i g . 3 -A and B . E f f e c t o f bud s c a l e r e mova l o n t he bud b ur s t
a t t he 1 5 t h January.
A
=
p h o t o graph taken o n 22/2/75.
B
=
p h o t ograph t a ken o n 2 9/2/75 .
28
C and D . E f f e c t o f heav·r ra i n a f t e r drought s e a s o n on
t h e b u d bur s t .
Fig. 4 .
C
=
B u d b ou r s e o f Granny S m i th , pho t ographe d o n 1 3/3/70 .
D
=
Spur b u d s o f Sturm e r P i ppin , pho t ographed on 1 3/3/70.
Exp e r i men t a l t r e e s and d e f i ni t i o n f o r or i e n t at i on and
c la s s i f i c a t i on of who r l g r o ups.
A.
3 year o l d t r e e.
B.
4 year o l d t r e e .
C.
D e f i n i t i o n for o r i e nt a t i on o f t he sho o t s i n t he
34
whorl group .
D.
La t e ral sh o ot; c la s s i f i ca t i on o f pos i t i on ,
o r i e n t a t ion , and w h o r l group.
F i g. 5.
S9asonal c hange o f sh o o t growth i n h o r i zontal a n d
v e r t i cal t r e a t me nts a t d i ff e re nt t i me .
A.
Fig. 6 .
B.
D orma n t pe r i o d bend i ng (Augu s t )
.
Pe tal fall hendi n g ( Oc tob e r )
C.
S e c on d c ov e r hor i z o n ta l ben d i n g ( Novemb e r )
D.
V e r t i c a l t reatme n t ( C on t r o l )
42 -a
E f f e c t o f hori z ontal b e ndi n g on sh o o t gr owth and
f lowe r i ng a t d i f f e r e n t o r i e n ta t i on , whorl , and c lass
of sh o o t s i n 2 9 posi t i o n s o f He d D e l i c ious ( Exp. 1 ) .
F i g . 7-
A.
D orma n t p e r i o d h o riz ontal b e n d i ng ( Augu s t ) .
42
B.
P e t a l fall hor i z o n t a l bend i n g ( O c t ob e r ) .
43
C.
S e c ond c ov e r hor i z o n t a l b e n d i n g ( Novemb e r ) .
44
D.
V e r t i c a l t reatme n t ( C on t r ol ) .
45
The e ffe c t o f ben d i n g f o r R e d D e l i c i ous ( Exp . 1 · and 2 )
a n d Granny S m i t h ( Exp . 3 ) o n sho o t gr owth a n d f l o we r i n g a t t h e whorl group and ori e nt a t i on .
A.
R e d D e l i c i ous ( 2 9 ) i n Exper i m e n t 1 .
B.
R e d D e l i cious ( 45 ) i n Exper i m e n t 2 .
c.
Granny S m i t h
( 2 9 ) i n Exper i m e nt 3.
48
xix
Fi g . 8.
E f fe c t o f p e n du lous be nd ing on sho o t gr owth a n d
f lo we r i ng at d i f f e r e n t o r i e n tat i o n, whor l , a n d
c la s s o f sho o t s in 45 p o s i t i on s o f R e d De l1c i ou s ( Exp . 2 ) .
Fig. 9 .
A.
D or mant p e r i od pendulous b en d i n g ( Au gust )
51
B.
P e t a l fall pendulous b e ndi ng ( Oc t o b e r )
52
C.
S e c ond c ov e r pe ndulous b e nding ( No v e mbe r )
53
D.
V e r t ical t reatment ( C on t r o l )
E f f e c t o f hor i z ontal b e nd i n g o n sh o o t growth and
f l owe r ing at d i f fe r e n t o r ientati o n , whorl, a n d c la s s
o f s h o o ts in 2 9 pos i t i on s o f Granny Smi t h (Exp . 3 ) .
A.
D o r mant p e r i o d hor izontal b e ndi n g ( A ugus t )
57
B.
P e t a l fall h o r i z ontal b e nding ( Oc t ob e r )
58
c.
V e r t i cal t r e a t ment ( C on t r o l )
59
F i g . 1 0 . E f fe c t o f h o r izontal b e nding o n leaf area at d i f f e r e n t
o r i e nta t i on , whorl , a n d c la s s o f s h o o t s i n 2 9 p o sitions
o f Re d D e l i c i ous ( Exp . 1 . ) .
Fig. 1 1 .
A.
D or mant p e r i od hor i z ontal b e nd i n g ( Augus t )
67
B.
P e t a l fall hor i z on t a l b e n d i n g ( Oc t ober )
68
C.
Sec ond c ov e r h o r i z ontal b e nding ( Novemb e r )
69
D.
V e r t i ca l t r e a t ment ( C on t r ol )
70
Re la t i onship o f the s e a s o na l hori z ontal treatme n t s on
the sho o t le n g t h and le a f a r e a .
Fi g . 1 2 .
S h o o t g r ow th a t the d i f fe re n t angle ( A fter E d m i n s t e r ,
76
1 91 7 ) .
Fig . 1 3 .
71
D e n s i t y o f e thyl a c e ta t e i n c h l o r o f or m a n d me thanol in
e th y l a c e ta t e in e a c h 1 0 ml f ra c t i on, leav i n g t h e B
f la sk t o the A f la sk i n gra d i e n t e lu t i o n .
Fig . 1 4 -A .
1 4-B .
Fig . 1 5 .
134
C u t t e r of c ol e op t i le se c t i o n s .
1 40
Turn- table for c o le op t i le bi oa s sa ys .
1 40
D e n s i t y of d i e thyl e t her i n hexane , e thyl a c e ta t e i n
die t hyl e th e r , and m e t ha n o l i n e thyl a c e ta t e i n e a c h
1 0 ml frac t i o n leaving t h e B c yli n d e r f lask t o t he
A c yl in de r .
1 46
14
Fig . 1 7 .
C - IAA a f t e r c e l lu l o s e TLC .
3
Paper and t h i n layer c hroma t o graphy o f H- z e atin .
Fig . 1 8 .
S tandard c a l ib r a t i on o f t h e oat and wheat c ole o p�ile
Fig . 1 6 .
Re c overy of
1 58
167
b i oas s a ys e it h�r s ingly o r c omb ine d in the t e s t tube
f o r the r e s p o n s e t o IAA a n d ABA .
Fig. 1 9.
169
C ompar i s on o f dwa r f r i c e s e e d l i n g s a t d i f f e r e n t c on c e n ­
t ra t i o n s o f GA
me d i um .
3
i n t h e gla s s tub e s a n d agar c ul t u r e
1 71 -a
XX
F i g . 20 .
C ompa r i s o n o f t he l e t tu c e b i oa s say a t d i f f e r e n t
c on c e n tr a t i on s of GA
Fig . 2 1 .
b e t we e n di f f e r e nt varie t i e s
1 73
3
The a c t i v i t i e s o f IAA and ABA - like s ub s tance s in a pple
.a
leaves a f t e r s i l i ca ge l-c e l i t e co lumn , and c e llu l o s e
t h i n laye r chroma t o graphy .
F i g . 22 .
1 75
C e llulose TLC o f g ibbe r e lli n - l ike sub s ta nc e s i n apple
leav e s be for e and a ft e r e lu t in g from t he sys t e m A
s i l icc
F i g . 23 .
ge l- c e l i t e c olumn .
1 77
G i b b e r e l l i n - like sub s tances i n the fra c t i on e lu t e d
from t h e silica ge l - c e l i t e c o lumn w i t h d i ffe r e n t
s o l v e n t s ys t e ms i n t h e gra d i e n t e lu t i o n .
F ig . 24 .
1 81
C ytok i n i n- like activ i tie s a f t e r Se pha d e x LH- 2 0 c olumn
and DEAE c e llulose thin layer chromatography as d e t e c t e d by t he rad i sh c o t yl e d o n b i oassay;
F i g . 25 .
G LC c hroma t ogram of me t hyla t e d plan t mat erials and
c o- c hr o ma t ograms of authe n t i c plant h ormone s .
Fi g . 2 6 .
1 85
1 89
Auxin- and ABA - l ike sub s t a n c e s o f apple leav e s i n t he
a c i d i c frac t i on on c e l lul o s e thin layer chroma t o graphy . 1 9 1
F i g . 27 .
S tandard c a l i b ra t i on o f a u t he ntic p lant hormon e s b y
GLC .
Fig . 2 8 .
A and
1 92 -3
B.
A b e hav i our o f au t he nt i c p lant hormone s
i n c e l lu l o s e thin laye r chr omat ography dete c t e d b y
GLC .
Fig. 29 .
1 96
R e cove r y o f a u t h e n tic plant h ormone s b y GLC f r om
c e l lu l o s e t h i n laye r c hr oma t ography .
Fig . 30 .
1 97
C o mpar i s on o f chroma t o grams in me t h yl e s ters and TMS
d e r i va t ive s of seve ral a u t he n t i c p la n t hormon e s
14
Fi g . 3 1 .
Loss of
F i g . 3 2G
Fra c t i o nat i o n o f ABA in S e phadex G - 1 0 c olumn
Fig . 3 3 .
C yt okinin- like a c t iv i t y in the s h o o t tips of apple
C -IAA in va c uum d r y
1 98
2 04
213
var i e t y S turmer Pippin ( Si l i c a ge l TLC deve lope d b y
wa t e r satur a t e d butano l wit h t wo r e plic at e s )
Fig . 3 4 .
A R e la t ive Plant Hormone I nd e x in t he apple leav e s
taken o n t h e 1 2 th o f N ov e mb e r
Fig . 3 5 .
213
228
A n i n t e rpre t a ti on of Khan's h ypoth e t i c a l mode l i�
s e e d d ormanc y and germina t i o n in t e rms o f Re lativ e
Pla n t Hormone Index
231
xxi
Abbrevia t i o n s
ABA
Absc isic acid
A lar
S u c c i ni c a c i d 2 , 2 - d i me thyl hydraz i de
AW-DMCS
A c i d washe d Dime t hyl d i c h l o r os i la ne
Axs
Auxi ns
BSA
( b i s ( tr ime thyls i lyl ) ac e t a m i de
BSTFA
N . O. -b i s ( tr i�e t hyls i l yl ) tr i flur oac e tami d e
BuOH
n-bu tanol
CKs
C yt ok i n i n s
C HC 1 3
C hl o r o f orm
CH Cl
3
DC - 1 1
Me thyl chloride
S i l i c one gr ease
EDTA
E t hylene d iamine t e t raa c e t i c ac i d
Et OH
E t h yl a l c ohol
Epon 1 00 1 Exoxy r e s i n
GA
G ib b e re ll i n
GAs
G i b b e r e llins
GLC
G a s l i qu i d c hromat ography
HC l
Hydr o c h l or i c a c i d
HIEFF= 8BP C y c l ohexane d i me tha n o l A p i pa t e
HJviDS
Hexa me t hyl d i s i lazane
IAA
I n d olyl - 3 - a c e t i c a c i d
IAA p
Ind o lyl-a spa r t a t e
ICA
I n d o lyl - 3 - c ar b oxy l i c a c i d
ILA
DL - 3 - ( 3 - i ndo lyl ) la c t i c a c i d
IPA
3 - ( 3 - i nd olyl ) - pr opi on i c a c i d
I PA
6 -(3 -me thyl-2- bute nylam in o ) ad e n o s ine ( pp. 2 0 7 )
msiPA
6 - ( 3 - me thyl- 2-b u t e n ylamin o ) -2 - me thylthi oa d e n o s i n e
I p yA
Ind o le p yruvi c a c i d
I
I od i ne
Ib s
I nhib i t or s
K HP04
2
KI
P o t a s s ium phospha te
KOH
P o t a s s ium hyd�ox ide
LSD
Lea s t S i gni fi cant D i f f e r e n c e
MAA W
Me thyl a c e ta t e : A c e t on i t ri l e : Ammonium hydr oxide: wa t e ::·
Me OH
Me thyl a l c ohol
NAA
Naphtha l e ne ac e t i c a c i d
Na c o 3
2
A nh ydro us s o dium c a r b o na t e
P o ta s s ium iodi de
xxii
Na H C 03
S o d iu m b i ca rb o na t e
NaOH
S odium hyd rox i d e
Na 2so4
Na s o
2 2 5
NH oH
4
A nh ydr ous s o d iu m su lpha t e
OV-1
M e thyl s i l i co n e
PYR
P yri d i n e
PVP
P ol yv in ylpyr r o l i d one
QF-1
5 0% t r if lu r opropyl me t h y l s il i c on e
SE-30
M e th y l s il i c one
SE-52
M e th yl s il i c o ne
TLC
T h i n la yer c h r o ma t o�ra p h y
TMCS
T r i me t h yl c hl or o s ilane
TNH
Trypta m in e
S o d ium pyr o s u lfa t e
Ammonium hydr oxi de
2
OH
T
T r yp t ophol
t-RNA
t ra n sf e r r ib onu cl e i c a c i d
TTP
T r yp t opha n
UC-W98
U nkn own % v i nyl methyl s i l i c o n e
V
V o lu me