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LEAF  AND  STEM  ANATOMY  THEIR  OF  SEVERAL  COMPACT  ALAR  APPLE  MUTANTS,  TREATED  CULTIVARS,  AND  PLANTS  by ALICE  CHEN-MIAO  LIU  B.S.A., Taiwan P r o v i n c i a l Chung-Hsing U n i v e r s i t y ,  A  THESIS THE  SUBMITTED  IN  REQUIREMENTS MASTER  PARTIAL FOR  OF  THE  FULFILMENT DEGREE  OF  SCIENCE  IN THE DEPARTMENT of PLANT SCIENCE  We accept t h i s t h e s i s as to the r e q u i r e d standard  THE  UNIVERSITY  OF  March, 1970  conforming  BRITISH  COLUMBIA  1967  OF  In p r e s e n t i n g t h i s t h e s i s  in p a r t i a l  f u l f i l m e n t o f the requirements  an advanced degree at the U n i v e r s i t y of B r i t i s h C o l u m b i a , I agree the L i b r a r y  s h a l l make i t f r e e l y  I f u r t h e r agree tha  available for  r e f e r e n c e and  permission for extensive copying of t h i s  It  i s understood that copying o r  thesis  permission.  Department The U n i v e r s i t y o f B r i t i s h Columbia Vancouver 8, Canada  or  publication  o f t h i s t h e s i s f o r f i n a n c i a l g a i n s h a l l not be a l l o w e d w i t h o u t my written  that  study.  f o r s c h o l a r l y purposes may be g r a n t e d by the Head of my Department by h i s r e p r e s e n t a t i v e s .  for  ABSTRACT  Naturally occurring  compact (spur  type) a p p l e t r e e s  s y l v c s t r i s L.) were compared w i t h s t a n d a r d and A l a r Stem anatomy r e c e i v e d  (Malus  treated  trees.  s p e c i a l a t t e n t i o n because no comparisons had  been done o f the stem anatomy Anatomical studies of leaves  i n s t a n d a r d and compact a p p l e i n compact mutants and i n A l a r  Red D e l i c i o u s a r e more d e t a i l e d than i n e a r l i e r Four c u l t i v a r s were used i n the s t u d i e s .  trees. treated  reports. They were H a r r o l d  Red  D e l i c i o u s , a compact mutant o f Red D e l i c i o u s , namely S t a r k r i m s o n , and s t a n d a r d Golden D e l i c i o u s , and i t s compact mutant S t a r k s p u r .  There  were t h r e e main s t u d i e s i n t h i s i n v e s t i g a t i o n . I n the f i r s t study, a n a t o m i c a l e x a m i n a t i o n s were made on the f o u r c u l t i v a r s without A l a r treatment.  S t a r k s p u r Golden D e l i c i o u s was  to have the t h i c k e s t l e a f and p a l i s a d e tivars  studied.  palisade  The compact type was  parenchyma and g r e a t e r  w i t h the s t a n d a r d  parenchyma among the f o u r found to have a t h i c k e r  mean p a l i s a d e  cul-  leaf,  number when compared  type.  The e f f e c t of A l a r a t c o n c e n t r a t i o n s same c u l t i v a r s was i n v e s t i g a t e d of t e r m i n a l  found  o f 0 and 1000 ppm on the  i n the second study.  growth by A l a r v a r i e d among the c u l t i v a r s .  The  suppression  The response  to A l a r was g r e a t e s t w i t h S t a r k s p u r and 50 per cent i n h i b i t i o n of shoot growth was o b s e r v e d . atment.  Microscopic  S t a r k r i m s o n was n o t a f f e c t e d by A l a r  examination revealed  cant d i f f e r e n c e s i n c e l l  t h a t t h e r e were no  tre-  signifi-  l e n g t h of c o l l e n c h y m a , parenchyma and  pith  ii c e l l s o r i n c e l l d i a m e t e r s and t i s s u e t h i c k n e s s taken from the f i r s t I n study t h r e e ,  when the samples were  i n t e r n o d e under the growing t i p . the e f f e c t o f A l a r and i t s i n t e r a c t i o n w i t h  g i b b e r e l l i c a c i d on Red and Golden D e l i c i o u s were c o n s i d e r e d .  In this  study, comparisons were a l s o made w i t h the u n t r e a t e d compact mutants. Alar  t r e a t m e n t s o f Red D e l i c i o u s were found t o i n c r e a s e  total  l e a f , spongy parenchyma and the l e n g t h  latter  two a c c o u n t e d f o r the i n c r e a s e  t r e a t e d Red D e l i c i o u s Gibberellic acid and  of palisade  i n t o t a l thickness  cells.  The  of Alar-  leaves. stimulated  t h e shoot growth o f Golden  S t a r k s p u r by 29 per c e n t , b u t t h i s s t i m u l a t i n g  t e d by A l a r .  thickness of  Delicious  e f f e c t was preven-  ACKNOWLEDGEMENTS  I w i s h t o express my deepest g r a t i t u d e t o Dr. G. W. Eaton, Associate  Professor,  Department o f P l a n t S c i e n c e ,  University of  B r i t i s h Columbia, under whose s u p e r v i s i o n t h i s p r o j e c t was undertaken, f o r h i s t e c h n i c a l advice the p r e p a r a t i o n  during  the r e s e a r c h ,  and f o r h i s guidance i n  of t h i s thesis.  S i n c e r e a p p r e c i a t i o n i s e s p e c i a l l y expressed t o Dr. N. E. Looney, P o m o l o g i s t , Canada Department o f A g r i c u l t u r e R e s e a r c h S t a t i o n , Summerland, B r i t i s h Columbia, who k i n d l y provided  many o f the p l a n t  m a t e r i a l s used i n t h i s p r o j e c t and who a l s o gave v a l u a b l e assistance  counsel and  i n s e v e r a l ways.  A l s o a p p r e c i a t i o n i s extended t o Drs. K. Beamish, C. A. Hornby, and V. C. R u n e c k l e s f o r t h e i r h e l p f u l s u g g e s t i o n s d u r i n g  this  endeavor.  T h i s Research was supported by NRCC O p e r a t i n g Grant A2023 awarded to Dr. G. W. Eaton.  V  TABLE  OF  CONTENTS Page  I. II. III. IV.  INTRODUCTION LITERATURE MATERIALS  1  REVIEW AND  .'  3  METHODS  12  RESULTS Experiment  20 1:  Comparisons  o f U n t r e a t e d Compact  and Standard A p p l e C u l t i v a r s Experiment  11:  .  .  .  E f f e c t s o f A l a r on Compact and  Standard A p p l e C u l t i v a r s Experiment  111 ( a ) :  Comparisons  30 o f Compact  w i t h A l a r t r e a t e d Red D e l i c i o u s Experiment  20  38  111 ( b ) : E f f e c t s o f A l a r and  G i b b e r e l l i c A c i d on Compact and Standard Golden D e l i c i o u s V. VI.  43  DISCUSSION  48  SUMMARY  53  BIBLIOGRAPHY  55  APPENDIX  59  vi LIST  OF  TABLES  Table  Page  1.  T h i c k n e s s o f L e a f T i s s u e s o f Four C u l t i v a r s  21  2.  T h i c k n e s s o f L e a f T i s s u e s of.Red D e l i c i o u s and Golden D e l i c i o u s  3.  .  25  T h i c k n e s s o f L e a f T i s s u e s of Standard and Compact Types  4.  26  C e l l Lengths o f Stem Collenchyma, Parenchyma and P i t h o f Four Apple C u l t i v a r s  5.  T h i c k n e s s o f Stem C o r t e x , V a s c u l a r T i s s u e ,  29 Diameters  of P i t h and Stem o f Four Apple C u l t i v a r s 6.  31  C e l l Diameters o f Stem Collenchyma, Parenchyma and P.ith o f Four A p p l e C u l t i v a r s  7.  32  E f f e c t o f A l a r on Shoot Growth of Red D e l i c i o u s and S t a r k r i m s o n  8.  33  E f f e c t o f A l a r on L e a f Number o f Red D e l i c i o u s and S t a r k r i m s o n  9.  34  E f f e c t o f A l a r on Average L e a f Length and Width of Red D e l i c i o u s and S t a r k r i m s o n  10.  35  I n f l u e n c e o f A l a r on Shoot Growth o f S t a r k s p u r and Golden D e l i c i o u s  .  11.  E f f e c t o f A l a r on L e a f Number o f Golden D e l i c i o u s  12.  I n f l u e n c e o f A l a r on T h i c k n e s s of L e a f T i s s u e s of Four A p p l e C u l t i v a r s  3§  a  .  .  .  37  39  vii Table 13.  Page Influence  of A l a r on  Parenchyma and 14.  Influence Tissues  15.  E f f e c t of A l a r on and  16.  Red  Influence  Influence  40  the T h i c k n e s s of L e a f  Delicious  41  Shoot Growth of  Starkrimson  Delicious of A l a r and  of L e a f T i s s u e s 17.  P i t h of Four A p p l e C u l t i v a r s  of A l a r on of Red  the C e l l Lengths of Collenchyma,  45 G i b b e r e l l i c A c i d on  of S t a r k s p u r and  of A l a r and  Thickness  Golden D e l i c i o u s  . . .  46  G i b b e r e l l i c A c i d on Mean  Shoot Growth of S t a r k s p u r and  Golden D e l i c i o u s  . . . .  47  viii LIST  OF  FIGURES  Figure  Page  1.  T h i c k n e s s of L e a f T i s s u e s of Four Apple C u l t i v a r s  2.  The L e a f C r o s s S e c t i o n of S t a r k s p u r Showing Longer  .  .  .  .  22  P a l i s a d e C e l l s and G r e a t e r Mean P a l i s a d e Number Than i n the Standard 3.  Golden D e l i c i o u s  23  The L e a f C r o s s S e c t i o n of S t a r k r i m s o n Showing G r e a t e r L e a f T h i c k n e s s Than i n Red D e l i c i o u s  4.  24  L o n g i t u d i n a l S e c t i o n o f Red D e l i c i o u s Stem Showing Collenchyma,  Parenchyma, V a s c u l a r T i s s u e and  P a r t of P i t h 5.  C r o s s S e c t i o n of Red D e l i c i o u s Stem Showing P a r t o f  6.  Collenchyma, Parenchyma, V a s c u l a r T i s s u e and P i t h Red D e l i c i o u s Leaves T r e a t e d w i t h A l a r a t 4000 ppm Were T h i c k e r Than U n t r e a t e d Leaves  27  . . .  28  42  INTRODUCTION  Due t o the c o s t o f l a b o r i n p r u n i n g and h a r v e s t i n g on  l a r g e t r e e s , c o n t r o l l i n g the s i z e o f f r u i t  fruit  t r e e s has h e l d the  i n t e r m i t t e n t i n t e r e s t o f pomologists f o r hundreds o f y e a r s .  A  r e c e n t approach t o a t t a i n i n g t r e e s i z e c o n t r o l has i n v o l v e d growth r e t a r d a n t s . trees  One showing c o n s i d e r a b l e  grown  chemical  promise f o r use on f r u i t  i s s u c c i n i c a c i d 2,2-dimethyl h y d r a z i d e commercially known and  h e r e a f t e r r e f e r r e d t o as A l a r .  The f i r s t p u b l i s h e d  of t h i s c h e m i c a l as a growth r e t a r d a n t  r e p o r t of the use  on p l a n t s was made by R i d d e l l  et a l . (31) i n 1962. Studies been r e p o r t e d  on the movement and f a t e of A l a r i n a p p l e s e e d l i n g s (25).  Other s t u d i e s have shown t h a t A l a r a p p l i e d to  a p p l e t r e e s caused many d e s i r a b l e e f f e c t s without a major of r o o t growth ( 3 ) .  bloom one to t h r e e days and has a l s o been r e p o r t e d  fruit  s e t ( 1 4 ) , improve f r u i t - k e e p i n g q u a l i t y ( 1 4 ) , reduce  drop ( 1 4 ) , and i n c r e a s e  fruit  to increase pre-harvest  c o l o r ( 2 1 , 23, 3 3 ) . Treatments  r e s u l t i n s m a l l e r and f i r m e r f r u i t  (5) and greener and t h i c k e r  often leaves  16, 17). I n s p i t e of such wide-spread a t t e n t i o n on a d i v e r s i t y o f  subjects,  there  is little  detailed information  on a n a t o m i c a l e f f e c t s o f  A l a r (16) o r a n a t o m i c a l comparisons between s t a n d a r d The and  suppression  A l a r suppresses t e r m i n a l growth ( 3 , 4, 14, 16),  delays  (14,  have  object  standard  of the p r e s e n t  and compact  types.  study was to f u r t h e r compare compact  types and the e f f e c t s o f a l a r on the morphology  . 2 and anatomy of a p p l e leaves and stems u s i n g s t a n d a r d and compact s t r a i n s of Red D e l i c i o u s and Golden D e l i c i o u s a p p l e s .  LITERATURE  The  REVIEW  development of p l a n t growth r e t a r d a n t s has  s i n c e the r e p o r t i n 1949  t h a t a new  been v e r y r a p i d  c l a s s of c h e m i c a l s ,  t i n i u m s , reduced stem e l o g a t i o n of bean p l a n t s without changes.  The  most a c t i v e compound was  When a p p l i e d i n one  per cent  l a n o l i n paste,  the f i r s t  found to be one-quarter of the  control (28).  year  a number of quaternary  l a t e r , Wirwile  other  internode  l e n g t h of  of  the  the  and M i t c h e l l (39) r e p o r t e d  that  ammonium carbamates r e t a r d e d the growth and  development of a broad v a r i e t y of p l a n t s p e c i e s without ment of malformed  formative  2,4-dichlorobenzyInicotinium.  t r e a t e d p l a n t s was One  the n i c o -  l e a v e s , stems, r o o t s and  flowers.  the  develop-  4-Hydroxy-5~iso~  propyl-2-methylpheny1 t r i m e t h y l ammonium c h l o r i d e , 1 - p i p e r i d i n e c a r boxy l a t e (Amo-1618) was group of chemicals  found to be  tested.  the most a c t i v e compound i n t h i s  I n 1958,  2,4-dichlorobenzyl-tributylphosphonium the growth of more w i d e l y in  1960  (2-chloroethy1)  P r e s t o n and L i n k (30)  found  c h l o r i d e (Phosfon) a f f e c t e d  d i f f e r e n t species  than d i d Amo-1618.  trimethylammonium c h l o r i d e (CCC)  was  to r e t a r d the growth of a l a r g e r number of s p e c i e s than any e a r l i e r compounds ( 3 6 ) . sprays  I n 1962,  that  R i d d e l l e t a l . (32)  Then  found of  reported  the that  of N-dimethylamino maleamic a c i d (CO 11) r e t a r d e d the growth of  legumes, v i n e c r o p s , potatoes  and  C011  i n aqueous s o l u t i o n , i t s analogue,  was  found to be u n s t a b l e  ornamental p l a n t s .  dimethylaminosuccinamic a c i d (B995), was  s t a b l e and  However, whereas  retarded  the  N-  growth of the B995 was discoverer, United B-9,  same s p e c i e s  as d i d C011  the o r i g i n a l e x p e r i m e n t a l code number g i v e n by  Uniroyal,  ornamentals and  Later  the name was  O r i g i n a l l y the m a t e r i a l was  the  shortened to B-Nine, i n t e n d e d f o r use  s o l d f o r t h i s purpose under the name of B-Nine.  l a t e r commercial p r e p a r a t i o n , species  the  then the Naugatuck Chemical D i v i s i o n of  S t a t e s Rubber Company.  DMAS and A l a r .  several  (13).  of f r u i t  Alar-85,  crops.  The  i s r e g i s t e r e d f o r use  on The  on  c h e m i c a l s t r u c t u r e i s as  follows:  0  II CHo  -  I  CH  -  2  C C  - NH  -  N' CHr;  -  OH  II 0  Among the growth r e t a r d a n t s  tested, Alar  been s t u d i e d by many i n v e s t i g a t o r s . sweet c h e r r i e s , a p p l e s e e d l i n g s have been s t u d i e d  (33,  new  sweet c h e r r y ,  leaves  of the  26).  and  the  Martin  i n P h a r b i t i s plants et al.  seedlings. resistant  (26)  were a b l e  movement and  in  f a t e of A l a r  short-day p l a n t , P h a r b i t i s n i l , r e s i d u a l A l a r was  Prunus avium, i n the  spring  (40).  persistence  following  A l s o , by  to f o l l o w  using  labeled A l a r ,  the movement of A l a r i n a p p l e  to breakdown i n the p l a n t and t r a n s p i r a t i o n stream.  was  of t h i s growth r e -  From c h e m i c a l a n a l y s i s they concluded t h a t i t  r a p i d l y i n the  found i n  With r a d i o a c t i v e B995, Z e e v a a r t  to demonstrate the m o b i l i t y and  tardant  has  Ryugo r e p o r t e d  a l a t e f a l l a p p l i c a t i o n (33). able  The  seems v e r y p r o m i s i n g , and  was  Due  absorbed and  was translocated  to i t s r a p i d a b s o r p t i o n  and  5, h i g h m o b i l i t y , coverage should be o f l e s s importance and a more approach t o a p p l i c a t i o n may be i n o r d e r . over many o t h e r  chemical  casual  T h i s i s an advantage o f A l a r  sprays.  Once w i t h i n the p l a n t , A l a r causes a number o f e f f e c t s .  The  e f f e c t on r o o t growth of one year o l d a p p l e t r e e s was s t u d i e d by Barden (3) who r e p o r t e d retardants  t h a t the m e r i t o f A l a r over some o t h e r  growth  i s t h a t A l a r causes many d e s i r a b l e e f f e c t s on the above  ground p o r t i o n s of an a p p l e t r e e w i t h o u t a major s u p r e s s i o n of r o o t growth.  Other workers have s t u d i e d the e f f e c t o f A l a r on  Zeevaart reported  t h a t treatment o f the short-day  B995 r e s u l t e d i n s h o r t , t h i c k i n t e r n o d e s the (16)  plant P h a r b i t i s with  The e f f e c t of A l a r on  shoot diameter o f a p p l e t r e e s has a l s o been noted. reported  t h a t A l a r treatment i n c r e a s e d  Golden D e l i c i o u s and Y o r k I m p e r i a l  apples.  former was due t o an i n c r e a s e i n r a d i a l and  (40).  cortex.  thicker.  F o r York I m p e r i a l ,  Longitudinal  shoots.  Halfacre et a l .  stem r a d i u s o f both The i n c r e a s e d  thickness  r a d i u s i n the  o f the p i t h ,  i t was due to p i t h and c o r t e x  phloem being  sections of A l a r - t r e a t e d plants of both apple  c u l t i v a r s had fewer and s h o r t e r c e l l s per i n t e r n o d e . was a f f e c t e d more than c e l l  Cell  division  expansion t r a n s v e r s e l y and l o n g i t u d i n a l l y .  S e v e r a l workers have noted e f f e c t s of A l a r on f l o w e r i n g o f f r u i t trees.  B a t j e r et al. (4) r e p o r t e d  t h a t a p p l e and c h e r r y  trees  w i t h A l a r i n the e a r l y summer o f 1962 produced more f l o w e r s than unsprayed t r e e s . " V i o l e t " , flower  sprayed  i n 1963  However, i n the short day p l a n t P h a r b i t i s n i l ,  formation  was i n h i b i t e d by the a p p l i c a t i o n o f A l a r  6 via  the r o o t s  night  (40).  f o r a period  o f 24 hours p r i o r to one i n d u c t i v e  E d g e r t o n e t a_l. r e p o r t e d  promoted on t h r e e year o l d D e l i c i o u s  long  t h a t f l o w e r bud f o r m a t i o n was trees  sprayed w i t h B995.  The  pre-bloom a p p l i c a t i o n o f B995 on mature t r e e s d e l a y e d bloom one to t h r e e days and r e s u l t e d i n h i g h e r f r u i t c o n t r o l when f r o s t s o c c u r r e d f o l l o w i n g reports  indicated  fruit.  E d g e r t o n _et aj.. found t h a t  Delicious  trees  s e t as compared w i t h unsprayed the treatments (14).  Several  t h a t A l a r a l s o a f f e c t e d k e e p i n g q u a l i t y o f the sprays of B995 on three year o l d  e a r l y i n the growing season reduced f r u i t  size at  h a r v e s t (14) and p r e - h a r v e s t s p r a y s o f B995 to more mature M c i n t o s h apple trees resulted  i n firmer  fruit  than on u n t r e a t e d p l a n t s .  Other  workers have demonstrated the e f f e c t s o f A l a r on enhancing a p p l e q u a l i t y a t h a r v e s t and a f t e r storage ( 1 5 ) . When A l a r was a p p l i e d a t a c o n c e n t r a t i o n cherry the  o f 2000 ppm to sweet  Prunus avium, e a r l y p r o d u c t i o n of the a n t h o c y a n i n pigments i n  f r u i t was observed and Ryugo (33) concluded t h a t a l t h o u g h A l a r  enhanced the b i o s y n t h e s i s  o f a n t h o c y a n i n s , i t d i d not measurably  advance the p h y s i o l o g i c a l m a t u r i t y o f c h e r r i e s . reports  that e a r l y  season a p p l i c a t i o n of A l a r  meters o f sweet c h e r r y The  Looney, however,  promotes s e v e r a l  para-  maturity (24).  e f f e c t of post-bloom a p p l i c a t i o n of A l a r on a p p l e  r i p e n i n g has a l s o been s t u d i e d .  Looney (21) r e p o r t e d  that  fruit the amount  of c h l o r o p h y l l i n p e e l and f l e s h o f a p p l e was lower through the season when a spray o f 4000 ppm of A l a r was a p p l i e d  i n mid-May, two weeks  a f t e r bloom but he delay maturity.  concluded t h a t A l a r d i d not n o t i c e a b l y advance or  He  also  studied  under storage  c o n d i t i o n s and  2000 ppm  significantly  Alar  the r e s p i r a t o r y b e h a v i o r of a p p l e s  found t h a t a m i d - J u l y a p p l i c a t i o n of reduced r e s p i r a t i o n of s t o r e d f r u i t  0° C, whereas, a mid-May 2000 ppm r e p o r t from the was  d e l a y e d by  same l a b o r a t o r y ,  spray, d i d not  treatments of A l a r a p p l i e d two  Looney suggests t h a t A l a r  w i t h i n the f r u i t and maturity  (23).  One  this  counteracted  ppm  of  ethy-  biosynthesis  r e l a t e d to  B a t j e r _et aJL.  drop.  fruit  drop and  reported  that  Winesap a p p l e t r e e s reduced  d e l a y e d the development of w a t e r c o r e .  Treated  somewhat lower i n s o l u b l e s o l i d s ' ( 5 ) .  et aJL. thought A l a r might have c e r t a i n s p e c i f i c e f f e c t s  than a g e n e r a l  e f f e c t on the  fruit  (8) because they found  the a p p l i c a t i o n of A l a r to a p p l e t r e e s i n c r e a s e d delayed best harvest do  100  and  of the most noteworthy e f f e c t s of A l a r i s i t s  f r u i t s were f i r m e r and  rather  by  suppressed e t h y l e n e  the a p p l i c a t i o n of A l a r to D e l i c i o u s and  Blanpied  later  weeks a f t e r bloom,  suppression.may not be  e x c e l l e n t c o n t r o l " of p r e - h a r v e s t  pre-harvest  In a  t h e . r i p e n i n g of M c i n t o s h a p p l e s  t h i s i n h i b i t o r y e f f e c t of A l a r was lene.  (21).  at  fruit  firmness  date f o r three v a r i e t i e s i n I r e l a n d , but  so f o r M c i n t o s h i n New  York.  They a t t r i b u t e the  that and  did  not  conflicting  r e s u l t s to d i f f e r e n c e s i n the i n t e r a c t i o n of v a r i e t y , season, l o c a t i o n and  the  s p e c i f i c e f f e c t s of the  Various ported.  e f f e c t s of A l a r on  material. l e a f c h a r a c t e r i s t i c s have been r e -  Edgerton and Hoffman r e p o r t e d  spray to t h r e e - y e a r - o l d  t h a t A l a r a p p l i e d as a  D e l i c i o u s t r e e s i n mid-June produced  foliar leaves  8 normal and,  i n some cases,  d a r k e r green and  Barden i n v e s t i g a t e d the  concentrations.  appeared leaves  arrangement of the leaves,  production  and  same workers r e p o r t e d  s t u d i e d i n other All  leaves of  treated  stimulated  a l s o f r e s h and  E f f e c t s of A l a r on  c u l t i v a r s such as Red  because of the i n t i m a t e  transverse  (16).  leaves  found t h a t the  thickness  r e l a t e d to the p h o t o s y n t h e t i c  photosyn2  (g/cm  fresh cells  respira-  a l s o r e l a t e d to the  growth  (7).  e f f e c t of a growth s t i m u l a n t , i n p l a n t s has  and  and of  Beakbane concluded t h a t the number of p a l i s a d e  t o r y a c t i v i t y of the p a l i s a d e m e s o p h y l l and  elongation  be  leaf structure  McClendon s t u d i e d the  a f u n c t i o n of t h e i r d e n s i t y  p o t e n t i a l of a p p l e r o o t s t o c k s  area  orchard  ,  per u n i t l e a f s u r f a c e was  on  Delicious.  have s i g n i f i c a n c e i n the  rate.  and  Later,  l e a f anatomy should  twenty-three d i f f e r e n t s p e c i e s of p l a n t and  The  palisade  dry weights per u n i t  r e l a t i o n s h i p between the  i t s maximum p h o t o s y n t h e t i c  weight) (27).  looser  I n Golden D e l i c i o u s  used on York I m p e r i a l  the above f i n d i n g s may  t h e t i c r a t e was  a  t h a t A l a r treatment decreased l e a f area  a p p l e t r e e s and  of l e a f t i s s u e (17).  treated  expansion whereas A l a r i n h i b i t e d c e l l d i v i s i o n  expansion a t a l l c o n c e n t r a t i o n s  York I m p e r i a l  of A l a r  apple  p a l i s a d e c e l l s and  spongy parenchyma c e l l s .  the lower c o n c e n t r a t i o n s  (14).  l e a f a n a t o m i c a l responses of  They found the  p l a n t s were t h i c k e r as a r e s u l t of longer  the  leaves  year o l d t r e e s of Golden D e l i c i o u s and York I m p e r i a l  with A l a r at various  cell  The  t h i c k e r i n texture, than the u n t r e a t e d  H a l f a c r e and one  larger i n size.  g i b b e r e l l i c a c i d (GA),  o f t e n been i n v e s t i g a t e d .  As  on  t h e r e was  stem no  evidence of c e l l  elongation  f o r a t l e a s t 72 hours a f t e r a p p l i c a t i o n  of g i b b e r e l l i n to the v e g e t a t i v e , p l a n t s of the b i e n n i a l short-day Hyoscyamus and  of  t h a t the i n i t i a l in  the  long-day p l a n t Salmolus, Sachs _et al.  increase  c e l l number (34).  i n stem l e n g t h was  due  Beakbane suggested t h a t GA  p o s s i b l e expanding agent i n l e a f t i s s u e subjected (6), 0.5  because i n a p p l e l e a f d i s c s t r e a t e d ppm  GA,  great  d i s t r i b u t i o n and  concluded  s o l e l y to an might be  used as a  to shade  i n a b a s i c medium  expansion of the epidermal c e l l s was  increase  conditions plus  observed.  shape of m e s o p h y l l c e l l s were a l s o a f f e c t e d by  The GA.  S e v e r a l workers have examined the i n t e r a c t i o n of A l a r w i t h GA plants.  Z e e v a a r t found the  i n h i b i t i o n of flower  P h a r b i t i s n i l by A l a r c o u l d be g i b b e r e l l i n A3  to the  cucumber s e e d l i n g s , Moore (29) GA3  applied  to the  was  shoot t i p was  e f f e c t of 25/ug of A l a r a p p l i e d old 1000  able  long n i g h t  a p p l i c a t i o n of  (40).  By  to demonstrate t h a t  ppm  and  s u f f i c i e n t to completely n u l l i f y s i m u l t a n e o u s l y (29).  of potassium g i b b e r e l l a t e (KGA)  reduced to l e s s than 50 per  t r e a t e d w i t h KGA  a l o n e (14).  a c t i o n between A l a r and Further GA  the  When  The  a t 200  ppm,  the  three-year-  the  cent of t h a t made by  at shoot  shoots  However, the r e a l mechanism of i n t e r gibberellins s t i l l  s t u d i e s are needed to e l u c i d a t e the i n the  using  1 / i g of  D e l i c i o u s a p p l e t r e e s were sprayed w i t h a m i x t u r e of A l a r  growth was  and  formation i n  completely overcome by  plumule b e f o r e the  in  remains unknown.  r e l a t i o n s h i p between A l a r  apple.  d i f f e r e n c e i n growth h a b i t ,  chemical c o n t e n t , and  p o s i t i o n between S t a r k i n g D e l i c i o u s and  the n a t u r a l  l e a f com-  compact mutant  10 S t a r k r i m s o n have been s t u d i e d e x t e n s i v e l y .  I n g e n e r a l , when compared  w i t h S t a r k i n g , the mutant i s r e p o r t e d to have s h o r t e r s h o o t s ,  fewer  l a t e r a l s h o o t s , more f r u i t i n g s p u r s , more nodes per f o o t and t h i c k e r leaves w i t h a g r e a t e r depth of p a l i s a d e parenchyma, more dry weight, c h l o r o p h y l l , N, and Ca ( 1 , 37, 3 8 ) . l i z e about the a n a t o m i c a l  B e f o r e i t i s p o s s i b l e to g e n e r a l  d i f f e r e n c e s between s t a n d a r d and compact  t y p e s , comparisons must be made i n o t h e r c u l t i v a r s D e l i c i o u s and i t s compact mutant, S t a r k s p u r . s i d e r e d these  such as Golden  Only A r a s u  (1) has con-  l a t t e r two s t r a i n s and he o n l y r e p o r t s data f o r t o t a l  l e a f and p a l i s a d e parenchyma t h i c k n e s s .  Stem anatomy  comparisons  between compact and s t a n d a r d growing types have not been made i n any cultivar. S i n c e the n a t u r a l compact mutants have many m e r i t s over the s t a n d a r d growing s t r a i n s and s i n c e A l a r i s r e p o r t e d to cause s i m i l a r changes i n s t a n d a r d a p p l e t r e e s , a c r i t i c a l  comparison of the d i f f e -  rences and s i m i l a r i t i e s between these two approaches to s i z e c o n t r o l i s required.  I n t r i e r i r e p o r t e d t h a t spur type t r e e s and those t r e a t e d  w i t h r e t a r d a n t s showed many a n a l o g i e s i n morphology, p h y s i o l o g y , b i o c h e m i s t r y and anatomy  ( 1 9 ) . T h i s was based on an e x t e n s i v e  review and not c r i t i c a l  literature  comparisons w i t h i n any s i n g l e experiment.  This  r e p o r t t h e r e f o r e must be regarded as s e t t i n g up a number of hypotheses which demand i n v e s t i g a t i o n . larities  L i k e w i s e , Looney r e p o r t e d t h e r e were s i m i -  between compact mutants and s t a n d a r d D e l i c i o u s a p p l e t r e e s  t r e a t e d w i t h A l a r ( 2 2 ) . He found S t a r k r i m s o n was a p p r o x i m a t e l y  t h a t the net a s s i m i l a t i o n r a t e of  twelve p e r cent h i g h e r than t h a t of  S t a r k i n g D e l i c i o u s and a s i m i l a r d i f f e r e n c e was S t a r k s p u r and Golden D e l i c i o u s ( 2 2 ) .  r e v e a l e d between  According retardant  to the  studied  t i c s such as biochemical concerning  l i t e r a t u r e , A l a r i s the most p r o m i s i n g growth  to d a t e .  While i t s e f f e c t s on c e r t a i n c h a r a c t e r i s -  growth h a b i t , e x t e r n a l appearance, morphology f u n c t i o n have been s t u d i e d , t h e r e i s no  of  main p o i n t s a r i s i n g from t h i s  stem anatomy i n Red l e a f and  Delicious. and  geneticccompossible  l i t e r a t u r e review w i l l  be  D e l i c i o u s and and  leaf  d e t a i l e d a n a t o m i c a l comparisons  compact types of Red  and  Golden  While some a n a t o m i c a l e f f e c t s of A l a r on Golden D e l i c i o u s l e a f anatomy i n compact and  D e l i c i o u s have a l r e a d y  been r e p o r t e d ,  standard  Golden and  Red  these w i l l be i n c l u d e d i n the  study to a l l o w more d i r e c t comparison w i t h the r e s u l t s of  earlier studies. any  The  time i n t h i s study; e f f e c t s of A l a r on  stem i n standard  comparisons of  present  at a l l  c a r e f u l study.  i n v e s t i g a t e d f o r the f i r s t and  and  t o o l s f o r s i z e c o n t r o l would i n d i c a t e t h a t any  i n t e r a c t i o n merits Two  information  the e f f e c t of A l a r on the n a t u r a l compact mutants.  h o r t i c u l t u r a l importance of both growth r e t a r d a n t s p a c t i o n as  and  Previous  compact mutant and  superficial.  With t h i s  workers have not  l e a f anatomy s t u d i e s information  s t u d i e d stem anatomy i n to date have been r a t h e r  i t should  be p o s s i b l e to  generalize  about the a n a t o m i c a l e f f e c t s of A l a r or n a t u r a l compaction upon a p p l e varieties.  MATERIALS  AND  METHODS  Experiment I Comparisons  of U n t r e a t e d Compact and Standard C u l t i v a r s  The l e a f and  shoot samples of a p p l e t r e e s were o b t a i n e d from the  o r c h a r d of the Canada Department at  Summerland.  Two  of A g r i c u l t u r e Research S t a t i o n  v a r i e t i e s , Red D e l i c i o u s and Golden D e l i c i o u s ,  w i t h s t a n d a r d and compact t y p e s , i . e . H a r r o l d Red D e l i c i o u s  each  (standard),  S t a r k r i m s o n (compact), Golden D e l i c i o u s ( s t a n d a r d ) and S t a r k s p u r (comp a c t ) , were used i n t h i s experiment. old  t r e e s were randomly  On June 11, 1968,  chosen from each c u l t i v a r .  North and South s i d e o f each t r e e was  One  shoot from the  sampled by t a k i n g two n e i g h b o r i n g  l e a v e s from the middle p a r t of the c u r r e n t y e a r ' s shoot. was  t h r e e nine year  Each  sampled by t a k i n g d i s c s from t h r e e d i f f e r e n t p o s i t i o n s ,  middle and b a s a l on each s i d e o f the m i d r i b . first  i n t e r n o d e below  the shoot t i p was  l e a f and  shin F l u i d  apical,  From the same shoot, the  taken.  Each i n t e r n o d e was  t r a n s v e r s e l y i n t o two p a r t s f o r b o t h c r o s s and l o n g i t u d i n a l The  S o l u t i o n B:  cut  sections.  shoot t i s s u e s were f i x e d i n B e l l i n g s M o d i f i e d Nava-  (20) f o r which the formula i s as f o l l o w s :  S o l u t i o n A:  leaf  Chromic a c i d c r y s t a l s  5g  Glacial acetic acid  500cc  D i s t i l l e d water  320cc  Formalin  200cc  D i s t i l l e d water  175cc  Saponin  3g  After fixation, drated  the samples were washed w i t h running tap water, dehy-  i n an e t h a n o l  s e r i e s and embedded i n p a r a p l a s t  according  procedure of Johansen (20) which i s shown below: Dehydration: 1.  5%, e t h y l a l c o h o l  2.  ll%'.;ethyl a l c o h o l  2 hours  3.  18% e t h y l a l c o h o l  2 hours  4.  30% e t h y l a l c o h o l  2 hours  5.  approximate 50% a l c o h o l  2 hours o r  6.  .  .....  .  .  5 parts  95% e t h y l a l c o h o l  4 parts  Tertiary butyl alcohol  1 part  approximate 70% a l c o h o l  overnight  D i s t i l l e d water  3 parts  957 e t h y l a l c o h o l  5  Tertiary butyl alcohol  2 parts  approximate 85% a l c o h o l  95% e t h y l a l c o h o l Tertiary butyl alcohol approximate 95% a l c o h o l  Tertiary butyl alcohol approximate 100%-:alcbh6l.  or  longer  parts  3  parts  10  parts  7 parts a t l e a s t 1 hour  95% e t h y l a l c o h o l  9.  longer  a t l e a s t 1 hour  D i s t i l l e d water  8.  hours  D i s t i l l e d water  0  7.  .2  . . . .  Tertiary butyl alcohol  9  parts  11  parts  a t l e a s t 1 hour 3  parts  to the  14 100% 10.  ethyl alcohol  1 part  Tertiary butyl alcohol  3 changes (one of which should remain o v e r n i g h t )  Infiltration: 1.  m i x t u r e of equal and  2.  fill  Tertiary butyl alcohol  s o l i d i f y but not  container  cool  paraplast,  the b u t y l a l c o h o l - p a r a f f i n o i l m i x t u r e and  the process  place  sunk to the bottom of  replace with  twice d u r i n g  d i n g each change of f i n a l l y replace with w i l l be  just the  i n the oven a t once.  t r a c e s of a l c o h o l remain and  6.  solidified  pour o f f the e n t i r e mixture of p a r a f f i n o i l and  repeat  l e t the  completely.  about 1 hour a f t e r the m a t e r i a l has vial,  5.  of melted p a r a p l a s t and  put the m a t e r i a l on top of the cover w i t h  4.  a t l e a s t 1 hour  a v i a l three-fourth f u l l  paraplast 3.  p a r t s of p a r a f f i n o i l  pure melted  the  what paraplast.  the next 6 hours or so, d i s c a r -  paraplast. pure melted p a r a p l a s t and  the  material  ready f o r embedding w i t h i n the next 30 minutes.  Embedding: 1.  remove the v i a l from the oven, shake the m a t e r i a l to get i t o f f the bottom and  q u i c k l y pour i n t o the p l a s t i c mold.  2.  add more melted p a r a p l a s t from the  3.  w i t h a needle heated s l i g h t l y  stock c o n t a i n e r  i f necessary.  i n the flame, q u i c k l y d i s p o s e  p i e c e s of m a t e r i a l i n t o an o r d e r l y arrangement. 4.  as  soon as the mould can be moved w i t h o u t d i s t u r b i n g the  the  15 arrangement of the p i e c e s of m a t e r i a l , t r a n s f e r tbsa v e s s e l of c o l d water. 5.  l e t the mold f l o a t u n t i l the s u r f a c e of the p a r a p l a s t becomes sufficiently  f i r m to permit p l u n g i n g the mold  slowly beneath the  s u r f a c e o f the water. 6.  l e a v e the molds i n the water f o r h a l f an hour or u n t i l roughly  tho-  cooled.  A f t e r embedding, a l l the l e a f and stem samples were c u t w i t h a r o t a r y microtome a t t e n microns t h i c k n e s s .  S e c t i o n s were a f f i x e d to  s l i d e s w i t h Haupt's a d h e s i v e ( 1 8 ) , and s t a i n e d w i t h s a f r a n i n and  fast  green a c c o r d i n g to the schedule d e s c r i b e d by Johansen (20) and shown below: Staining: 1.  Xylene - 10 to 15 minutes.  2.  Xylene - 100% ETOH (1-1) - 5 minutes.  3.  95% ETOH - 5 minutes.  4.  70% ETOH - 5 minutes.  5.  50% ETOH - 5 minutes.  6.  water - wash w e l l .  7.  S t o c k w e l l ' s s o l u t i o n - a t l e a s t 24 hours, depending on m a t e r i a l .  8.  water - wash w e l l .  9.  t a n n i c a c i d - 10 to 15 minutes.  10.  water - wash w e l l .  11.  ferric  12.  water - wash w e l l .  c h l o r i d e - s e v e r a l minutes, depending on b l a c k i n g .  16 13.  safranin - at least overnight; usually  24 h o u r s .  14.  water - wash w e l l .  15.  95% ETOH w i t h 1/2% p i c r i c a c i d - no more than 10 seconds.  16.  95% ETOH w i t h 4-5 drops ammonia  17.  100% ETOH - about 30 seconds.  18.  f a s t green s t a i n i n g s o l u t i o n - s t a r t i n g w i t h a 10 second  per 100 cc - 2 minutes.  immersion. 19.  c l o v e o i l ( t o stop a c t i o n of f a s t green) - a few seconds.  20.  c l e a r f o r one or two minutes i n : clove  oil  -  50 cc  100% ETOH  -  25 cc  Xylene  -  25 cc 100 cc  21. The  X y l e n e - a t l e a s t 10 minutes.  s l i d e s were then examined m i c r o s c o p i c a l l y .  was used f o r measuring leaf  the c e l l  An o c u l a r  micrometer  l e n g t h and r a d i a l diameter of stem and  tissues. The expected mean squares f o r the a n a l y s i s of v a r i a n c e o f t h i s  experiment a r e shown i n the Appendix, Experiment  T a b l e s 1 to 4.  11  E f f e c t s of A l a r on Compact and Standard C u l t i v a r s (A)  A p p l e t r e e s of Golden D e l i c i o u s and S t a r k s p u r on EM V I I r o o t s t o c k s  were grown i n a growth chamber a t Summerland. a t 0 and 1000 ppm was a p p l i e d  to each c u l t i v a r .  D e l i c i o u s had an average shoot l e n g t h 10.6 cm.  I n December 1968, A l a r A t t h i s time Golden  of 14.25 cm and S t a r k s p u r had  There were s i x t r e e s of each treatment and c u l t i v a r .  Alar  sprays and measurements of shoot l e n g t h and l e a f number were made a t weekly  intervals.  The 'compact' S t a r k s p u r grew v e r y v i g o r o u s l y and i n  f a c t , was not n o t i c e a b l y d i f f e r e n t from Golden D e l i c i o u s when A l a r was not a p p l i e d .  However, A l a r appeared  S t a r k s p u r than on Golden Shoot  to reduce shoot growth more on  Delicious.  l e n g t h s were measured from the base o f the new shoot to the  base of the t i p l e a f c l u s t e r , and l e a v e s were counted from the base up, i n c l u d i n g h a l f - o p e n e d ones a t top of the shoot. were t e r m i n a t e d on F e b r u a r y 4, 1969.  The treatments  Samples were c o l l e c t e d a t t h a t  same time. Sampling and p r e p a r a t i o n procedures were e s s e n t i a l l y i n experiment  the same as  I , but the second and t h i r d f u l l y expanded l e a v e s below  the shoot apex were chosen f o r l e a f sampled were i n i t i a t e d w e l l a f t e r  sampling to ensure t h a t the l e a v e s  the A l a r  treatments were begun.  In  each t r e e one o r two shoots were used, and t h e r e were s i x t r e e s of each c u l t i v a r randomly  a s s i g n e d to each of the treatments and the con-  trols . (B)  A t o t a l of twenty-four t r e e s of H a r r o l d Red D e l i c i o u s and S t a r k -  rimson were used i n t h i s experiment, a l s o a t Summerland.  Technical  grade A l a r a t 0 o r 1000 ppm was a p p l i e d to s i x t r e e s o f each When f i r s t  t r e a t e d , the average  of S t a r k r i m s o n , 8 cm.  cultivar.  shoot l e n g t h of H a r r o l d was 13 cm and  A l a r treatments, shoot l e n g t h and l e a f  number  d e t e r m i n a t i o n s were done once a week f o r e i g h t weeks b e g i n n i n g A p r i l 8, 1969 w i t h H a r r o l d and A p r i l  22, 1969 w i t h S t a r k r i m s o n .  Sampling and p r e p a r a t i o n procedures were the same as d e s c r i b e d i n  18 experiment I .  The expected mean squares f o r the a n a l y s i s o f  variance  of t h i s experiment a r e shown i n the Appendix, T a b l e s 5 to 13. Experiment I I I (a) Comparisons of Compact w i t h A l a r In A p r i l ,  t r e a t e d Red D e l i c i o u s  1969, the s c i o n s o f Red D e l i c i o u s , and S t a r k r i m s o n were  g r a f t e d on EM 11 r o o t s t o c k s  planted  i n p l a s t i c pots.  A l l lateral  shoots were removed and o n l y one shoot was a l l o w e d to develop on each tree.  Twenty-one Red D e l i c i o u s and seven S t a r k r i m s o n t r e e s were used  i n t h i s experiment.  On June 12, the Red D e l i c i o u s t r e e s had an ave-  rage shoot l e n g t h of 35 cm.  Technical  grade A l a r w i t h a small  o f T r i t o n added, was a p p l i e d a t 0, 1000 and 4000 ppm. seven r e p l i c a t e s w i t h i n each treatment.  There were  The seven S t a r k r i m s o n  w i t h an average shoot l e n g t h o f 14.3 cm were l e f t u n t r e a t e d parison. block.  amount  trees  f o r com-  A l l t w e n t y - e i g h t t r e e s were randomly arranged w i t h i n one Shoot l e n g t h s were measured a t weekly i n t e r v a l s .  ment was t e r m i n a t e d f i v e weeks a f t e r  The e x p e r i -  treatment.  Experiment I I I (b) E f f e c t s of A l a r and G i b b e r e l l i c A c i d on Compact and Standard Golden D e l i c i o u s Four t r e e s each of Golden D e l i c i o u s and S t a r k s p u r were randomly a r r a n g e d w i t h i n each of two b l o c k s . rootstocks  grafted i n A p r i l ,  These t r e e s were a l s o on EM 11  1969 and prepared as d e s c r i b e d  above.  On  June 12, 1969, A l a r a t 0 and 1000 ppm, GA a t 1000 ppm, and GA a t 1000 ppm combined w i t h A l a r a t 1000 ppm were a p p l i e d i n d i v i d u a l l y to single trees within  the same b l o c k .  Shoot l e n g t h s  were measured once  19 weekly f o r s i x c o n s e c u t i v e weeks. lected ion  a t the end o f the experiment.  The same c o l l e c t i o n and p r e p a r a t -  procedures were used as i n experiment I .  t h a t o n l y two d i s c s i n experiment I , tions  L e a f and shoot samples were c o l -  a l o n g the m i d r i b of each l e a f were taken because  no s i g n i f i c a n t d i f f e r e n c e s  on a l e a f had been found.  analysis  The one e x c e p t i o n was  among the d i f f e r e n t  posi-  The expected mean squares f o r the  of v a r i a n c e of t h i s experiment a r e shown i n the Appendix,  T a b l e s 14 to 17.  RESULTS  Experiment  I  Comparisons of U n t r e a t e d  Compact and Standard A p p l e C u l t i v a r s  There were no d i f f e r e n c e s among H a r r o l d Red D e l i c i o u s , Golden D e l i c i o u s and S t a r k s p u r epidermis  creased  i n the t h i c k n e s s of the upper and lower  and spongy parenchyma.  parenchyma than the o t h e r  Starkrimson,  The S t a r k s p u r  three c u l t i v a r s  had a t h i c k e r p a l i s a d e  (Table 1, F i g . 1 ) .  The i n -  t h i c k n e s s of the p a l i s a d e parenchyma was found to be due to  g r e a t e r mean p a l i s a d e c e l l  l a y e r number and l e n g t h ( T a b l e 1, F i g .  A l t h o u g h the a n a l y s i s of v a r i a n c e  2).  showed no d i f f e r e n c e s i n the t o t a l  l e a f t h i c k n e s s between H a r r o l d Red D e l i c i o u s and Starkrimson,  discri-  minant f u n c t i o n a n a l y s i s showed t h a t the c u l t i v a r s ( F i g . 3) d i f f e r e d s i g n i f i c a n t l y a t the f i v e per cent l e v e l .  Red and Golden D e l i c i o u s  were compared w i t h r e s p e c t to l e a f t i s s u e t h i c k n e s s , p o o l i n g the compact and standard  types.  D e l i c i o u s was probably and  The t h i c k e r p a l i s a d e parenchyma of Golden  due to b o t h the g r e a t e r mean number of l a y e r s  the l e n g t h o f p a l i s a d e c e l l s  (Table 2).  A l s o compact and  types were compared p o o l i n g the two c u l t i v a r s . found to have t h i n n e r  l e a v e s , t h i n n e r p a l i s a d e parenchyma and  mean number of p a l i s a d e c e l l In order  was smaller  l a y e r s than the compact mutant ( T a b l e 3 ) .  to i n v e s t i g a t e the cause f o r the t h i c k e r shoots and  shorter internodes made o f c e l l  Standard type  standard  i n the compact apple mutants, measurements were  s i z e i n the p i t h and c o r t e x , the t h i c k n e s s o f v a s c u l a r  t i s s u e as w e l l as stem t h i c k n e s s .  F i g u r e s 4 and 5 show l o n g i t u d i n a l  TABLE 1 THICKNESS  OF  LEAF  Harrold Red D e l i c i o u s  TISSUES  OF  Starkrimson  FOUR  CULTIVARS  IN  MICRONS  Golden D e l i c i o u s  Starkspur  Significance Level  12a  12a  13a  0.1566  75a  82a  0.1999  100b  112a  0.0028  Lower Epidermis  12a  Spongy Parenchyma  75a  75a  P a l i s a d e Parenchyma  84c  90bc  Upper E p i d e r m i s  14a  14a  14a  15a  0.5218  Total  186b  191b  202b  222a  0.0047  Number of P a l i s a d e C e l l Layer  3.0b  3.1b  3.0b  3.3a  0.0015  29 b  29 b  34a  0.0413  Average P a l i s a d e C e l l Length  X  33ab  w i t h i n a row, means having a l e t t e r i n common a r e not s i g n i f i c a n t l y 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  different  a t the  Leaf  thickness  in  microns  FIGURE 2 The l e a f c r o s s s e c t i o n of S t a r k s p u r showing longer p a l i s a d e c e l l s and g r e a t e r mean p a l i s a d e number ( l e f t ) than i n the standard Golden D e l i c i o u s ( r i g h t ) . (381X) u>  The  leaf  c r o s s s e c t i o n of S t a r k r i m s o n ( l e f t ) showing g r e a t e r l e a f Red D e l i c i o u s ( r i g h t ) . (300X)  thickness  than  TABLE 2 THICKNESS  OF LEAF TISSUES OF RED DELICIOUS GOLDEN DELICIOUS IN MICRONS  Red D e l i c i o u s  x  Golden D e l i c i o u s  x  AND  Significance Level  Lower E p i d e r m i s  12  13  0.0456  Spongy Parenchyma  75  78  0.2317  P a l i s a d e Parenchyma  87  106  0.0008  Upper E p i d e r m i s  14  15  0.2107  188  212  0.0018  Total Number o f P a l i s a d e C e l l Layer Average P a l i s a d e C e l l Length  x  3.02  29  mean of standard and  3.19  33 compact types.  0.0039  0.0078  TABLE 3 THICKNESS  OF LEAF TISSUES OF STANDARD COMPACT TYPES IN MICRONS  AND  Standard  Compact  Lower Epidermis  12  12  0.6533  Spongy Parenchyma  75  79  0.1923  Palisade  92  101  0.0350  14  14  0.7804  195  206  0.0345  Parenchyma  Upper Epidermis Total Number o f P a l i s a d e C e l l Layer Average P a l i s a d e C e l l Length  3.0  31  3.2  32  xx  Significance Level  0.0013  0.3983  ^ e a n s of theRed D e l i c i o u s and Golden D e l i c i o u s . 'means of the Starkrimson and S t a r k s p u r .  27  COLLENCHYMA  PARENCHYMA  VASCULAR  PITH  FIGURE 4 L o n g i t u d i n a l s e c t i o n o f Red D e l i c i o u s stem showing c o l l e n c h y m a , parenchyma, v a s c u l a r t i s s u e , and p a r t o f p i t h . (118 X)  TISSUE  COLLENCHYMA) ) CORTEX PARENCHYMA )  VASCULAR  TISSUE  PITH  FIGURE 5 C r o s s s e c t i o n o f Red D e l i c i o u s stem showing p a r t o f c o l l e n c h y m a , parenchyma, v a s c u l a r . t i s s u e and p i t h . (60 X) [S3 00  TABLE 4 CELL  LENGTHS PITH OF  Cultivar  OF STEM COLLENCHYMA, PARENCHYMA AND FOUR APPLE CULTIVARS IN MICRONS  Collenchyma  Harrold  48a  Starkrimson  Parenchyma  Pith  61a  41a  44a  67a  38a  Golden  45a  57a  37a  Starkspur  41a  51a  38a  X  W i t h i n a column, means having a l e t t e r i n common are not s i g n i f i c a n t l y d i f f e r e n t a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  30 and c r o s s s e c t i o n s r e s p e c t i v e l y o f Red D e l i c i o u s shoot t i s s u e . were no s i g n i f i c a n t d i f f e r e n c e s i n c e l l  There  l e n g t h of collenchyma,  chyma or p i t h c e l l s among the f o u r c u l t i v a r s ( T a b l e 4 ) .  paren-  I n stem  cross  s e c t i o n s , S t a r k r i m s o n had a t h i c k e r v a s c u l a r t i s s u e than the o t h e r cultivars  ( T a b l e 5 ) , but the t h i c k n e s s of the c o r t e x , p i t h and  stem were not s i g n i f i c a n t l y d i f f e r e n t i n the c u l t i v a r s .  The  cell  meter o f the collenchyma of Red D e l i c i o u s and S t a r k r i m s o n Red was  g r e a t e r than t h a t of Golden D e l i c i o u s and S t a r k s p u r .  total dia-  Delicious  Starkspur  had a s m a l l e r p i t h c e l l diameter than the o t h e r t h r e e c u l t i v a r s , but the  mean diameter of the parenchyma c e l l s d i d not d i f f e r among  vars (Table 6).  The data p r e s e n t e d i n T a b l e 5 were measured as  cultithough  each t i s s u e were c i r c u l a r . Experiment  11  E f f e c t s o f A l a r on Compact and Standard A p p l e The  shoot growth of Red D e l i c i o u s was  by A l a r u n t i l was  first  1000  ppm  s i x weeks a f t e r  Cultivars  not s i g n i f i c a n t l y  affected  treatment, but growth r a t e s u p p r e s s i o n  suspected f o u r weeks a f t e r treatment ( T a b l e 7).  A l a r at  d i d not show any i n h i b i t i n g e f f e c t on the shoot growth of  S t a r k r i m s o n ( T a b l e 7). A l a r d i d not s i g n i f i c a n t l y a f f e c t  l e a f number ( T a b l e 8 ) , l e a f  l e n g t h or w i d t h i n e i t h e r Red D e l i c i o u s or S t a r k r i m s o n ( T a b l e 9 ) .  How-  ever, i n Golden D e l i c i o u s ,  was  s i g n i f i c a n t i n h i b i t i o n on shoot l e n g t h  observed from the second week a f t e r A l a r treatment. to  treatment i n c r e a s e d g r a d u a l l y t i l l  ment ( T a b l e 10).  The d i f f e r e n c e  the t e r m i n a t i o n o f the e x p e r i -  The e f f e c t of A l a r on S t a r k s p u r shoot growth  was  due  TABLE 5 THICKNESS OF STEM CORTEX, VASCULAR TISSUE, DIAMETERS AND STEM OF FOUR APPLE CULTIVARS IN MICRONS  Cultivar  Cortex  X  OF  PITH,  Vascular Tissue  Pith Diameter  Shoot Diameter  H a r r o l d Red D e l i c i o u s  256a  248b  1073a  2160a  Starkrimson  263a  414a  9 73a  2273a  Golden D e l i c i o u s  233a  327ab  908a  2006a  Starkspur  288a  279b  939a  1950a  w i t h i n a column, means s h a r i n g the same l e t t e r a r e not s i g n i f i c a n t l y d i f f e r e n t a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  TABLE 6 CELL  DIAMETERS OF STEM OF FOUR APPLE  Cultivar  COLLENCHYMA, PARENCHYMA AND PITH CULTIVARS IN MICRONS  Collenchyma  H a r r o l d Red D e l i c i o u s  20a  Starkrimson  Parenchyma  Pith  29a  35a  21a  27a  33a  Golden D e l i c i o u s  18b  25a  30a  Starkspur  18b  25a  26b  X  W i t h i n a column, means s h a r i n g the same l e t t e r a r e n o t s i g n i f i c a n t l y d i f f e r e n t a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  TABLE 7 EFFECT OF ALAR RED DELICIOUS  Cultivar  Alar  Concentration (ppm)  Weeks A f t e r 1  Red  Delicious  ON SHOOT GROWTH OF AND STARKRIMSON (CM)  Treatment  2  3  4  5  6  7  8  15g  23f  29 e  35d  43b  50a  55a 44b  0  8h  1000  8h  15g  21f  28e  32de  37cd  41b c  X  Significance Level  0.0135  Starkrimson  0  12a  18a  26a  33a  40a  45a  45a  1000  14a  24a  30a  38a  42a  45a  47a  Significance Level  0.9389  w i t h i n each c u l t i v a r , means having a l e t t e r i n common a r e not s i g n i f i c a n t l y a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t . x  different  TABLE 8 EFFECT  Cultivar  Red  Delicious  Alar  OF  ALAR  ON LEAF NUMBER OF AND STARKRIMSON  Concentration (ppm)  RED  DELICIOUS  Weeks A f t e r 3 4  Treatment 5 6  7  8  0  6  8  11  14  16  18  21  23  1000  6  8  11  14  16  18  20  22  Significance Level  0.5581  Starkrimson  0  8  12  15  18  20  22  23  1000  10  13  17  19  20  23  24  Significance Level  0.7543 Co  •P-  TABLE 9 EFFECT OF ALAR ON AVERAGE LEAF LENGTH OF RED DELICIOUS AND STARKRIMSON  Cultivar  Red  Delicious  A l a r Concentration (ppm)  Significance Level  L e a f Width  0  10.1  5.3  1000  10.0  4.9  Significance Level  Starkrimson  Leaf Length  AND WIDTH (CM)  0.8042  0.2430  0  12.4  6.1  1000  12.0  5.7  0.2673  0.1551  TABLE 10 INFLUENCE OF ALAR ON SHOOT GROWTH OF STARKSPUR AND GOLDEN DELICIOUS (CM)  Cultivar  Golden D e l i c i o u s  Alar  Concentration (ppm)  0 1000  Significance Level  0.0000  Starkspur  0 1000  Significance Level  2  16i  Weeks A f t e r Treatment 3 4 5  6  20h  26f  33d  40c  46b  51a  141  16i  19h  23g  27f  30e  33d  13hij  16gh  22ef  31d  41c  47b  52a  9i  llij  14hi  16hi  21fg  24ef  26de  X  0.0000  w i t h i n each c u l t i v a r , means having a l e t t e r i n common a r e not s i g n i f i c a n t l y a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  different  Co  TABLE EFFECT  Cultivar  Golden D e l i c i o u s  OF  ALAR  Alar  ON  LEAF  Concentration (ppm)  0; 1000  Significance Level  11  NUMBER  OF  1  14f 12g  X  GOLDEN  DELICIOUS  Weeks A f t e r Treatment 2 3  4  16.6d  19b  22a  15e  17cd  17.6b  0.0007  ^Sneans having a l e t t e r i n common a r e not s i g n i f i c a n t l y d i f f e r e n t a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  38 q u i t e d i f f e r e n t from e x p e c t a t i o n . during  the  during  the f o u r t h week.  hibited  1000  25%  (Table  ppm  number of  one  of  11).  control.  no  10).  Four weeks a f t e r treatment,  show any  s i g n i f i c a n t e f f e c t on  by  the  leaf  thinner  and  Starkrimson leaf  lamella  s i g n i f i c a n t d i f f e r e n c e s between treatments i n I n Golden D e l i c i o u s , the  lower e p i d e r m i s and S t a r k s p u r had  less total  data f o r c e l l  fewer p a l i s a d e  thinner  l e n g t h i n the  cells  (Table  than  stem l o n g i t u d i n a l s e c t i o n s  the  significantly  and  from the u n t r e a t e d Experiment I I I  than the  showed longer  compared w i t h the t r e a t e d p l a n t s .  treated Harrold  Golden D e l i c i o u s were not  The  signifiin  untreated parenchyma  cell  lengths  cell in  significantly different  plants.  (a)  Comparisons of Compact w i t h A l a r t r e a t e d Red A l a r was  of  l e n g t h of parenchyma c e l l  shorter  S t a r k s p u r Golden D e l i c i o u s p l a n t s  c o r t e x as  pali-  d i f f e r e n t treatments i s shown i n  collenchyma c e l l s but  p i t h were not  the  12).  A p p a r e n t l y the A l a r t r e a t e d S t a r k r i m s o n p l a n t s had  shorter  any  treated  spongy parenchyma, fewer  leaf thickness  c u l t i v a r s , each w i t h two  c o r t e x and  in  (Table  examination of H a r r o l d  Treated  T a b l e 13.  ones.  50%  seven v a r i a b l e s measured.  sade c e l l s and  cantly  found  However, s i x weeks a f t e r treatment, A l a r i n -  A l a r d i d not  revealed  p l a n t s had  four  l e s s r e t a r d a t i o n was  Starkspur.  the  The  noted  s i g n i f i c a n t l y reduced Golden D e l i c i o u s l e a f number  Microscopic sections  r e t a r d a t i o n e f f e c t was  second week a f t e r treatment, but  shoot growth by  Alar  The  found to i n c r e a s e  the  thickness  Delicious  of t o t a l  l e a f , spongy  TABLE INFLUENCE  Cultivar  Red D e l i c i o u s  Starkrimson  Golden D e l i c i o u s  A l a r Concentration (ppm)  THICKNESS CULTIVARS  Lower Spongy E p i d e r m i s Parenchyma  OF LEAF TISSUES (MICRONS)  Palisade Parenchyma  Upper Epidermis  OF  T o t a l Pa 1 i sade Avera ge•-• Number Palisade  0  12  76  124  16  226  3  41  1000  11  73  114  15  213  3  39  0  11  81  100  16  208  3  33  1000  11  90  113  15  229  3  36  0  11  77  96  14  198  3  32  10**  81  90  14  195  2.8*  33  0  11  80  97  14  203  3  33  1000  11  71*  89  14  184*  2.9*  31  1000  Starkspur  OF ALAR ON FOUR APPLE  12  - • s i g n i f i c a n t a t 5 per cent * * s i g n i f i c a n t a t 1 per cent  level. level.  TABLE 13 INFLUENCE  OF ALAR ON THE CELL LENGTHS OF COLLENCHYMA, PARENCHYMA AND PITH OF FOUR APPLE CULTIVARS (MICRONS)  Alar Concentration (ppm)  Pith  Red  Delicious  Starkrimson  Golden D e l i c i o u s  Starkspur  0  33  46  38  42  1000  35  31**  43  33  0  59  54  50  56  1000  53  50  54  42*  0  35  37  37  41  1000  38  34  39  32  - ' s i g n i f i c a n t a t 5 per cent * * s i g n i f i c a n t a t 1 per cent  level. level.  o  TABLE 14 INFLUENCE  Cultivar  x  Spongy Parenchyma  Palisade Parenchyma  TISSUES  Upper Epidermis  Total  Palisade Number  Average Palisade  63ab  78b  16a  168b  3a  26b  10a  71a  87a  16a  184a  3a  30a  4000  10a  69a  90a  16a  186a  3a  30a  0  10a  59b  75b  16a  161b  3a  27b  Delicious  Red  Delicious  1000  Red  Delicious  Significance Level  ALAR ON THE THICKNESS OF LEAF OF RED DELICIOUS IN MICRONS  Alar Concentration Lower (ppm) Epidermis  Red  Starkrimson  OF  0  11a  0.8043  0.0342  0.0039  0.7688  0.0062  0.0907  0.0085  W i t h i n each column, means having a l e t t e r i n common a r e not s i g n i f i c a n t l y d i f f e r e n t a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  43 parenchyma and (Table ppm,  14).  l e n g t h of p a l i s a d e  The  were not  c e l l i n H a r r o l d Red  r e s u l t s from two  concentrations  d i f f e r e n t from each o t h e r .  palisade  parenchyma and  were not  d i f f e r e n t from u n t r e a t e d  slightly  t h i c k e r spongy parenchyma was  and  average p a l i s a d e  lower e p i d e r m i s and  The  i . e . 1000  total  leaf  parenchyma of  H a r r o l d Red  Delicious and  4000  thickness,  Starkrimson  D e l i c i o u s , although  found i n the  latter.  The  upper  p a l i s a d e number were not d i f f e r e n t from t r e a t -  ment to treatment, hence the r e s u l t e d from an i n c r e a s e  t h i c k e r l e a f of A l a r  i n the  thickness  of  t r e a t e d Red  Delicious  spongy parenchyma  and  p a l i s a d e parenchyma ( F i g . 6). A l a r i n h i b i t e d shoot growth of Red i n h i b i t i n g e f f e c t was  greater  4000 ppm  The  (Table  15).  second week's growth and experiment.  A f t e r two  D e l i c i o u s a p p l e t r e e s and  a t the h i g h e r c o n c e n t r a t i o n ,  treatment e f f e c t was was  still  p r e s e n t a t the  i . e . at during  termination  of  the the  weeks, the growth r a t e of A l a r - t r e a t e d Red  l i c i o u s was  l e s s than t h a t of S t a r k r i m s o n Red  krimson was  still  Experiment I I I  noticeable  the  smaller  D e l i c i o u s although  DeStar-  i n total size.  (b)  E f f e c t s of A l a r and Golden D e l i c i o u s There were no  G i b b e r e l l i c A c i d on Compact and  Standard  s i g n i f i c a n t d i f f e r e n c e s between means f o r the  thick-  ness of Golden D e l i c i o u s l e a f t i s s u e s as a r e s u l t of treatment w i t h A l a r or GA  (Table  16).  Golden D e l i c i o u s and end  However, GA  increased  mean shoot growth of  S t a r k s p u r Golden D e l i c i o u s by  of the experiment (Table  17).  A l a r a t 1000  ppm  29  per  cent by  or the A l a r and  the GA  44 combination d i d not a f f e c t the shoot growth o f Golden D e l i c i o u s S t a r k s p u r Golden D e l i c i o u s ( T a b l e 17). s t i m u l a t i n g e f f e c t of  GA.  and  Apparently A l a r n u l l i f i e d  the  TABLE 15 EFFECT OF ALAR STARKRIMSON AND  Cultivar  Alar  Concentration (ppm)  ON SHOOT GROWTH OF RED DELICIOUS (CM)  Red  Delicious  0  36h  Red  Delicious  1000  Red  Delicious  Starkrimson  Weeks A f t e r 2 3  1  Treatment 4 5  6  47ef  54c  59b  63a  66a  36h  45 gh  48ef  50de  52cd  54c  4000  34i  42gh  45fg  46f  47ef  48ef  0  14m  211  25kl  28jk  31j  33i  Significance Level f o r Interaction  x  0.0000  ^ e a n s h a v i n g a l e t t e r i n common a r e not s i g n i f i c a n t l y d i f f e r e n t a t the 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  TABLE 16 INFLUENCE OF LEAF  Cultivar  Golden  Treatment (ppm)  Lower Epidermis  Spongy Parenchyma  Palisade Parenchyma  Upper Epidermis  control  11  70  93  15  189  3  32  A - 1000  11  65  89  15  180  3  33  GA - 1000  11  54  85  16  166  3  28  A - 1000, GA - 1000  10  58  86  15  169  3  31  control  11  66  90  14  182  3  31  A - 1000  11  60  88  15  174  3  31  GA - 1000  12  61  87  15  175  3  29  A - 1000, GA - 1000  12  72  94  14  191  3  34  Delicious  Starkspur  Significance  either  OF ALAR AND GIBBERELLIC ACID ON THICKNESS TISSUES OF STARKSPUR AND GOLDEN DELICIOUS*  Level  0.4403  0.2.187  0.6475  0.8584  Total  Number o f Palisade Layer  0.1668  There was no s i g n i f i c a n t e f f e c t of any o f these t r e a t m e n t s on l e a f t i s s u e s cultivar.  0.9695  thickness of  Average Pa l i s a d e C e l l Length  0.4339  TABLE 17 INFLUENCE  Treatment (ppm)  OF  ALAR AND GIBBERELLIC ACID ON MEAN SHOOT OF STARKSPUR AND GOLDEN DELICIOUS (CM)  1  X  2  Weeks A f t e r Treatment 3 4  GROWTH  5  6  Control  23h  31g  33fg  35cdefg  38cdef  40cd  A l a r - 1000  24h  34efg  38cdef  39cdef  39cdef  41c  GA  23h  34efg  41c  48b  56a  58a  19 h  32g  35defg  38cdef  40cd  41c  - 1000  A l a r - 1000, GA - 1000  S i g n i f i c a n c e L e v e l of I n t e r a c t i o n  0.0001  means h a v i n g a l e t t e r i n common a r e not s i g n i f i c a n t l y 5 per cent l e v e l by Duncan's M u l t i p l e Range T e s t .  different  a t the  DISCUSSION  S t a r k s p u r was  found  to have a t h i c k e r l e a f , p a l i s a d e parenchyma  and a g r e a t e r mean p a l i s a d e number and tivars. Arasu  s i z e than the other  three  These r e s u l t s agree w i t h the c o n c l u s i o n s of Westwood  (1) and Westwood and Z i e l i n s k i  have t h i c k e r l e a v e s and  (38)  palisade tissue.  have s h o r t e r i n t e r n o d e s , g r e a t e r l e a f  t h a t the spurtype They a l s o found  cul-  (37),  mutants  the mutant to  s u r f a c e per f o o t of shoot,  fewer 2  s i d e branches but more spurs, and of l e a f .  g r e a t e r c h l o r o p h y l l content  These a t t r i b u t e s of spurtypes  l i g h t d i s t r i b u t i o n , bearing fruit-bearing potential. t h i c k e r l e a f and examination  f a v o r them w i t h r e g a r d  surface, photosynthetic e f f i c i e n c y  The  present  study has demonstrated  of the l e a f anatomy of the spurtype mutants.  i n t h i c k n e s s of the p a l i s a d e parenchyma was  t h i s d i f f e r e n c e was By  due  both  increase longer  The  Star-  t h i c k e r than those of Red D e l i c i o u s  a t t r i b u t e d to t h i c k e r p a l i s a d e parenchyma.  comparing the t h i c k n e s s of l e a f t i s s u e s of Red  cent t h i c k e r i n terms of t o t a l l e a f t h i c k n e s s .  Delicious with  Apparently  s i g n i f i c a n t d i f f e r e n c e s i n t h i c k n e s s of lower e p i d e r m i s ,  dermis and  and  to  Golden D e l i c i o u s ( T a b l e 2), i t i s e v i d e n t t h a t the l a t t e r was  no  to  careful  The  a g r e a t e r mean p a l i s a d e l a y e r o f them.  krimson l e a v e s were o n l y s l i g h t l y and  found  cm  the  p a l i s a d e t i s s u e s of the spurtype mutants by  p a l i s a d e c e l l s and  per  spongy parenchyma between these two  varieties.  11.3  per  t h e r e were upper e p i The  dif-  f e r e n c e i n t o t a l l e a f t h i c k n e s s i s a t t r i b u t a b l e to d i f f e r e n c e s i n p a l i sade parenchyma which was  18 per cent t h i c k e r i n Golden D e l i c i o u s than  49 i n Red  Delicious.  18 per cent h i g h e r  Looney r e p o r t e d t h a t Golden D e l i c i o u s had net a s s i m i l a t i o n r a t e than Red  an  Delicious (22).  Compact mutants have been r e p o r t e d to have a t h i c k e r stem and shorter internodes study  i t was  than s t a n d a r d v a r i e t i e s  vars studied (Table 4 ) . tex, p i t h and  p i t h c e l l s among the f o u r  cell culti-  I n stem c r o s s s e c t i o n s , the ..thickness of c o r -  t o t a l stem of mutants were a l s o not d i f f e r e n t from  (Table 5).  types were not d i f f e r e n t from the spur  These r e s u l t s r e p r e s e n t types  the  Furthermore, i n c e l l diameter of d i f f e r e n t t i s s u e s  ( T a b l e 6 ) , standard  standard  However, i n t h i s  found t h a t t h e r e were no s i g n i f i c a n t d i f f e r e n c e s i n  l e n g t h of collenchyma, parenchyma and  standard  (37, 38).  types.  the f i r s t comparisons between compact  w i t h r e s p e c t to stem anatomy.  used i n t h i s experiment was  Since  and  the shoot m a t e r i a l  taken from the f i r s t i n t e r n o d e under  the  growing t i p , m a n i f e s t a t i o n of the reasons f o r the r e p o r t e d t h i c k e r and s h o r t e r shoots may should  not occur i n t h i s r e g i o n of the shoot.  c o n s i d e r mature t i s s u e s .  F u t u r e work  I t i s q u i t e p o s s i b l e t h a t the  of A l a r treatment have not shown up completely  effects  on t h i s premature t i s -  sue . In experiment I I , Red not show any with ppm  12).  However, i n experiment I I I , A l a r a t  found to i n c r e a s e the t h i c k n e s s of t o t a l  chyma and  ppm  did  s i g n i f i c a n t d i f f e r e n c e i n l e a f t h i c k n e s s when compared  the c o n t r o l ( T a b l e was  D e l i c i o u s t r e a t e d w i t h A l a r a t 1000  l e n g t h of p a l i s a d e c e l l s  r e s u l t s might be due  (Table  15).  1000  l e a f , spongy parenThese c o n f l i c t i n g  to the d i f f e r e n c e s of season, l o c a t i o n and  s p e c i f i c e f f e c t of the m a t e r i a l used i n the experiment.  the  These a r e  the  50 first  r e s u l t s reported  on the e f f e c t s of A l a r on the anatomy of e i t h e r  compact or s t a n d a r d Red  Delicious.  I n Golden D e l i c i o u s thinner  (Table  lower epidermis and  spur Golden D e l i c i o u s had leaf thickness,  and  12),  the  fewer p a l i s a d e  thinner  The  c e l l s and  cells.  luence the e f f e c t of A l a r t r e a t m e n t .  (17)  total infthat  differently  concentration.  p r e s e n t study a l s o r e v e a l e d  t h a t s u p p r e s s i o n of I t was  terminal found t h a t  of  f o u r c u l t i v a r s used i n experiment I I , S t a r k r i m s o n t r e e s were not  a f f e c t e d by A l a r treatment ( T a b l e most s u s c e p t i b l e  to the A l a r  7, F i g . 7) and  treatment ( T a b l e  among c u l t i v a r s , the above r e s u l t s l a b e l e d m a t e r i a l had  c o u l d be  than the Golden D e l i c i o u s  cent ( T a b l e  GA  stimulated  GA  was  applied  treatment.  ( 1 6 ) , and  Golden D e l i c i o u s A l a r a t 19 per  explained  differences  i f incorrectly  However, even b e f o r e the app-  treatment much d i f f e r e n t l y  l e a f number, l e n g t h or w i d t h of Red  w i t h those o f H a l f a c r e  Besides  the  trees.  krimson were a f f e c t e d by A l a r  by  10).  sampled grew v e r y v i g o r o u s l y  l i c a t i o n of A l a r , they responded to the  Neither  S t a r k s p u r was  been used i n the experiment.  though the S t a r k s p u r t r e e s  r  Stark-  a l s o reported  responded to A l a r  growth by A l a r v a r i e d among the c u l t i v a r s used. the  had  the t r e a t e d  Thus c u l t i v a r might  Halfacre  York I m p e r i a l  when t r e a t e d a t the same  of t r e a t e d p l a n t s  spongy parenchyma, a s m a l l e r  fewer p a l i s a d e  the Golden D e l i c i o u s and  leaves  1000  11),  but  ppm no  the was  D e l i c i o u s and  Star-  These f i n d i n g s do not  f i n d i n g s need v e r i f i c a t i o n . found to reduce the  e f f e c t was  observed on  shoot growth of Golden D e l i c i o u s and  agree In  l e a f number Starkspur.  S t a r k s p u r but when  i n combination w i t h A l a r , the s t i m u l a t i n g e f f e c t  was  51 cancelled  c o m p l e t e l y by A l a r .  E d g e r t o n and  t h a t the s t i m u l a t i n g e f f e c t of GA be  c a n c e l l e d by  on Red  Hoffman (14)  also  found  D e l i c i o u s apple trees  could  Alar.  S i n c e the n a t u r a l compact mutants have many m e r i t s over the dard growing s t r a i n s , i t may  be  d e s i r a b l e to induce s i m i l a r changes i n  s t a n d a r d a p p l e t r e e s or i n t e n s i f y the d e s i r a b l e t r a i t s by means of treatment w i t h growth r e g u l a t o r s there a r e o n l y a few  workers s t u d y i n g  n a t u r a l compact mutants and worker, I n t r i e r i  (19),  spur type t r e e s r a n d larities  9.5  per c e n t .  parenchyma.  active s i t e within dant.  dings h e l p  due  of p a l i s a d e  increase  of s e v e r a l c u l t i v a r s had growth r a t e of Red immediate and  weeks a f t e r the a p p l i c a t i o n of A l a r .  of H a r r o l d  Red  ppm  and  4000 ppm,  the  a  noted.  e x p l a i n the r e p o r t of E d g e r t o n and  1000  that  Delicious  i n t o t a l thickness  to an  parenchyma was  i n h i b i t o r y e f f e c t was  t r a t i o n s of  Red  of  i n thickness  l e a f t i s s u e which responds to the  A l a r i n h i b i t e d the This  increase  of H a r r o l d  Hence i t seems t h a t t h i s t i s s u e i s the  the  t r e a t e d a p p l e trees  literature,  One  t r e a t e d w i t h A l a r showed many s i m i -  A f t e r the treatment w i t h A l a r 4000 ppm,  i n thickness  However,  induced compact h a b i t .  leaf thickness  A p p a r e n t l y the  t i s s u e a f t e r A l a r treatment was palisade  compacts  anatomy.  A l a r treatment i n c r e a s e d by  such as A l a r .  upon r e v i e w i n g the  standard trees  of the  the r e l a t i o n s h i p s among the  the c h e m i c a l l y  reported,  i n morphology and  stan-  leaf of  primary  growth r e t a r -  16 per cent  increase  These a n a t o m i c a l  Hoffman (14) thicker  the  fin-  that A l a r  leaves.  Delicious apple shoots.  l a s t e d f o r more than s i x  When A l a r was  a p p l i e d a t concen-  i n h i b i t i o n s of the shoot growth  D e l i c i o u s were found to be  18 per  cent and  27  per  cent  52 respectively.  This  ( 3 , 4,  16,  14,  15,  r e s u l t concurs w i t h the f i n d i n g s of o t h e r 17)  that A l a r i n h i b i t s  the t e r m i n a l  workers  growth of  I n t e r e s t i n g l y , A l a r d i d not appear to i n f l u e n c e the  l e n g t h of any  the stem c e l l types examined i n the c u r r e n t  As  e a r l i e r , t h i s may  have been due  study.  support the f i n d i n g s of M a r t i n jit a l . (24) who  had  a greater  of  discussed  to the sampling procedure but  may  apples.  i t also  found t h a t A l a r  e f f e c t on c e l l d i v i s i o n i n a p p l e f r u i t s  than on  cell  size. U n t r e a t e d H a r r o l d Red continuously.  D e l i c i o u s shoots have a tendency to grow  T h i s would r e s u l t i n a t a l l e r  S t a r k r i m s o n and A l a r t r e a t e d H a r r o l d curves a r e q u i t e s i m i l a r and These r e s u l t s a r e cluded  study has  considering  review of I n t r i e r i (19) who  the c o n c e n t r a t i o n  spur type t r e e s  to the u n d e r s t a n d i n g of  A p p a r e n t l y both the nature of of A l a r a r e  the use of t h i s growth  con-  anatomy.  contributed  e f f e c t s of A l a r on a p p l e t r e e s . c u l t i v a r and  D e l i c i o u s the shoot growth  a p p l e t r e e s t r e a t e d w i t h A l a r and  a r e q u i t e s i m i l a r i n morphology and present  However, i n the  become l e v e l a t about the f o u r t h week.  consistent withtthe  t h a t standard  This  Red  tree.  the the  important f a c t o r s when  retardant.  SUMMARY  A two and  y e a r study was  conducted to i n v e s t i g a t e  a n a t o m i c a l changes i n a p p l e l e a f and  shoot  There i s no  p r e v i o u s a n a t o m i c a l comparisons of  compact a p p l e t r e e s or between A l a r  t r e e s of Red  Delicious.  and  t r e a t e d Red  numbers of p a l i s a d e  two  and  the  ppm  From A p r i l  was  shoot  stems between  applied  reported.  D e l i c i o u s and  Golden D e l i c i o u s ,  A l a r were a p p l i e d  experiment A l a r a t the  c o n c e n t r a t i o n s of 0,  to c u l t i v a r s which had  1000  each  treatments.  c o n c e n t r a t i o n s of  to a d d i t i o n a l t r e e s of the  to June 1969,  untreated  spongy parenchyma  compact t y p e s , were s t u d i e d without A l a r  second phase of the  1000  ana-  l a y e r s i n compact mutants or i n A l a r  first  v a r i e t i e s , Red  w i t h s t a n d a r d and I n the  width,  t r e a t e d and  D e t a i l e d measurements of  D e l i c i o u s are  I n 1968,  and  Factors  length.  s t a n d a r d and  thickness  morphological  stem t i s s u e s .  l y z e d were l e a f anatomy, l e a f number, l e a f l e n g t h anatomy and  the  0  same c u l t i v a r s . and  been g r a f t e d  4000 ppm on EM  II  of root-  stocks . Without A l a r treatment, i t was l e a f and  thicker palisade  s t u d i e d and  Red  mutants had greater The shorter  parenchyma than the o t h e r t h r e e  D e l i c i o u s was  than Golden D e l i c i o u s .  found t h a t S t a r k s p u r had  Results  thickness  a l s o i n d i c a t e d t h a t compact a p p l e palisade  parenchyma  number as compared w i t h standard  compact mutants have been r e p o r t e d internode.  thicker  cultivars  found to have l e s s t o t a l l e a f  on average t h i c k e r l e a v e s and  mean p a l i s a d e  a  types.  to have t h i c k e r  However, m i c r o s c o p i c e x a m i n a t i o n of  and  stems  and  samples taken  54 from the f i r s t  internode  differences i n c e l l  under the growing t i p r e v e a l e d no  l e n g t h of collenchyma, parenchyma, p i t h  c e l l diameter or t h i c k n e s s The vars.  suppression  The  significant  i n the  same t i s s u e s .  of t e r m i n a l growth by A l a r varied.among  response to A l a r was  h i b i t i o n of shoot growth by 50  greatest with Starkspur per  cells,  cent was  not a f f e c t e d by A l a r treatment i n the  observed.  culti-  where an i n Starkrimson  was  same experiment.  I n the t h i r d phase of the experiment, A l a r t r e a t e d l e a f b l a d e s p l a n t s were found to i n c r e a s e i n t o t a l t h i c k n e s s , i n t h i c k n e s s spongy parenchyma and from two  concentration  to d i f f e r  i n the l e n g t h of p a l i s a d e c e l l s . of A l a r , i . e . 1000  from each o t h e r .  These data  and  4000 ppm,  i n d i c a t e t h a t the  The  of  of  results  were not  found  s i t e which i s  most a f f e c t e d by A l a r i s i n the p a l i s a d e parenchyma c e l l s of the  leaf  tissue. GA by  stimulated  29 per c e n t , but  the  shoot growth of Golden D e l i c i o u s and  t h i s s t i m u l a t i n g e f f e c t was  Starkspur  prevented by A l a r .  BIBLIOGRAPHY  1.  A r a s u , N. T. 1968. Spur-Type s p o r t s i n A p p l e s . M a i l i n g Res. S t n . f o r 1967. 113-119.  Ann.  Rept. E .  2.  Ashby, D. L . and N. E . Looney. 1968. 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Planta (Berl.) 71:68-80.  59  APPENDIX A n a l y s i s o f V a r i a n c e Models (Table numbers correspond t o those i n t e x t )  Table 1 Line Number  Line number of F denominator  Degrees o f freedom  Source o f V a r i a t i o n  Expected mean squares  Clone  (c-1)  =  3  T r e e s w i t h i n Clones  c(t-l)  =  8  Shoots w i t h i n Trees  ct(s-l)  = 12  Leaves w i t h i n Shoots  cts(e-l)  = 2<4  5  Position  p-1  =  2  6  P o s i t i o n x Clone  (p-D(c-l)  =  6  7  P o s i t i o n x Trees w i t h i n Clones  c(p-lHt-l)  = 16  0  8  P o s i t i o n x Shoots w i t h i n Trees  ct(p-lXs-l)  = 24  0  9  P o s i t i o n x Leaves w i t h i n Shoots  cts(p-l)(l-l)=  Disc  ctspl(d-l)  = 144  ctspld(m-l)  = 864  ctspldm-1  =1151  10 E  Measurements  Total  w i t h i n Discs  48  o 2  E  + 2 o 2  ° E 2  0  10  0  0  + 2 a 2  2 E 2 E 2 E  + 2  + 2  + 2  2 ° E 2 E  +1 2 o 2  + 1 2 a 2  S(TC)  S(TC)  + 2 l  + 1 2  + 2 4 o 2  T(C)  + 7 2 e 2  C  * T(C) o 2  2 ° S(TC)  2 2 ° D ( T C S L P ) ° L(TCS) +6  2 2 2 2 ° D(TCSLP) ° PL(TCS) * PS(TC) ° T P ( C ) +2  + 2  + 2  2  + 6  2  0  +6  D(TCSLP)  2 E  2  D(TCSLP) ° L(TCS)  2  E  ° L(TCS)  + 6  2 2 ° D(TCSLP) ° L(TCS)  2  10  D(TCSLP)  + 2o  2 ° D(TCSLP) '  +1  PL(TCS)  a  +4o  ,+l(o  PL(TCS)  + 8  PS(TC)  + 8a  PS(TC)  + 8o  2 2 2 ° D ( T C S L P ) ° P L ( T C S ) ° PS(TC)  + 2  +2  2 2 ° D ( T C S L P ) ° PCLCTCS) +2  2 + 2  + 4  D(TCSLP)  + 9 6 6  2 P.  .•24o' TP(C) PC TP(C)  60  Table  2, 3  Line Number  Source o f V a r i a t i o n  Line number of F denominator  Degrees o f freedom  1  Variety  (v-1)  =  1  2  Type  (t-1)  =  1  3  V a r i e t y x Type  (v-D(t-l)  =  1  4  Tree w i t h V a r i e t y and Type  vt(r-l)  =  8  E  Measurements w i t h Type  vtr(m-l)  =1140  vtrm-1  = 1151  Total  Variety,  Expected mean squares  E  o 2  + 9 6  ° R(VT) 2  ° E 2  + 9 6 ( j 2  R(VT)  ° E  + 9 6 < j 2  R(VT)  ° E  + 9 6  2  2  +  5  7  + 2 8 8  ° R(VT) 2  + 5 7 6 6 2  6  e  2  V  T  ° VT 2  61  T a b l e U, 6  Line Number  Line number of F denominator  Degrees o f freedom  Source o f V a r i a t i o n  Expected mean squares  (v-1)  =  3  v(t-l)  =  8  a 2  E  + 5 o 2  C(VTS)  Shoot w i t h i n Tree  vt(s-l)  =  12  a 2  E  + 5 a 2  C(VTS)  Section within  vts(c-l)  =  24  a  vtsc(m-l)  = 192  vtscm-1  = 239  Variety Tree w i t h i n  Variety  Shoot  Measurements w i t h i n Total  Shoot  ° E 2  + 5 o 2  C(VTS)  + 1 0 o 2  + 1 0  V ° C<VTS) 5  2  + 1 0 o 2  S(VT)  S(VT)  ° S(VT) 2  + 2 0  + 2 0 o 2  T(V)  ° T(V) 2  + 6 0 e 2  V  Table 5, 9 Line number of F denominator  Line Number 1 2  Expected mean s q u a r e s  Source o f V a r i a t i o n 2  0  3  0  E  0  Variety Tree w i t h i n  Variety  3  Shoot w i t h i n Tree  E  Measurements w i t h i n Total  Shoot  V  3 o 2  2 E 2 E  S(VT)  + 6  ° T(V) 2  +  + 3  °  2 2 S ( V T ) ° T(V)  + 3  °  2 S(VT>  + 6  1 8 9 2  V  63  Table 7, 8, 10, 11  Line Number 1 2 3 4 E  . Line number of F denominator  Degrees o f freedom  Expected mean squares  Source o f V a r i a t i o n (t-1)  = 1  2  E  t(s-l)  =10  E  ° E  (d-1)  = 7  E  c  2  (d-D(t-l)  =  7  E  a  2  t(d-lMs-l)  =  70  o  2  tds-1  =  95  a 2  + 8  ° S(T)  + 8  ° S(T)  2  Treatment Shoot w i t h i n  2  2  Treatment +126  E  2 D  Date E  Date x Treatment Date x Shoot Total  E  +12a  2 D T  + 4 8 6 2  T  64  T a b l e 12  Line Number  Source o f V a r i a t i o n Treatment  (t-1)  Shoot w i t h i n Treatment  t(s-l)  Leaf w i t h i n Shoot  ts(l-l)  Disc within  Leaf  Section within  Disc  Measurements w i t h i n Total  Section  Line number of F denominator  Degrees o f freedom  Expected mean squares  2  o 2  E  + 2  ° C(TSLD)  3  a 2  E  + 2  ° C(TSLD)  10  4  tsl(d-l)  = 100  5  0  tsld(c-l)  = 120  E  0  tsldc(m-l)  -240  tsldcm-1  :479  ° E 2  2 E 2 E 0  +  2  2  + 2 o 2  2  + 2  2 E  a  °  + 4 o 2  + 4 o 2  C(TSLD)  + 4 o 2  D(TSL) D(TSL) D(TSL)  2 2 C ( T S L D ) ° D(TSL) + 4  2 C(TSLD)  + 2 4 o 2  + 2 4 o 2  + 2 4  L(TS)  L(TS)  + 4 8 o 2  ° L(TS) 2  + 4 8 o 2  S(T)  S(T>  + 2 1 , 6 2  T  T a b l e 13  Line Number  Source o f V a r i a t i o n  Treatment Shoot w i t h i n  (t-1) Treatment  Section within  Shoot  Measurements w i t h i n Total  Line number of F denominator  Degrees o f freedom  Section  Expected mean squares  a  s(t-l)  V ° C(TS) 5  ° E  + 5 o 2  ° E  + 5  2  st(c-l)  10  stc(m-l)  80  stcm-1  99  2  2  +1 0 a 2  C(TS)  ° C(TS) 2  + 1 0  S(T)  +5 0 8  ° S(T) 2  2  T  66  Table  14  Line Number  Source o f V a r i a t i o n Treatments Tree w i t h i n  Treatment  Line number of F denominator  Degrees of freedom (t-1)  =  3  t(r-l)  =  16  Leaf w i t h i n Tree  tr(l-l)  D i s c w i t h i n Leaf  trl(d-l)  =  "»0  Section within  trld(c-l)  =  80  trldc(m-l)  = 160  trldcm-1  = 319  Disc  Measurement w i t h i n Total  Section  20  =  Expected mean squares 2  2 E  0  + 2  °  4.1  2  S(TRLD) 2  E+"2 " o  o  S(TRLD) ° + 4  r  2  2 E  a  + 2  °  2  S(TRLD) ° + 4  .  2 E  S(TRLD)  2.-2 ° E  S(TRLD)  0  + 4 a  h 0  2 2 D(TRL) ° L(TR) + 8  + 1 6  °  2 R(T)  , + 8o + 16o' D(TRL)* L(TR) R(T) o u  2 2 D ( T R L ) ° L(TR) + 8  2 D(TRL)  2 + 8 0 6  T  67  T a b l e 15  Line Number  Source o f V a r i a t i o n Treatment Tree w i t h i n  (t-1) Treatment  Line number of F denominator  Degrees o f freedom  t(r-l)  3  2  a  24  E  2 ° E  5  E  2 « E  Date  d-1  Date x Treatment  (d-l)(t-l)  =  t(d-l)(r-l)  = 120  tdr-1  = 167  Date x T r e e w i t h i n Total  Treatment  Expected mean squares  15  V  6 < j 2  + 6  R(T)  2 ° R(T)  + 2 8  2 ° D  2 2 a +7o E  D  T  + 4 2 9 2  T  68  T a b l e 16  Line Number  Source o f V a r i a t i o n  L i n e number of F denominator  Degrees o f freedom  Expected mean squares  Block  (r-l)  =  1  Variety  (v-1)  =  1  Treatment  (t-1)  =  3  0  V a r i e t y x Treatment  (v-lXt-1)  =  3  0  B l a c k x V a r i e t y and Treatment  (r-D(vt-l) =  7  Leaf w i t h i n  vtr(l-l)  =  16  0  vtrl(p-l)  =  32  0  Section within Position  vtrlp(s-l)  =  64  0  Measurement w i t h i n  vtrlps(m-l) =  128  vtrlpsm-1  255  Treatment  Position within  Total  Leaf  Section  =  ° E 2  + 2 c 2  2 0  E 2 E  +  2  a  °  + 2  2 E  +  2  a  S ( V T R L P ) '* p(VTRL) ' L ( V T R ) +  E  + 2 a  2 E  + 2  °  E  + 2  °  2  +8(  2  + 1 2 8 o 2  2 2 2 S(VTRLP) "° P ( V T R L ) ° L ( V T R ) +  + 8  2 S(VTRLP)  2 2 P(VTRL) ° L(VTR)  +l t a  +8  +  S(VTRLP) v  + 8  +  4 a  ° P(VTRL)  + 8o u  + 8  2 2 S(VTRLP)* * P(VTRL) 1  2 S(VTRLP)  0  + 1 2 8 8  2 V  T  + 1 5  °  2 RVT  L(VTR)>+16o' RVT  2 2 2 S ( V T R L P ) ° P ( V T R L ) ° L(VTR) + 4  R  2  +6 1 4 8  2 2 2 S(VTRLP) "*° P ( V T R L ) ° L ( V T R )  2 2 a ,-+2a 2  o2  + 3 2  2 ° VT  69  T a b l e 17  Line Number 1 2 3 H 5 6 7 8 9 E  L i n e number of F denominator  Degrees o f freedom  Expected mean squares  Source o f V a r i a t i o n (b-1)  = 1  E  o  +48o  2  (v-1)  = 1  E  o  2  *U8e  2  E  (t-1)  = 3  E  c  2  +248  2  E  (t-D(v-l)  =  5  o  2  (b-D(tv-l)  = 7  E  o .+6o  (d-1)  = 5  E  o .+16e  (d-D(v-l)  =15  E  o  2  (d-lXt-l)  =15  E  o  2  (d-l)(t-l)(v-l)alS  E  o  2  2 E  B  Block v  Variety T  Treatment 3  E  +6o  „+12o  2  T  B  V  V a r i e t y x Treatment B l o c k x Treatment,  2  r  E  +  6  2  °  T B V  Variety 2  r  E  +  1  6  E  2  2  D  Date E  *8o  2  •to  2  + 2„ +2o  22  +  8  °  2  D V  Date x V a r i e t y E ' ^ D T  Date x Treatment E  Date x Treatment x V a r i e t y Date x B l o c k w i t h i n Total  vt(d-lXb-l)  =  HO  vtdb-1  =  95  Treatment  o  2  E  1 0 D  T  V  2  T  V