The Anatomy of the Pigeonpea

S.S. Bisen and A . R . S h e l d r a k e

Research Bulletin No. 5

ICRISAT
International Crops Research Institute for the Semi-Arid Tropics
ICRISAT Patancheru P.O., Andhra Pradesh 502 324. India

March 1981
T h e I n t e r n a t i o n a l C r o p s R e s e a r c h I n s t i t u t e f o r the S e m i - A r i d T r o p i c s ( I C R I S A T ) i s a n o n p r o f i t
scientific educational Institute receiving support f r o m a variety of donors. Responsibility for
the i n f o r m a t i o n i n t h i s p u b l i c a t i o n r e s t s w i t h I C R I S A T . W h e r e t r a d e n a m e s a r e u s e d t h i s does
not constitute e n d o r s e m e n t of or d i s c r i m i n a t i o n against any p r o d u c t by the Institute.

Correct citation: B I S E N , S.S., and S H E L D R A K E , A . R . 1981. T h e anatomy o f the pigeonpea.

Research Bulletin No. 5 Patancheru, A. P., India: International Crops Research Institute for
the S e m i - A r i d T r o p i c s . 2 4 p p .

(This publication was previously issued i n 1977 a s I C R I S A T D e p a r t m e n t a l P r o g r e s s R e p o r t :

Pulse Physiology 1. It reports r e s e a r c h w o r k now completed and is reissued in the present
f o r m because of sustained demand.)
 CONTENTS

Introduction 1

M a t e r i a l s and Methods 1
Fixation
Dehydration and embedding in w a x
Sectioning
Slide preparation
Stains
Clearing of leaves
Stomatal preparations

Anatomical Observations 3

Stems 3
Xylem
Tension wood
Secretory ducts

Leaves 4
Hairs
Stomata
Petioles
Pulvini
Abscission zones

Roots 7
Nodules

Reproductive Structures 7
The development of flowers
The development of pods
Pedicels

Seeds 8

References 9

Figures 10
Abbreviations used in the figures
Abbreviations used in the figure captions
Figures 1-56 11
 Introduction
D u r i n g the 3 y e a r s 1974-77 we studied the anatomy of m o s t of
the tissues and organs of the pigeonpea and, in the course of
this w o r k , have built up a collection of permanent microscope
slides. These are retained in the A n a t o m y L a b o r a t o r y at
I C R I S A T as a reference collection and m a y be consulted by
anyone w h o is interested.
This report contains a b r i e f and p r e l i m i n a r y description
of pigeonpea anatomy. We have studied the anatomy of s e v e r a l
different c u l t i v a r s ; unless otherwise indicated, the f o l l o w i n g
general descriptions apply to all cultivars investigated. We
have not noticed any s t r i k i n g qualitative anatomical differences
among cultivars; no doubt quantitative differences exist, but
these are difficult to establish w i t h anatomical methods i n v o l v -
ing very small samples.
M a n y of the features of the anatomy of the pigeonpea are
s i m i l a r to those of other dicotyledonous plants, described in
standard textbooks of anatomy. We have not attempted to
duplicate these descriptions. Some aspects of the anatomy of
the pigeonpea have been c o v e r e d i n d e t a i l b y D r . P. Venkate-
shwara Rao in his P h . D . thesis u n d e r r e f e r e n c e (no date). A
copy of this thesis is available in the I C R I S A T l i b r a r y .

Materials and Methods

M o s t observations w e r e made o n m a t e r i a l f r o m plants g r o w n i n the f i e l d a t the I C R I S A T

Center; a few samples were also taken f r o m plants grown in pots.

Fixation. The m a t e r i a l was fixed for at least 48 hr either in F P A ( f o r m a l i n 40%:

propionic acid : ethyl alcohol 70%, in the p r o p o r t i o n 5:5:90 by v o l u m e ) or in
glutaraldehyde (3% s o l u t i o n in phosphate buffer, pH 6. 8). In this report Figures 20,
21, 22, 29, 32, 53, 54, 55, and 56 are of m a t e r i a l f i x e d in glutaraldehyde; the
r e m a i n i n g figures are of m a t e r i a l fixed in F P A , unless otherwise stated.

Dehydration and embedding in wax. A f t e r fixation the m a t e r i a l was dehydrated b y

i m m e r s i o n in a series of alcohol solutions, followed by a m i x t u r e of absolute alcohol
and tertiary butanol, and finally brought to pure tertiary butanol. The material was
kept in a m i x t u r e of tertiary butanol and paraffin wax (melting point 60°C) in an
oven for 24-72 hr before infiltration in pure paraffin wax. The material was then
embedded in wax.

Sectioning. Sections of the wax-embedded m a t e r i a l w e r e cut on a C a m b r i d g e R o t a r y

M i c r o t o m e at a thickness of 8-10 y f o r soft m a t e r i a l such as buds, and 12 u f o r h a r d e r
material. Sections of woody stems and roots w e r e cut on a s l i d i n g m i c r o t o m e at a t h i c k -
ness of 12-15 u after the m a t e r i a l had been boiled in w a t e r to expel a i r bubbles and to •
soften i t ; this material was cut directly without embedding in wax. The sections shown
in Figures 3, 4, 5, 6, 7, and 9 w e r e prepared in this way.

Slide preparation. The ribbons of serial sections f r o m the r o t a r y m i c r o t o m e were

floated on w a r m water (50°C) before being arranged on slides, w h i c h had been smeared
w i t h Haupt's adhesive (1 g Knox gelatine, 2 g phenol, and 15 ml g l y c e r o l in 100 ml
water). The slides w e r e kept on a hot plate at 50°C and a few drops of 4% f o r m a l i n
w e r e added to coagulate the gelatine in the adhesive.
The slides w e r e dewaxed in pure xylene for 5-60 m i n before staining. They were
passed through 100%, 90%, 70%, and 50% alcohol solutions and finally water before
staining in aqueous solutions; f o r alcoholic stains, this dehydration was not necessary.
A f t e r staining the slides w e r e dehydrated in alcohol, passed to an alcohol-xylene m i x -
ture, and cleared in xylene before mounting in D P X mountant, Permount or Canada
Balsam. Some of the sections stained w i t h aniline blue and mounted in D P X faded after
several months.

Stains, (a) T o l u i d i n e B l u e . This relatively new stain is polychromatic, giving

different colors w i t h different tissues. Staining was c a r r i e d out in a 0.05%
aqueous solution as d e s c r i b e d by F e d e r and O ' B r i e n (1968).
(b) S a f r a n i n a n d A n i l i n e B l u e . The slides were first stained in safranin
( 1 % in 70% alcohol), washed in 90% alcohol, and counter-stained in aniline blue
( 1 % in absolute alcohol).
(c) T r y p a n B l u e . F o r the detection of m y c o r r h i z a e in cleared root
m a t e r i a l we f o l l o w e d the p r o c e d u r e of P h i l l i p s and H a y m a n (1970). After
c l e a r i n g i n 10% p o t a s s i u m h y d r o x i d e the roots w e r e stained w i t h t r y p a n
blue in lactophenol.
(d) H a e m a t o x y l e n e . The sections w e r e kept in a 3% aqueous solution
of iron alum for 0.5-1 hr, washed, kept in a 1% aqueous solution of haematoxylene
for 0.5-1 hr, washed, and differentiated in i r o n a l u m or in a saturated solution
of p i c r i c acid in 70% alcohol. Some sections were counterstained in safranin, as
described above.
(e) P i c r i c A c i d a n d I o d i n e . Staining f o r p r o t e i n s i n seeds was c a r r i e d
out w i t h a saturated solution of p i c r i c acid in 70% alcohol; after washing they
were stained w i t h iodine in potassium iodine solution to reveal starch. The
sections w e r e mounted in glycerine.

Clearing of leaves. T h e p r o c e d u r e w a s that o f A l o n i and Sachs (1973). The leaves

were boiled in lactic acid before staining in 0.2% lacmoid in lactic acid, followed by
i m m e r s i o n in phosphate buffer (pH 7.5) and mounted in 70% s o d i u m lactate. The
material shown in Figures 11 and 12 was prepared by this method.

Stomatal preparations. Standard methods of preparing impressions of the epidermis

using nail varnish and silicone rubber failed w i t h pigeonpeas, owing to the hairiness

2
of the leaves. H o w e v e r it was possible to o b t a i n s o m e s t o m a t a l i m p r e s s i o n s of the
upper epidermis only, using Quickfix.
An alternative method was devised, using Scotch tape. T h e tap w a s f i r m l y f i x e d o n
both sides of the leaf, and then gently peeled off. T h e e p i d e r m i s stuck to the tape and
in this way good peels could be obtained d i r e c t l y . Even better results were obtained by
keeping the peels in 100% a l c o h o l to e x t r a c t c h l o r o p h y l l , and then passing them through
an alcohol series to w a t e r and staining t h e m in a 0. 05% aqueous solution of toluidine
blue for a few minutes. At this stage the e p i d e r m a l peel c o u l d be separated f r o m the
Scotch tape and mounted on a slide in w a t e r or g l y c e r i n e . The stomata were stained
pink and were clearly visible (Fig. 17). T h i s m e t h o d can be used f o r s t u d y i n g the
d i s t r i b u t i o n and frequency of stomata, but not for the study of stomatal apertures,
since these probably change d u r i n g the p r o c e s s i n g of the e p i d e r m i s .

Anatomical Observations

Steins

T h e internodes of the s t e m develop by elongation of the tissue between the leaf i n i t i a l s

in the apical m e r i s t e m ( F i g . 1).
The p r i m a r y v a s c u l a r tissue of the stem is organized in strands connecting
the nodes; each s t r a n d is associated w i t h a r i d g e on the s t e m . These ridges are clearly
visible even in old, secondarily thickened stems. CoUenchymatous bundle caps underlie
the e p i d e r m i s of the ridges ( F i g . 2).
A l t h o u g h the x y l e m and p h l o e m are o r g a n i z e d into f a i r l y d i s t i n c t strands i n y o u n g
stems, the f i b e r s t o w a r d s the e x t e r i o r of the p h l o e m are not confined to these p r i m a r y
vascular strands but f o r m a continuous r i n g . Immediately outside this r i n g of fibers
is a one-cell-thick ring of thin-walled cells, some of which contain rhomboidal crystals.
T h e cortex in between the bundle caps is usually three or four cells thick. The
vacuoles of some of these cells stain densely w i t h m o s t stains, and in unstained f r e s h
m a t e r i a l t u r n blue soon after sectioning. At a l a t e r stage some of the c o r t i c a l cells
contain reddish anthocyanin pigment in their vacuoles. It is possible that the densely
staining m a t e r i a l in the vacuoles of young c o r t i c a l cells is an anthocyanin p r e c u r s o r .
T h e e p i d e r m i s of the s t e m bears h a i r s of the same types that are found on leaves,
described below.
T h e s t e m thickens as a r e s u l t of the a c t i v i t y of the v a s c u l a r c a m b i u m , w h i c h
p r o d u c e s a continuous r i n g of x y l e m tissue t o w a r d s the inside and p h l o e m t o w a r d s
the outside ( F i g . 2). The continuation of these processes results in thick woody
stems (Figs. 3 and 4). In the outer p a r t of the b a r k there are w e l l - d e v e l o p e d
lenticels (Fig. 5).

Xylem. W i t h i n the x y l e m the vessels are either s o l i t a r y o r i n r a d i a l o r , more rarely,

tangential multiples (Fig. 3). The vessels are surrounded by parenchymatous cells,
and tangential bands o f p a r e n c h y m a r u n between the vessels ( i . e., in an aliform-
confluent pattern). M u c h o f the r e m a i n d e r o f the x y l e m tissue b e t w e e n the m e d u l l a r y
rays consists of x y l e m fibers.

3
 T h e m e d u l l a r y rays are either one c e l l t h i c k (uniseriate) or several cells t h i c k
(multiseriate). M e d u l l a r y rays can be seen in transverse section in Figures 3, 4, and-
8, in tangential longitudinal section in Figure 6, and in radial longitudinal section in
Figure 7.
D u r i n g the vegetative phase the p a r e n c h y m a t o u s cells in the x y l e m , including the
medullary rays, contain large quantities of starch (Fig. 8). These starch reserves can
be detected v e r y s i m p l y in the f i e l d , by applying a few drops of iodine to the cut ends
of stems or branches, which t u r n blue or black if starch is present. D u r i n g the r e p r o -
ductive phase, these starch reserves disappear; but in plants f r o m w h i c h flowers are
continuously removed, the s t a r c h r e s e r v e s in the s t e m are n o t depleted, indicating that
these reserves are m o b i l i z e d as a consequence of pod development.

Tension wood. D u r i n g the monsoon season the strong w e s t e r l y winds bend the stems
and affect the pattern of branching, w i t h a p e r m a n e n t effect on the m o r p h o l o g y of the
plants (see N a r a y a n a n and Sheldrake 1975, chapter 8). T h e bending of the stems
results i n a n a s y m m e t r i c development o f w o o d , w h i c h becomes t h i c k e r o n the w i n d w a r d
(west) side of the s t e m . W i t h i n the x y l e m on this side, bands of fibers w i t h gelatinous
thickenings are formed (Fig. 9). Such thickenings are characteristic of "tension w o o d "
in dicotyledons.

Secretory ducts. W i t h i n the p h l o e m r e g i o n of the stems and also in the outer p a r t of

the p i t h near to the p r i m a r y x y l e m tissue ( F i g . 2) are cells containing material that
stains densely w i t h a l l the dyes we have used; even in unstained sections the contents
of these cells appear reddish b r o w n . In longitudinal sections these cells are seen to be
elongated and joined end to end to f o r m ducts (Fig. 5). These cells differentiate at an
e a r l y stage w i t h i n p r i m a r y tissues (Figs. 1 a n d 10) a n d a r e a l s o f o r m e d w i t h i n s e c o n d -
ary phloem tissue (Fig. 4). They are found in leaves, petioles, roots, flowers, and
pod w a l l s ; indeed they occur throughout the plant body.
W h e n young tender shoots are cut, a d r o p of f l u i d exudes onto the cut surface.
T h i s f l u i d , w h i c h has an e x t r e m e l y astringent taste, is colorless at first, but turns
r e d on exposure to the a i r . The result of this process is that wounds on stems, leaves
pods, etc., become covered w i t h red m a t e r i a l that dries to f o r m a sort of varnish or,
if exuded in greater quantities, transparent globules. This solid material, l i k e the
fluid f r o m which it is derived, has a b i t t e r a s t r i n g e n t taste.
The chemical nature of this material is unknown, but it seems probable that it
contains tannin-like polyphenolic compounds, w h i c h are oxidized on exposure to the
air. Possibly it plays a role in p r o t e c t i n g the plant against pests a n d / o r diseases.

Leaves

The leaves are trifoliate. T h e pattern of the m a i n veins w i t h i n the l a m i n a is c l e a r l y

visible to the naked eye. W i t h i n the areas enclosed by the m i n o r veins are veinlets
containing a single f i l e of tr acheids, m a n y of w h i c h end b l i n d l y ( F i g . 11).

4
 In the m i d r i b of the leaf, the v a s c u l a r tissue in the v e n t r a l h a l f o c c u r s in a c o n t i n -
uous a r c h e d band, w i t h p h l o e m o n the outside and x y l e m w i t h i n . In the v e n t r a l p a r t ,
there are two distinct strands mostly consisting of phloem tissue. The center of the
d o r s a l p a r t of the m i d r i b is occupied by f i b e r s , above w h i c h is a cap of coUenchymatous
cells (Fig. 13). C r y s t a l s are frequently seen in the c o r t i c a l cells i m m e d i a t e l y adjacent
t o the f i b e r s . These are clearly visible in minor veins, especially in polarized light
(Fig. 12). S m a l l e r crystals are found w i t h i n the p h l o e m p a r e n c h y m a .
In the leaf l a m i n a there is a d i s t i n c t palisade l a y e r , and in the l o w e r p a r t of the
leaf a spongy m e s o p h y l l w i t h l a r g e a i r - f i l l e d i n t e r c e l l u l a r spaces ( F i g . 14).

Hairs. The leaves of pigeonpeas are pubescent, m o r e so on the l o w e r than on the

upper surface. At l o w m a g n i f i c a t i o n , t w o distinct types of h a i r can be seen: s i m p l e ,
and glandular (Fig. 15). T h e glandular h a i r s are s p h e r i c a l and y e l l o w i n c o l o r . In
fresh sections of leaves, they resemble s m a l l balloons, distended w i t h yellow fluid
(Fig. 16). W h e n these hairs are damaged, the contents escape and a deflated, colorless
sac r e m a i n s . These sacs appear to contain an o i l y s e c r e t o r y m a t e r i a l , w h i c h is
p r o b a b l y responsible f o r the fragrance of the vegetative p a r t s of the pigeonpeas.
Essential o i l can be collected by the steam distillation of leaves, etc., of pigeonpeas;
it contains a m i x t u r e of compounds including the terpenoid a-Copaene (Gupta et a l .
1969).
These f l u i d - f i l l e d sacs seem to develop f r o m short m u l t i c e l l u l a r glandular hairs
found on young leaves (Fig. 17).
The m e a n frequency of these y e l l o w - f l u i d - f i l l e d glandular hairs on young mature
2
leaves of 10 m e d i u m - and late-duration cultivars was 1 . 9 / m m on the upper and
2
3.2/mm o n the l o w e r surface. The number per leaf declined w i t h age, presumably
o w i n g to the damage and rupture of the h a i r s . The variation in number f r o m leaf to
leaf w i t h i n a cultivar was too great to enable cultivaral differences in hair frequency
to be established.
S i m i l a r s i m p l e and glandular hairs are found on a l l a e r i a l parts of the plants,
except some p a r t s of the f l o w e r s such as petals and stamens. In addition, a further
type of glandular hair is v e r y frequent on pod walls (Fig. 43) b u t i s r a r e l y seen o n
other organs.

Stomata. T h e r e are o v e r 10 t i m e s as m a n y s t o m a t a on the l o w e r than on the upper

surface of the leaves (Table 1).

T a b l e 1 . Frequency of stomata on the upper and lower epidermis of fully

expanded leaves.

2
Number of stomata/mm
Cultivar
Upper surface Lower surface

ICP-1 55 +33.2 667 + 34.4

T-21 43 +44.4 609 + 103.9
HY-3C 19 + 1 7 . 1 447 + 94.3

5
 Stomata in the l o w e r e p i d e r m i s of a leaf are shown in F i g u r e 18. We have not so
far been able to make impressions of this e p i d e r m i s in silicone rubber or other
m a t e r i a l , owing, to hairiness of the surface. However, satisfactory impressions of
the upper epidermis w e r e obtained w i t h Q u i c k f i x ( F i g . 19).

Petioles. The petiole contains a number of distinct vascular strands (Fig. 20) o u t s i d e
w h i c h l i e bands of fibers ( F i g . 21). T h e two "flanges" p r o j e c t i n g f r o m the upper s u r -
face of the petiole contain s m a l l v a s c u l a r bundles. Not u n c o m m o n l y a few of the x y l e m
vessels in petioles w e r e found to be f i l l e d w i t h d a r k staining m a t e r i a l r e s e m b l i n g the
tanninlike material in secretory ducts (Fig. 21). We do not know the reasons f o r this.

Pulvini. At the p r o x i m a l end of the petiole is a s w o l l e n r e g i o n , the p u l v i n u s . S i m i l a r ,

b u t s m a l l e r , p u l v i n i a r e found a t the j u n c t i o n b e t w e e n each leaflet and the p e t i o l e .
The changes in t u r g o r on different sides of these p u l v i n i are responsible f o r the
m o v e m e n t s of the leaves and petioles. The canopy structure of pigeonpeas is con-
stantly changing as a result of these p u l v i n a r m o v e m e n t s . T h e u p r i g h t steep positions
of the leaves at night depend on the geotropic sensitivity of the p u l v i n i , and the m o v e -
m e n t s of the leaves throughout the day are influenced by intensity of the l i g h t f a l l i n g
o n the p u l v i n i a n d a l s o the w a t e r status o f the p l a n t (see N a r a y a n a n a n d S h e l d r a k e 1 9 7 5 ,
chapter 8).
T h e v a s c u l a r tissue is a r r a n g e d in a horseshoe shape in the center of the p u l v i n u s ,
w i t h the x y l e m on the inside; the p h l o e m is surrounded by a band of fibers ( F i g . 22).
As in the stems, leaf veins, and petioles, small crystals (presumably of calcium
oxalate) are found scattered throughout the p h l o e m , in parenchymatous cells; larger
crystals are found in the c o r t i c a l cells adjacent to the phloem fibers.
M o s t of the pulvinus consists of c o r t i c a l tissue. M a n y o f the c e l l s i n the c o r t e x
contain densely staining tanninlike m a t e r i a l w i t h i n their vacuoles ( F i g . 22). It is the
changes in the t u r g o r of these c o r t i c a l cells that are responsible f o r the movements of
the p u l v i n i . In other species, such changes have been shown to be due to active fluxes
of K+ and o t h e r ions.

Abscission zones. In senescent leaves an abscission zone develops at the j u n c t i o n

between each leaflet and the petiole and at the j u n c t i o n between the petiole and the
stem. In each case the abscission zone is p r o x i m a l to the pulvinus.
P r i o r to abscission, cells in the abscission zone d i v i d e , w i t h the plane of c e l l
d i v i s i o n p a r a l l e l to the plane of abscission. The weakening of the w a l l s of these cells
(which in other species has been shown to depend on the production of cellulase and
other h y d r o l y t i c enzymes) results in an easy separation of the two sides of the
abscission zone ( F i g . 23). Consequently the leaflet or petiole f a l l s .
A b s c i s s i o n zones are also f o r m e d at the j u c t i o n between the peduncle of the
inflorescence and the p e d i c e l of f l o w e r s or young pods ( F i g . 24). In pigeonpeas, pods
d e v e l o p f r o m o n l y a s m a l l m i n o r i t y o f the f l o w e r s (see S h e l d r a k e e t a l . 1979); m o s t of
the f l o w e r s abscind infructuously.

6
Roots

T h e p r i m a r y s t r u c t u r e of the roots is usually t e t r a c h ( F i g . 25). Secondary thickening

takes place as a r e s u l t of the a c t i v i t y of the c a m b i u m . In older roots, there are
numerous rhomboidal crystals in cells of the cortex (Fig. 26). As in stems and other
aerial organs, s e c r e t o r y ducts containing tanninlike m a t e r i a l are present w i t h i n the
phloem region.
M y c o r r h i z a e are often present in c o r t i c a l cells of the roots ( F i g . 27) a n d
occasionally f r u i t i n g bodies of the m y c o r r h i z a e can be observed ( F i g . 28).

Nodules. In the y o u n g nodules, g r o w t h occurs f r o m a m e r i s t e m a t i c zone a r c h i n g

around the apical end ( F i g . 29). T h e medulla of the nodules contains numerous
bacterioid-filled cells (Figs. 30 and 31). V a s c u l a r stands are present in the c o r t e x
(Figs. 29 and 30). Sometimes the bacterioidal cells are highly vacuolated, as in
Figure 29, but the nonvacuolate f o r m shown in F i g u r e s 30 and 31 is m o r e usual. We
do not k n o w the cause of this vacuolation.
In some nodules bacterial infection threads are clearly visible in cells near the
m e r i s t e m a t i c zone. These infectio n threads appear to m o v e t h r o u g h the cells ( F i g . 32).

Reproductive Structures

The development of flowers. A n e a r l y stage i n the development o f f l o w e r buds i s

s h o w n in l o n g i t u d i n a l section in F i g u r e 33 and a late stage in F i g u r e 34. E a r l i e r and
l a t e r stages are shown in t r a n v e r s e section in Figures 35 and 36, respectively.
Pollen m o t h e r cells develop and separate f r o m each other w i t h i n the e m b r y o n i c
anther (Fig. 37). Their nuclei undergo meiosis, r e s u l t i n g i n the f o r m a t i o n o f tetrads
containing four nuclei (Fig. 38); then tetrads separate into four distinct cells (Fig. 39),
w h i c h develop into pollen grains (Fig. 40).
We have not studied the process of megasporogenesis in d e t a i l . We think that the
stage s h o w n i n F i g u r e 4 1 represents the cells d e r i v e d f r o m the megaspore m o t h e r
cell by meiosis. One o r m o r e o f these c e l l s gives r i s e t o the e m b r y o sac ( F i g . 42),
and the others degenerate.

T h e development of pods. In the f i r s t 3 weeks after anthesis, the p o d w a l l g r o w s m o r e

r a p i d l y than the young seeds, b u t t h e r e a f t e r u n d e r g o e s l i t t l e f u r t h e r g r o w t h (see
Narayanan and Sheldrake 1975, chapter 4).
The pod w a l l is w e l l supplied w i t h secretory ducts containing tanninlike m a t e r i a l
(Fig. 43). T h e outer e p i d e r m i s of the pod contains stomata. This epidermis bears
simple hairs and globular secretory hairs containing y e l l o w o i l (Fig. 44) s i m i l a r t o
those found on leaves (Figs. 15-17). In addition, there are large numbers of a t h i r d
type of hair seen only occasionally on vegetative organs, with secretory cells towards
the base and a long tubular neck ( F i g . 43). These hairs produce a colorless liquid
exudate, w h i c h in f r e s h m a t e r i a l can be seen in droplets at the tips and on the outside
of the h a i r s .

7
 At the t i m e of anthesis, the ovules are present in an undivided space w i t h i n the
c a r p e l ( F i g . 34), but w i t h i n the 1st week of pod development c r o s s - w a l l s develop between
the d e v e l o p i n g seeds, d i v i d i n g the p o d into locules ( F i g . 46).
D u r i n g the 1st w e e k after anthesis the e n d o s p e r m undergoes r a p i d development
and becomes the p r e d o m i n a n t tissue in the developing seed ( F i g s . 45 and 46); by the
end of the 2nd week there are s t i l l l a r g e amounts of endospermous tissue ( F i g . 48).
W i t h i n the e m b r y o , distinct cotyledons are apparent (Fig. 47). Further development of
the seeds involves r a p i d g r o w t h of the cotyledons and degeneration of the r e m a i n i n g
nutritive tissues.

Pedicels. The pedicels of the flowers contain s m a l l vascular bundles surrounded by a

ring of fibers (Figs. 49 and 50). D u r i n g the 1st w e e k of p o d d e v e l o p m e n t , secondary
thickening begins and the c a m b i u m gives r i s e to a r i n g of x y l e m tissue towards the
inside of the p e d i c e l and to a r i n g of p h l o e m towards the outside ( F i g . 51). Secondary
t h i c k e n i n g continues d u r i n g the 2nd and 3 r d weeks but s l o w s d o w n i n the 4th w e e k ,
after w h i c h little m o r e vascular tissue develops. This thickening results in a great
increase in the c r o s s - s e c t i o n a l area of the vascular tissue supplying the pod ( F i g . 52).
D u r i n g the l a t e r stages of p o d d e v e l o p m e n t the p h l o e m tissue c o l l a p s e s , probably as a
r e s u l t of the m a t u r a t i o n and desiccation of the pod.
T h e p h l o e m f i b e r s , l i k e the x y l e m f i b e r s , contain large amounts of gelatinous w a l l
thickening and do not become heavily lignified.
Some of the cells of the p i t h , at f i r s t t y p i c a l l y t h i n - w a l l e d , begin to show r e t i c u -
late w a l l thickenings by the end of the 2nd week of p o d development. Wall thickening
takes place i n m o r e p i t h cells d u r i n g the 3 r d w e e k , and some of these cells begin to
lignify. B y the t i m e o f m a t u r a t i o n m o s t o f the p i t h cells possess thickened, p i t t e d ,
lignified w a l l s r e s e m b l i n g those of x y l e m tracheids.

Seeds

T h e large cotyledons of the e m b r y o fill m o s t of the seed ( F i g . 53). The seed coat
resembles that of other legumes, w i t h an outer palisade l a y e r of sclereids and a
subepidermal l a y e r of " p i l l a r c e l l s , " between w h i c h are large i n t e r c e l l u l a r spaces
(Fig. 54). A t h i n l a y e r of collapsed parenchymatous cells and the r e m a i n s of the
endosperm underlie the subepidermal cells.
At the h i l u m r e g i o n , a hole in the seed coat leads into a " t r a c h e i d island" ( F i g . 55).
The tissue outside the seed coat at the h i l u m is k n o w n as funicular tissue; this tissue
includes a palisade l a y e r adjacent to that of the seed coat ( F i g . 55).
The parenchymatous cells of the cotyledons contain large starch grains and
numerous protein bodies (Fig. 56). V a s c u l a r strands r u n throughout the ground tissue
of the cotyledons.
A f t e r g e r m i n a t i o n of the seeds, the p r o t e i n bodies and s t a r c h g r a i n w i t h i n the
cotyledons gradually disappear as the r e s e r v e s are m o b i l i z e d into the g r o w i n g
seedlings; the cotyledons are exhausted w i t h i n a week or so.

8
References

ALONI, R., and SACHS, T. 1973. The t h r e e - d i m e n s i o n a l s t r u c t u r e o f the p r i m a r y

phloem system. Planta 113:345-353.

FEDER, N . , and O ' B R I E N , T. P. O. 1968. Plant microtechnique: some principles

and new methods. A m e r i c a n Journal of Botany 55:123-142.

GUPTA, G. L . , NIGAM, S.S., SASTRY, S.D., and C H A K R A V A R T I , K. K. 1969.

Investigations o n the essential o i l f r o m Cajanus cajan ( L i n n . ) Millsp. Perfumery
and Essential O i l R e c o r d 60:329-336.

N A R A Y A N A N , A . , and S H E L D R A K E , A . R . 1975. Pigeonpea physiology. ICRISAT

Pulse Physiology annual report 1974-75, Part I, Patancheru, A. P., India.

P H I L L I P S , J . M . , and H A Y M A N , D . S . 1970. Improved procedures for clearing roots

and staining parasitic and vesicular-arbuscular m y c o r r h i z a l fungi for r a p i d
assessment of infection. Transactions of the B r i t i s h M y c o l o g i c a l Society 55:158-161.

SHELDRAKE, A.R., NARAYANAN, A., and V E N K A T A R A T N A M , N. 1979. The

effects of f l o w e r r e m o v a l on the seed y i e l d of pigeonpeas (Cajanus cajan). Annals
of Applied Biology 91:383-390.

V E N K A T E S W A R A R A O , P. (No date) Developmental and anatomical studies in

Cajanus cajan ( L i n n . ) M i l l s p . Thesis, Sardar Patel University, Vallabh Vidyanagar,
India. 118 p p .

9
 Figures

Abbreviations used in the figures

A A b s c i s s i o n zone O Ovary
AN Anther P Phloem
B Bacterioid P A L Palisade tissue
C A M Cambium PAR Parenchyma
COL Collenchyma PC Pillar cell
COT Cotyledon PET Petal
CP Carpel primordium PG Pollen grain
CRY Crystal PMC Pollen mother cell
E M B Embryo S Stomata
END Endosperm SCL Sclereids
EP Epidermis SD Secretory ducts containing
F Fiber tanninlike material
FB F r u i t i n g body SM Spongy mesophyll
FT Funicular tissue SP Stamen p r i m o r d i u m
GH Glandular hair ST Starch
H Simple hair T Tetrad
LI Leaf initial TAP Tapetum
M Mycorrhizae VB Vascular bundle
MR Medullary ray X Xylem
MZ M e r i s t e m a t i c zone XF X y l e m fiber
XV Xylem vessel

Abbreviations used in the figure caption

A. B. Aniline blue Saf. Safranin

Haem. Haematoxylene T.S. Tranverse section
L. S. Longitudinal section Tol. B. Toluidine blue

10
Fig. 1 : L.S. apical meristem, cv. ICP-1 (Saf. - A . B . ) x 113.
Fig. 2 : T . S . y o u n g s e c o n d a r i l y t h i c k e n i n g s t e m , c v . ST-1
(Saf. - A.B.) x 75.
Fig. 3 H T . S . woody m a i n - s t e m , c v . Hy-3C ( S a f . - A . B . ) x 3 0 .
Fig. 4 : T . S . woody m a i n - s t e m , b a r k r e g i o n , c v . HY-3C
(Saf. - A . B . ) x 113.

11
F i g . 5: L.S. b a r k r e g i o n o f woody stem, showing l e n t i c e l ,
c v . ICP-1 ( T o l . B) x 113.
F i g . 6: T a n g e n t i a l L.S. o f x y l e m i n woodv stem, showing
m e d u l l a r y r a y s , c v . ST-1 ( T o l . B) x 75.
F i g . 7: R a d i a l L.S. o f x y l e m i n woody stem showing m e d u l l a r y
r a y s , cv. ST-1 ( T o l . B) x 75.
F i g . 8: T.S. xylem i n woody stem s t a i n e d w i t h i o d i n e , showing
s t a r c h in m e d u l l a r y r a y s and xylem parenchyma, cv. ST-1 x 75.

12
Fig. 9 : T . S . xylem i n woody stem s h o w i n g g e l a t i n o u s t h i c k e n i n g
( a r r o w ) i n x y l e m f i b e r s , c v . ST-1 ( S a f . - A . B . ) x 3 0 0 .
F i g . 10 : L . S . a x i l l a r y b u d , s h o w i n g s e c r e t o r y d u c t s , c v . T -21
(unstained) x 75.

F i g . 11 : C l e a r e d l e a f s h o w i n g v e i n s ; many m i n o r v e i n s end
b l i n d l y ( a r r o w ) , c v . HY-3C ( s t a i n e d w i t h l a c m o i d i n
l a c t i c a c i d ) x 300.
F1g. 12 : Cleared l e a f photographed in p o l a r i z e d l i g h t showing
c r y s t a l s a s s o c i a t e d w i t h t h e v e i n s . c v . HY-3C x 3 7 5 .

13
F i g . 13 : T.S. l e a f in m i d - v e i n r e g i o n , cv. ST-1
( S a f . - A.B.) x 113.
F i g . 14 : T.S. l e a f l a m i n a , showing p a l i s a d e t i s s u e and
spongy m e s o p h y l l , c v . ST-1 ( S a f . - A.B.) x 375.
F i g . 15 : S u r f a c e v i e w o f edge o f t h e l a m i n a o f a f r e s h l e a f
showing s p h e r i c a l o i l - f i l l e d g l a n d u l a r h a i r s , c v .
HY-3C x 4 5 .
F i g . 16 : T.S. f r e s h l e a f lamina showing s i m p l e h a i r s and
s p h e r i c a l o i l - f i l l e d g l a n d u l a r h a i r s , CV.HY-3C
( u n s t a i n e d ) x 75.

14
F1g. 17 : T . S . f r e s h l e a f lamina showing g l a n d u l a r h a i r s ,
c v . HY-3C ( u n s t a i n e d ) x 450.
Fig. 18 : Peel o f l o w e r e p i d e r m i s o f l e a f l a m i n a showing
s t o m a t a , c v . ICP-6997 ( T o l . B . ) x 4 5 0 .
Fig. 19 : Q u i c k f i x i m p r e s s i o n of upper e p i d e r m i s of l e a f
l a m i n a , s h o w i n g s t o m a t a , c v . Pusa a g e t i x 4 5 0 .
F1g. 20 : T . S . p e t i o l e , c v . T-21 ( T o l . B.) x 38.

15
F1g. 21 : T.S. p e t i o l e , showing v a s c u l a r b u n d l e s . Some xylem
vessels are f i l l e d w i t h densely s t a i n i n g m a t e r i a l
( a r r o w ) , c v . T-21 ( T o l . B) x 113.
F i g . 22 : T.S. p e t i o l a r p u l v i n u s , cv. T-21 ( T o l . B) x 45.
F i g . 23 : L.S. a b s c i s s i o n zone o n p e t i o l e a f t e r t h e a b s c i s s i o n
o f a l e a f l e t . Note c e l l d i v i s i o n s p a r a l l e l t o
s u r f a c e ( a r r o w ) , c v . HY-3A ( T o l . B.) x 7 5 .
F i g . 24 : L.S. a b s c i s s i o n zone at base of p e d i c e l a f t e r t h e
a b s c i s s i o n o f a f l o w e r , c v . HY-3C ( T o l . B.) x 94.

16
F i g . 25 : T.S. s t e l e o f young r o o t showing t e t r a c h arrangement
o f p r i m a r y x y l e m , photographed i n p o l a r i z e d l i g h t ,
c v . ICP-1 ( T o l . B . ) x 300.
Fig. 26 : T.S. s e c o n d a r i l y t h i c k e n e d r o o t , w i t h numerous
c r y s t a l s i n t h e c o r t e x , photographed i n p o l a r i z e d
l i g h t , cv. ST-1 ( T o l . B.) x 38.
F i g . 27 : L.S. c o r t e x o f r o o t , showing m y c o r r h i z a e w i t h i n the
c e l l s , cv. ICP-1 ( T o l . B.) x 450.
F i g . 28 : Squashed c l e a r e d r o o t showing f r u i t i n g b o d i e s o f
m y c o r r h i z a e i n t h e c o r t i c a l r e g i o n ( t r y p a n b l u e ) x 450.

17
F1g. 29 : L.S. r o o t b e a r i n g two n o d u l e s . In t h e n o d u l e on t h e
r i g h t t h e m e r i s t e m a t i c zone can b e seen. The b a c t e r i o i d a l
c e l l s are vacuolated. The c o r t e x o f t h e r o o t c o n t a i n s
m y c o r r h i z a e . c v . ICP-1 ( T o l . B.) x 30.
F i g . 30 : L.S. n o d u l e showing b a c t e r i o i d a l c e l l s , c v . HY-3C
( S a f . - A.B.) x 75.
F i g . 31 : L.S. n o d u l e showing b a c t e r i o i d a l c e l l s , c v . HY-3C
( S a f . - Haem.) x 300.
F i g . 32 : L.S. c o r t i c a l r e g i o n o f n o d u l e showing Rhizobium
i n f e c t i o n threads passing through c e l l s ( a r r o w ) ,
c v . ICP-1 ( T o l . B.) x 4 5 0 .

18
F1g. 33 : L.S. young f l o w e r buds showing stamen, c a r p e l and
p e t a l p r i m o r d i a , cv. ST-1 ( S a f . - A.B.) x 38.
F i g . 34 : L.S. f l o w e r bud soon b e f o r e a n t h e s i s , cv. ST-1
( T o l . B.) x 38.
F1g. 35 : T.S. young f l o w e r bud showing d e v e l o p i n g a n t h e r s
and o v a r y , c v . ICP-1555 (Haem), x 113.
F1g. 36 : T.S. f l o w e r bud soon b e f o r e a n t h e s i s , cv. ICP-1555
( T o l . B.) x 75.

19
F i g . 37 : T.S. young a n t h e r showing p o l l e n mother c e l l s ,
c v . ICP-1555 (Haem.) x 750.
F i g . 38 : T.S. d e v e l o p i n g a n t h e r showing e a r l y t e t r a d s ,
c v . ICP-1555 (Haem.) x 600.
F1g. 39 : T.S. d e v e l o p i n g a n t h e r showing t e t r a d s , c v . ICP-1555
(Haem.) x 600.
F1g. 40 : T.S. a n t h e r soon b e f o r e a n t h e s i s c o n t a i n i n a p o l l e n q r a i n s
c v . ICP-1555 (Haem.) x 300.

20
Fig. 41 : L.S. d e v e l o p i n g o v u l e showing t h e c e l l s d e r i v e d from
t h e megaspore m o t h e r c e l l b y m e i o s i s ( a r r o w ) , c v . ICP-1555
( T o l . B.) x 113.
F1g. 42 : L . S . o v u l e , s h o w i n q embryo sac ( a r r o w ) , c v . ST-1
(Haem.) x 1 1 3 .
F1g. 43 : T . S . 8 - d a y - o l d pod w a l l s h o w i n g g l a n d u l a r h a i r s and
s e c r e t o r y d u c t s , c v . ST-1 ( S a f . - A . B . ) x 7 5 .
F i g . 44 : S u r f a c e o f f r e s h pod w a l l s h o w i n g s p h e r i c a l o i l - f i l l e d
g l a n d u l a r h a i r s , c v . NP(WR)-15 x 1 8 .

21
Fig. 45 : T.S. 7 - d a y - o l d pod, showing t h e endosperm w i t h i n
t h e d e v e l o p i n g seed, c v . ST-1 ( S a f . - A.B.) x 30.
F1g. 46 : L.S. 7 - d a y - o l d pod, showing t h e endosperm w i t h i n
t h e d e v e l o p i n g seed, c v . HY-3C (Haem.) x 3 8 .
F1g. 47 : T.S. 1 4 - d a y - o l d pod, showing d i s t i n c t c o t y l e d o n s
w i t h i n t h e d e v e l o p i n g seed, c v . ST-1 ( S a f . - Haem.) x 15.
F i g . 48. : L.S. 1 4 - d a y - o l d d e v e l o p i n g seed showing endospermous
t i s s u e and p a r t of t h e embryo, c v . ST-1 (Haem.) x 30.

22
F i g . 49 : T.S. p e d i c e l of f l o w e r , showing v a s c u l a r bundles
s u r r o u n d e d by a r i n g o f f i b e r s , c v . ST-1 (Haem.) x 30.
F i g . 50 : T.S. p e d i c e l of f l o w e r , showing p r i m a r y v a s c u l a r
t i s s u e , c v . ST-1 (Haem.) x 113.
F i g . 51 : T.S. p e d i c e l of 8-day-old pod, showing e a r l y stages
of secondary t h i c k e n i n g , cv. ST-1 ( S a f . - A.B.) x 30.
F i g . 52 : T.S. p e d i c e l of 4 9 - d a y - o l d m a t u r e pod, showing
secondary t h i c k e n i n g , c v . ST-1 (Haem.) x 94.

23
F i g . 53 : T.S. seed at end o p p o s i t e t h e h i l u m , showing t h e seed
c o a t and t h e c o t y l e d o n s , c v . T-21 ( T o l . B.) x 45.
F i g . 54 : T.S. seed c o a t showing o u t e r l a y e r o f s c l e r e i d s and i n n e r
l a y e r o f p i l l a r c e l l s , cv. T-21 ( T o l . B.) x 113.
F i g . 55 : T.S. seed i n h i l u m r e g i o n showing f u n i c u l a r t i s s u e ,
t h e opening i n t h e seed c o a t and t h e t r a c h e i d i s l a n d
( a r r o w ) , c v . T-21 ( T o l . B.) x 75.
F i g . 56 : L.S. seed showing s t a r c h g r a i n s and p r o t e i n b o d i e s
( a r r o w ) i n c e l l s o f t h e c o t y l e d o n , cv. T-21 ( s t a i n e d
w i t h i o d i n e and p i c r i c a c i d ) x 300.

24