Chapter 3

Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Rice is a self-pollinated crop, although outcrossing up to 6.8% has been observed in some varieties and under certain conditions (see Sahadevan and Namboodiri 1963). Among wild rices, O. sativa f. spontanea accessions have shown up to 50% outcrossing (Sakai and Narise 1959) and O. longistaminata and O. perennis accessions have shown up to 100% outcrossing (Sakai and Narise 1959; Oka and Morishima 1967). Oka (1988) listed outcrossing rates estimated in various types of wild and cultivated rices (Table 3.1). The Asian forms of the O. perennis complex or O. rufipogon showed a range from 7 to 56%, tending to be higher in perennial than in annual types. The African annual species, O. breviligulata, appeared to have a lower rate, ranging from 3% to 20%. O. sativa showed much lower rates.

3.1 Rice Floral Organs, Pollination and Fertilization Mechanisms

A rice plant bears perfect flowers in single-flowered spikelets borne on a panicle. A flower consists of six stamens, each composed of a two-lobed, four-Ioculed anther, borne oh a slender filament, and a pistil containing one ovule. The short style bears a feathery stigma with two branches. The flower is fully developed and the stigma fully receptive at the time when pollen sheds. At this stage lodicules become turgid and force the lemma and pale a apart. Anther dehiscence and extrusion occur more or less simultaneously so that the stigma of a flower receives pollen from the same flower, hence resulting in self-pollination. Lemma and palea close after about 50 to 90 min (Virmani and Athwal 1973).

Rice pollen grains after shedding from the anther are comparatively short­lived and generally lose their viability within 5 min under ordinary conditions; in some exceptional cases a few pollen grains remain viable for 15 min (Koga et al. 1971). On the other hand, wild rice pollen grains have a longevity of up to 9 min (Oka and Morishima 1967).

Soon after being deposited on the stigma, pollen grains start the germination process. The first step in the process is expansion of the pollen grains by the absorption of liquid from the moist surface of the stigma and the protrusion of the intine through a germ pore (Fig. 3.1). Germination occurs 2 to 3 min after

S. S. Virmani, Heterosis and Hybrid Rice Breeding© Springer-Verlag Berlin Heidelberg 1994

Tab

le 3

.1.

Out

cros

sing

rat

es e

stim

ated

in

wild

and

cul

tiva

ted

rice

spe

cies

by

diff

eren

t m

etho

ds.

(Oka

198

8)

00

0

Tax

a/ty

pe

Ori

gin

Met

hod

No.

of

Res

ults

R

efer

ence

po

pula

tion

s (%

)

Asi

an p

eren

nis

0 P

eren

nial

T

aiw

an

Mar

ker

gene

1

30.7

O

ka (

1956

) t::

T

hail

and

Mar

ker

gene

1

44.0

O

ka

and

Cha

ng (

1961

) ... n ....

Tha

ilan

d Is

ozym

e m

arke

rs

1 (N

E88

) 50

.6

Bar

bier

(19

87)

0 '" '" In

term

edia

te

Tha

ilan

d Is

ozym

e m

arke

rs

1 (C

P20)

55

.9

Bar

bier

(19

87)

S·

(]<

I

Per

enni

al

Indi

a V

aria

nce

rati

o 1

37.4

O

ka

and

Cha

ng (

1959

) ~

Sri

Lan

ka

Var

ianc

e ra

tio

2 22

.4-2

6.5

Sak

ai a

nd N

aris

e (1

959)

0 n ::r

A

nnua

l In

dia

Var

ianc

e ra

tio

1 21

.7

Ok

a an

d C

hang

(19

59)

I'> 2.

Indi

a V

aria

nce

rati

o 3

16.6

-33.

9 S

akai

and

Nar

ise

(196

0)

'" i3 In

dia

Mar

ker·

gen

e 1

7.9

Roy

(19

21)

'" T

hail

and

Isoz

yme

1 (N

E4)

7.

2 B

arbi

er (

1987

) I'>

t:I

C

o W

eedy

In

dia

Var

ianc

e ra

tio

2 17

.3-2

0.6

Ok

a an

d C

hang

(19

59)

::t:

'<

brev

ilig

ulat

a A

fric

a V

aria

nce

rati

o 2

3.2-

19.7

M

oris

him

a et

al.

(196

3)

cr- :=!.

Co

Indi

a M

arke

r ge

ne

34

0-6.

8 B

utan

y (1

957)

0

0

sati

va

0 0

Indi

ca

Afr

ica

Mar

ker

gene

2

0-1.

1 R

ober

ts e

t al

. (1

961)

C

o

'"ti

.... In

dica

T

aiw

an

Mar

ker

gene

4

0.1-

0.3

Ok

a (u

npub

l.)

0 Co

Japo

nica

T

aiw

an

Mar

ker

gene

5

0.6-

3.9

Ok

a (u

npub

l.)

t::

n :to

Indi

ca

Sri

Lan

ka

Var

ianc

e 1

3.6

Sak

ai a

nd N

aris

e (1

960)

0 t:I

'"ti

.... ~ g. 0 '" S·

::Id

('i.

0

Rice Floral Organs, Pollination and Fertilization Mechanisms 81

@ 1

2 4

Fig.3.1. Pollen germination and pollen tube penetration in rice. (Namai 1987). 1 Beginning of pollen germination; 2-3 three nuclei transferring from pollen to pollen tube together with whole pollen contents; 4 pollen with pollen tube penetrating into stigma with whole pollen contents

pollination. Two generative and a vegetative nuclei move into the pollen tube together with the pollen cytoplasm 3 to 5 min after pollination. After all the contents of a pollen grain move into the pollen tube, the tube makes its way between the stigmatic papillae into the tissues of the style. Only the distal part of the tube has living cytoplasm. About 9 h after pollination the pollen tube reaches the micropyle of the nucellus (Namai 1987).

Double fertilization starts after 9 to 12 h of pollination and finishes com­pletely 18 to 24 h after pollination. The tip ofthe pollen tubes penetrates into the embryo sac through a synergid, then it bursts and gives off the generative nuclei. One of them fertilizes the egg cell and the other fertilizes the polar nuclei. The fertilized polar nuclei is the primary nucleus of the endosperm, which begins cell division soon after fertilization. The fertilized egg undergoes embryonic develop­ment on the next morning (Namai 1987). Most of the endosperm cells are developed by only 4 days after fertilization and embryo development takes about 10 days t6 complete. Within 5 to 6 days after fertilization, the ovary grows longer and becomes the same length as the mature grain. It is possible to determine whether florets are fertilized or not 2 days after flowering and pollination. After about 15-16 days following fertilization, the young seed reaches the maximum width. The grain thickness then increases slowly until about 25 days ~fter fertilization.

Ripening of rice seeds involves the following three stages (cf. Namai 1987):

1. Milk grain stage - after 5-7 days of fertilization. The content of the young grain is a white liquid. All grains are green. The top of the panicle during the milk stage bends gently in an arc.

2. Dough grain stage - after 10 to 20 days (in the tropics) and 10-30 days (in temperate regions). The milky white liquid of the grain turns soft, and

82 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

gradually becomes a hard dough. The panicle during the dough stage bends completely from the neck.

3. Mature grain stage - after 25-35 days (in the tropics) and 40-50 days (in temperate regions). Grain color in the spikelets begins to change from green to yellow. When 90-100% of the filled spikelets turn yellow, the mature grain stage is complete. All grains become yellow and hard and the panicle arches further.

Seed sterility percentage is increased strictly by a low temperature during the 2 to 4 days after flowering and also by the surface drainage at full heading time.

Knox and Singh (1987) reviewed new perspectives in pollen biology and fertilization in crop plants. According to them, the role of pollen grains is to produce and transfer the sperm cells or their progenitors down the growing pollen tube to the embryo sac where fertilization occurs. Recent research has indicated that during pollen deVelopment several haploid, genome-specific genes are express~d. These genes are not expressed in any other tissues of the diploid sporophyte (Mascarenhas et al. 1985). These genes control steps in pollen­specific functions, for example, microspore and pollen development, germin­ation and tube formation, sperm cell formation and stigma/style recognition and penetration.

3.2 Natural Outcrossing Mechanisms in Rice

As mentioned earlier, cultivated rice cultivars show very limited natural out­crossing. However, a wide range (0-44%) of natural outcrossing has been observed on male sterile plants/lines (Stansel and Craigmiles 1966; Athwal and Virmani 1972; Carnahan et al. 1972; Azzini and Rutger 1982). In large-scale hybrid rice seed production plots in China, natural outcrossing on male sterile lines has been reported up to 74%, with a median value of25-35% (L. P. Yuan, pers. comm.). Xu and Li (1988) reported outcrossing rates ranging from 14.6 to 53.1 % in various experiments conducted at Changsha, Hunan, China. Vari­ability in extent of natural outcrossing on male sterile lines of rice can be attributed to variations in flowering behavior, floral characteristics of male sterile and pollen parents, and variations in environmental factors.

Some plant characteristics viz., plant height, flagleaf length and angle, panicle exsertion, etc., also affect natural outcrossing. All these factors are described and discussed in the following paragraphs.

3.2.1 Plant Characteristics in Relation to Outcrossing

A good panicle exsertion in a male-sterile parent would expose a higher number of spikelets for outcrossing compared to a male-sterile line showing incomplete

Natural Outcrossing Mechanisms in Rice 83

panicle exsertion. Similarly, incomplete panicle exsertion in a pollen parent would result in lower pollen release into the air. Therefore, a good panicle exsertion in both seed and pollen parent in a hybrid seed production plot is essential to attain high outcrossing rates.

Chinese rice scientists suggest that a small and horizontal flagleaf is useful to enhance the outcrossing rate compared to a long and erect flagleaf. Since the number of panicles per square meter and the number of spikelets per panicle are the main seed yield components, high tillering capacity and larger panicles with many spikelets should be the basic traits of the seed parent.

Synchronization of days to flowering of seed and pollen parent is the key to attaining good outcrossing on the seed parent. Chinese scientists (Yuan 1985; Xu and Li 1988) have outlined the criteria for synchronization of flowering in hybrid seed production plots (Fig. 3.2). The procedure involves staggering seeding dates of the male and female parents, sowing the male parent three times to extend the duration for which pollen is available and predicting and adjusting flowering dates. Seeding date is usually determined by leaf age, effective accumulated temperature (EAT), and growth duration. For example, parents of the rice hybrid Shan Van -2 are Zhen Shan 97A and IR24. EAT of IR24 from seeding to initial heading is 1 133°C, and that of Zhen Shan 97 A, is 791°C. The difference in EAT of the two parents is 342°C. IR24 is seeded first and when its EAT reaches 342°C, that is the date for seeding of Zhen Shan 97 A (Xu and Li 1988). Chinese scientists have also identified management practices which can adjust difference in flowering of male and female parents. For example, if one of the parents reaches panicle initiation stage earlier, treating this parent with quick releasing N fertilizer and spraying the later parent with potassium dihydrogen phosphate, adjusts development differences of 4-5 days in flowering.

Blooming period

1, 2, 3, 4,... Days after panicles emerged

~---------------------~ , ,

1 2 3 4:5 6 7 8 910 11 ~2 IR26 first sowing I I I I j I I I I I I I; I

, , , , 12[345678 9~0111213

IR26 second sowing I I ~ I I I I I I I I I I ,

IR26 third sowing ; 1 2 3 4 5 6 7 8 9 10 1112 13 ~ I I ! ! !! !!! I I I

,

V20A 1 2 3; 4 5 6 7 8 9 10 :1112 13 14 I I I , I I I I ! ! , I I I I

,_ - - - - __ - - - __ - - - - __ - - _.1

Fig. 3.2. The synchronization offtowering period ofthe male and female parents of Wei­You 6. (Yuan 1985)

84 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Draining water from the field delays the panicle development in the male parent, and higher standing water speeds up the panicle development (Xu and Li 1988).

Rutger and Carnahan (1981) discovered a recessive gene for the elongated uppermost internode (eui) in Japonica rice which effectively resulted in a recessive, tall, plant type. The gene produced a near-doubling in length of the uppermost internode, a 12% increase in panicle length and little or no effect on other internodes or plant characteristics. The eui gene, can be incorporated in the pollen parent of a hybrid where a semidwarf F 1 is desired. The tall paternal plant type is desirable for wind-blown pollen dispersal onto semidwarf female plants and the resulting hybrid plants would be semidwarf, unlike the usual case of tall hybrids from semidwarfs by tall crosses. The increased height due to the eui gene would also permit a co-mingling of the hybrid parent seed stocks to maximize crossing and would facilitate mechanical removal of the parent before mass harvest of the commercial hybrid seed. Virmani et al. (1988) incorporated the eui gene, identified by Rutger and Carnahan (1981), into Indica rice IR50 through backcrossing.

3.2.2 Flowering Behavior in Relation to Outcrossing

The flowering process in rice has been described by Rodrigo (1925) and Parmar et al. (1979b). Rice panicles emerge from the flagleaf 24-36 days after pri­mordium initiation irrespective of the duration of varieties. The degree of synchrony of flowering varies widely, ranging from highly protracted to highly synchronized behavior. The majority of the early maturing cultivars were relatively more synchronized in flowering as compared to late types (Parmar et al. 1979b). The period of flowering varies from 3-10 days (Grist 1953; Chandra­ratna 1964; Parmar et aI. 1979b).

In North-Western India, opening of the spikelet or blooming in rice starts as early as 083Q h and in a fertile cultivar it is terminated by about 1200 h on normal days during the month of September; however the termination of blooming was delayed up to 1500 h in November when days are somewhat cooler. In tropical conditions, blooming also usually begins at 0830 h and is terminated bt;tween 1100 h to 1200 h in Indica rice cultivars; a Japonica ms line: Wu lOA, however, bloomed only for an hour between 1000 h to 1100 h at IRRI (Fig. 3.3). Under similar conditions some male sterile lines bloomed up to 2100h.

Peak blooming in rice occurs between 1000 and 1030 h irrespective of the spikelet morphology (Parmar et al. 1979b). The majority of rice cultivars initiated blooming at 1000-1100 h in Northwest Indian conditions. At IRRI also, peak blooming in some rice cultivars was found to occur between 0930 and 1030 h. Early duration rice cultivars tended to initiate blooming earlier than the late duration rice cultivars (Table 3.2). Normally, rice blooms only once in a day irrespective of the weather conditions. Parmar et al. (1979b) found two cultures viz., IARI 6193-B, IARI 7216 to have two flushes in a day, the peak of the first flush between 1000 and 1030h and the second between 1730 and 1800h.

Natural Outcrossing Mechanisms in Rice 85

Frequency of blooming florets (%)

~ b =- ~:~~ I 40 . --- -- 978

30

20

10

0

VAl. A d P 203A I 40 P2038

30

20

10

0

:1 e Wul0A I 577A 40 . ,

5778 , , Wul08 ,

30

20

10

0 9 11 13 15 17 19 21 9 11 13 15 17 19 21

Time (hour)

Fig. 3.3. Pattern of anthesis in six eMS and maintainer lines of rice at IRRI

The period from opening to closing of florets is designated as the blooming duration. Rice cultivars were found to bloom for 46-93 min and this trait was strongly influenced by environmental conditions at the time of flowering (Virmani and Athwal 1973). Parmar et al. (1979b) observed the blooming duration to range from 28-78 min. It was 28-35 min in early varieties and 50-70 min in long duration varieties.

Grist (1953) stated that duration of blooming depended much on the pace of pollination. Delay or failure of pollination for instance had been reported to prolong the blooming interval in rice and wheat. Saran et al. (1971) reported that duration of floret opening was positively correlated with the percentage of sterility. At IRRI, several cytoplasmic male-sterile lines showing complete pollen sterility were found to bloom longer than their corresponding isogenic main­tainer lines possessing normal pollen fertility (Table 3.3). Parmar et al. (1979a)

86 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Table 3.2. Frequency of varieties with varied times of blooming initiation in relation to duration of varieties. (Parmar et a1. 1979a)

Group as per Days to Frequency of cultures at various times of duration flowering initiation of blooming (a.m.)

9:00 9:30 10:00 10:30

Very early 80 3 1 6 8 Early 90 12 56 125 108 Semi-early 100 6 42 139 91 Medium early 110 1 4 41 80 Medium late 120 1 14 22 Semi-late 130 1 Late 140 Very late 150

Total 22 105 325 310

Table 3.3. The duration of opening of florets of six eMS lines of rice and their maintainers. (Young 1983)

Line

97 V20 Yar Ai Zhao P 203 WulO 577

Duration of opening of florets (min)

A B

141 ± 36 56 ± 15 141 ± 45 37 ±4 92 ± 15 61 ± 11

124 ± 30 33 ±4 37 ± 6 48 ± 2 72 ± 16 58 ± 10

11 :00 12:00

55 41 53 2

149 2

Later Total

18 301 278 126 92 42

18 74 11 11

29 942

also reported that ethrel induced male-sterile plants of rice showed longer duration of blooming than the fertile plants. They also noted the tendency of stigma in the sterile florets to protrude out well to intercept airborne pollen. However, observations made at IRRI indicate that the exsertion of the stigma is a genetic trait, and not all male-sterile lines possess this trait; however, it can be induced through specific breeding efforts. Besides sterility, there are several other factors that could prolong the period of glume opening. For instance, changes in morphology or angle of lemma and palea could prolong the glume opening period (Gill et al. 1969; Narhari and Bora 1963; Jachuck and Sampath 1969).

In addition to duration and angle of floret opening, synchronization of anthesis of female and male parents is extremely important. At IRRI we observed certain eMS and maintainer lines showing synchronous anthesis while others showed nonsynchronous anthesis (Fig. 3.3).

Natural Outcrossing Mechanisms in Rice 87

3.2.3 Mechanism and the Angle of Floret Opening in Rice

Lodicules are known to play an important role in glume opening. Parthasarathy (1927) indicated that mechanical puncturing of lodicules prevented spikelet opening. Kadam (1933) inferred from his experiments that the extension of filaments was primarily responsible for glume opening. Parmar et al. (1979b), however, reported that simultaneous action oflodicules, filaments, anther, ovary and stigma helps the glumes to open in rice. According to them, the inner floral organs such as filaments and stigma on getting turgid exert lateral pressure, of low magnitude, on the linear joints of lemma and palea, while lodicules exerted leverage pressure at the basal joint of the lemma and palea and as a result lemma and palea get unlocked a little. Simultaneously, anthers tend to protrude and thereby help increase the angle (250 -350 ) of opening. Short, coarse spikelets with thick anthers, for instance, had wider angles, whereas the long slender spikelets with narrow anthers had narrow angles. Empirical observations made on some male-sterile and maintainer lines at IRRI suggested that the spikelet opening percentage was perhaps associated with the extent of development of vascular tissue in the lodicules.

With the opening of the glumes anthesis also ends. All organs except stigma and style lose their turgidity and, hence, the leverage force necessary to keep the florets open no longer exists and the flower closes. Wide opening of the glumes for long intervals in the male sterile parent enables the stigma to intercept with ease the airborne pollen. Wide and prolonged opening of glumes in the pollen parent allows the anthers to protrude well and release their pollen fully into the air. All wild and cultivated rices are wind pollinated, although flowers of some scented varieties attract bees with an aroma (Oka 1988). Rice pollen grains are short-lived, remaining functional only for 3 min after being emitted in O. sativa (Nagao and Takano 1938). Pollen grains of a perennial wild rice (0. rujipogon, W120 from India) remained alive more than twice as long as those of a culti­var (Oka 198,8). Possibly, wild rices showing a higher rate of outcrossing have a higher disseminating ability of their pollen grains than do cultivars (Oka 1988).

From the foregoing review it is apparent that wide variability exists in rice for components of flowering behavior; there is an imperative need to determine the ideal degree of synchronization, the period of flowering and weather conditions that would facilitate a high percentage of outcrossing for successful commercial hybrid seed production. Parmar et al. (1979b) observed that the time of blooming initiation varied with the grain type to some extent. Rice varieties with long, bold spikelets showed flowering initiation somewhat later than rices with medium and short, bold spikelets. Blooming duration in the female parent was an important floral attribute for commercial hybrid seed production. An ideal female parent in the commercial hybrid rice seed produc­tion programs should have relatively early flowering initiation, and long duration of blooming. Since the genetics ofthese traits is not understood clearly, there is a need to conduct such studies to support the breeding program.

88 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

3.2.4 Floral Traits Influencing Outcrossing in Rice

The most important floral trait influencing outcrossing in rice is male sterility. Male-fertile plants show very little, if any, outcrossing due to the self-pollinating nature ofthe rice flower. Extent of outcrossing on a male-sterile line is influenced by its floral traits (viz., stigma size and exsertion, length of style, and angle and duration of spikelet opening) and those (viz., anther size, pollen number per anther, filament length, and duration of spikelet blooming) of the pollen parent. Significant varietal differences for these traits exist in cultivated and wild rices (Copeland 1924; Sampath 1962; Oka and Morishima 1967; Virmani and Athwal 1973; Lyakhovkin and Singil Din 1975; Parmar et al. 1979c; Virmani et al. 1980; IRRI 1983b; Virmani and Edwards 1983). The range of variation for these traits and the sources of desirable floral traits as reported in the literature are given in Table 3.4. Suh (1988) also reported the significance of a multiple pistillate (polycaryoptic) trait in increasing hybrid rice seed production.

Cultivated rices showed smaller anther and stigma sizes than wild rices (Table 3.5). However,. the anther/stigma ratio in the cultivated types was invariably higher than in the wild species. Generally, cultivated rices showed lower stigma exsertion rate compared to wild rices, with the exception of O. staffii and O. rujipogon (E. Africa) most of the wild rices showed 75-100% stigma exsertion, indicative of open pollination. These observations suggest that with domestication and shift from vegetative to the sexual mode of

Table 3.4. Range of variation and varietal sources possessing maximum value of floral traits influencing outcrossing in rice. (Virmani and Edwards 1983)

Trait Range of variation

Stigma length (mm)

Stigma breadth (mm)

Style length (mm)

Extent of stigma exsertion (%) Anther length (mm)

Anther breadth (mm)

Filament length (mm)

Pollen number/anther

a Parmar et al. (1979b). b IRRI (1983a).

Cultivated, Wild, Cultivated, Wild, Cultivated,

Wild, Cultivated, Wild, Cultivated, Wild, Cultivated, Wild, Cultivated, Wild, Cultivated,

o Virmani and Athwal (1973). d IRRI (unpubl.).

0.2-2.6 0.3-5.0 0.2-1.0 0.4-1.3 0.6-3.2

1.2-2.3 0.2-87.8

0-100 0.9-3.7 1.6-5.4 0.2-0.9 0.2-1.0

0-2.3 0-14

463-3833

Varietal source possessing maximum value

IARI 6637, IARI 17332, IARI 10979Aa Genetic stock 6209-3b

Not reporteda

Genetic stock 6209-3 IARI 7332, IARI 10754, IARI 10871, IARI 10979A, IARI 10205" Oryza sativa f. spontaneaa

BPI 76-2 (n.s.)O Genetic stock 6209-3b

IARI 5819, IARI 5823a Oryza australiensis a IARI 5819, IARI 5823a

Oryza sativa f. spontaneaa

Not known Not known IRI 3526-41-1-2d •

Tab

le 3

.5.

Var

iati

on i

n si

ze o

f an

ther

, st

igm

a an

d th

eir

rati

o (A

jS)

in r

elat

ion

to t

he d

egre

e o

f st

igm

a ex

sert

ion

in c

ulti

vate

d an

d w

ild r

ice.

(P

arm

ar e

t al

. 19

79b)

Mat

eria

l N

umbe

r A

nthe

r S

tigm

a A

nthe

rj

Fre

quen

cy o

f ty

pes

wit

h st

igm

a S

easo

n an

d pl

ace

of s

tudy

of

si

ze

surf

ace

stig

ma

exse

rted

to

the

leve

l o

f cu

ltur

es

mm

2 m

m2

rati

o 1

5%

50

%

75%

10

0%

Tot

al

A.

Cul

tiva

ted

rice

: 1.

N

.E.

Indi

an C

olle

ctio

n (R

abi

1973

, 44

65

1.08

8 0.

403

2.69

9 7.

32

2.26

0.

61

0.25

10

.44

Adu

thur

ai)

2.

N.E

. In

dian

Col

lect

ion

(Rab

i 19

71,

942

1.05

6 0.

422

2.50

2 16

.67

6.79

1.

00

0.10

24

.64

IAR

I D

elhi

) 3.

O

ther

var

ieti

es (

Rab

i 19

72 a

t di

ffer

ent

262

0.98

9 0.

558

1.77

2 15

.65

9.92

1.

15

26.7

2 ce

nter

s)

4.

F 1

hybr

ids

(Rab

i 19

72 a

t di

ffer

ent

cent

ers)

11

45

0.92

2 0.

661

0.39

5 19

.48

8.21

3.

76

1.20

33

.01

B.

Wil

d re

lati

ves

of

rice:

1.

O

. ru

fipog

on (

E.

Afr

ica)

(R

abi

1971

) 1

1.62

0 0.

600

2.70

0 V

ery

rare

exs

erti

on o

f st

igm

a 2.

O

. st

apfi

i (R

abi

1971

) 1

1.68

0 0.

800

2.10

0 50

.00

50.0

0 10

0.00

3.

O.

aust

rali

ensi

s (R

abi

1971

) 1

4.32

0 4.

100

1.05

4 75

.00

25.0

0·

100.

00

4.

O.

niva

ra (

Rab

i 19

71)

15

1.71

0 1.

760

0.97

2 25

.00

50.0

0 25

.00

100.

00

5.

O.

sati

va f

. sp

onta

nea

(Rab

i 19

71)

65

2.44

0 2.

613

0.93

4 50

.00

50.0

0 10

0.00

6.

O

. gl

aber

rim

a (R

abi

1971

) 8

0.96

0 1.

404

0.68

4 50

.00

50.0

0 10

0.00

7.

O

. of

fici

nali

s (R

abi

1971

) 1

2.10

0 3.

200

0.65

6 10

0.00

10

0.00

8.

O

. al

ta (

Rab

i 19

71)

1 1.

100

2.07

0 0.

531

100.

00

100.

00

9.

O.

rufip

ogon

(S

udan

) (R

abi

1971

) 1

0.88

0 2.

160

0.40

7 10

0.00

10

0.00

• V

aria

tion

bet

wee

n pl

ants

wit

hin

spec

ies.

Z

I» .... ~ .... e:. 0 ~ .... ('

) .... 0 CIl S· ()

Q ~

('I)

(') ::r

I» 2.

CIl 3 CIl S· !f ('I

)

00

\0

90 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

reproduction, the cultivated rice had become adapted to self-pollination (Parmar et al. 1979b). Among the wild rices, O. rujipogon and O. sativa f. spontanea, by virtue of their close genetic affinity to cultivated rices, it might be of immediate value in breeding for increased outcrossing potential in rice.

Oka (1988) stated that outcrossing in rice depends upon the capacity of stigmas to receive alien pollen before self-pollination and the capacity of anthers to emit much pollen to pollinate other plants in the proximity. The former capacity would be a function of the time interval from flowering to pollen emission, stigma size and extrusion of stigmas from the flower as conditioned by the style plus stigma length. The latter capacity would depend on the number and disseminating ability of the pollen grains. Oka and Morishima (1967) made an attempt at indirect estimation of outcrossing rate by combining three measurements [viz., time interval from flower opening to pollen emission (T), the stigma plus style length (S), and anther length (A)] to constitute a dis­criminaltt formula X = T + 0.78S + l.31A. The X score was found to be well proportiomil to the outcrossing rates directly estimated in a part of the strains, and a regression equation was obtained to convert the X values to outcrossing rate.

Parmar et al. (1979c) reported a distinct positive correlation between stigma surface and the frequency of varieties with exserted stigma. However, stigma surface tended to show a negative correlation with anther size. These results indicate that improvement in floral traits of female and male parents has to be done separately to increase outcrossing potential in rice in relation to hybrid rice breeding and seed production programs.

Kato and Namai (1987a) also studied inter-varietal variations for floral characteristics viz., number of protruding stigmata per spikelet and pistil length for seed parent and number of residual pollen grains in the anther protruding stage. They observed that tropical Japonica rices possess the highest mean number of protruding stigmata (0.517) and pistil length (1.84 m). On the other hand, most of the temperate Japonicas showed a smaller number of protruding stigmata. Some of the temperate Japonica local rices in Japan are designated as "Majiri" (meaning contamination) and natural outcrossing is considered a cause of the contamination. Such varieties were found to have a larger number of protruding stigmata. Therefore, Kato and Namai (1987a) considered that stigma protrusion contributes to a high natural outcrossing rate in rice. Tropical Japonica rices were also found to possess a higher residual pollen number compared to temperate Japonicas. Therefore, Kato and Namai (1987a) con­cluded that floral characteristics of temperate Japonicas can be improved by introducing promising floral characteristics (viz., protruded stigma, higher residual pollen) from tropical Japonicas to attain a high outcrossing rate.

Inheritance studies (Virmani and Athwal 1974) for floral traits, such as anther length, stigma length and stigma exsertion in rice, indicated that these traits were governed by polygenes. Huang and Huang (1978) reported that stigma exsertion was dominant, partially dominant, or recessive depending on the cross. They also found a negative correlation between stigma exsertion and pollen fertility. Similar observations were also made at IRRI while improving

Natural Outcrossing Mechanisms in Rice 91

stigma size and stigma exsertion in rice. Hassan and Siddiq (1984) reported that long anthers were monogenically dominant over short anthers and fully ex­sertedstigma were dominant over partially exserted stigma.

Virmani and Athwal (1974) found both additive and non-additive effects to be important in the inheritance of floral traits in rice. Prevalence of duplicate­type epistasis was considered a barrier to fixing these traits at higher levels of manifestation through conventional breeding. Therefore, recurrent selection in biparental progenies was proposed by Virmani and Athwal (1974) to improve these traits. Attempts to transfer the long stigma (5 mm) trait from a wild rice genetic stock 6209-3 (IRRI 1983) into the genetic background of some elite maintainer lines have been only partly successful. The undesirable linkage between long exserted stigma of wild rices and improved agronomic character­istics of cultivated rices is quite strong and needs to be broken to incorporate these traits into selected genotypes. Taillebois and Guimaraes (1988) used two approaches (viz., successive backcross and pedigree selection and recurrent selection procedures) to transfer long stigma of the wild species o. long­istaminata to o. sativa. Results indicated that

1. the long and exserted stigma of o. longistaminata is a dominant trait; 2. stigma length of o. longistaminata was recovered in less than 10% of the

plants of a backcross, while the rate was 5-10% in the F 2 generation of a backcross plant;

3. all plants in BC I and BC2 with a stigma length of o. longistaminata were partially or completely male-sterile and seed shattering was high; the sterility and seed shattering problem was, however, considerably reduced in the BC3

generation onwards.

3.2.5 Natural Outcrossing Mechanism in Rice

The success of hybrid seed production in rice depends on the deposition of a sufficient number of pollen grains on the stigma lobes of each spikelet of the seed parent. This is achieved through sufficient pollen flow from the pollen parents to the seed parents:ln this context, a good understanding of the flowering behavior and floral and other morphological traits of the seed and pollen parents is important. Natural outcrossing is also affected by the macro- and microenviron­ment in which the seed and pollen parents are grown. Namai and Kato (1987b) determined the number of pollen grains deposited on the stigma to assure seed set of a male-sterile line and concluded that three to four pollen grains are practi­cally sufficient for seed set on a male-sterile line. With regard to flowering duration of the spikelets, 30 min or more are required to deposit two or more pollen grains on the stigma lobes of more than 80% of spikelets in a Japonica male-sterile line (Nihonmasari) used for the studies. A considerable number of spikelets could set seed when only one pollen grain was deposited on the stigma lobe (Table 3.6).

The number of pollen grains deposited on the stigma is affected by the pollen load in the air. Namai and Kato (1987a) established simple methods for

92 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Table 3.6. Effect of stigma exsertion on seed setting percentage of a Japonica eMS line Reimei. (Namai and Kato 1988)

Kind of spikelet Total Spikelets Seed set (%)

Filled Unfilled

With exserted stigma 37 20 17 54.1 Without exserted stigma 218 69 149 31.7

X2 = 7.02*.

estimating airborne pollen grains per liter. A rough estimate of the airborne pollen grains was made from the number of pollen grains per cm2/h (Y) on vaseline-coated glass slides of the Durham's pollen sampler (Fig. 3.4), by means of the following formula

N = 0.0291 Y + 0.1699,

where N = number of pollen grains per liter of air in the flowering paddy field and Y = number of pollen grains per cm2 of slide surface exposed in the flowering paddy field for an hour.

Subsequently, a rotary pollen sampler was devised which simplified proced­ures for estimating airborne pollen grains per liter (Namai and Kato 1987a). Studies conducted by using this pollen sampler showed that anther length did not correlate with percentage or number of residual pollen grains per anther exserted from the spikelets (Kato and Namai 1987a). However, residual pollen grains per anther and blooming spikelets per unit area directly affected pollen shedding (Kato and Namai 1987b). Therefore, pollen parents for hybrid rice need numerous residual pollen grains per exserted anther and a large number of spikelets constantly blooming per plant (Namai and Kato 1988).

According to Namai and Kato (1988), wind velocity during flowering time of the spikelets was correlated significantly with the number of airborne pollen grains per liter in the paddy field (r = 0.868**) as well as seed set percentage (r = 0.827*). The number of airborne pollen grains per liter was also correlated with the seed,set percentage of the eMS seed parent (r = 0.918**). The daily nutpber of airborne pollen grains per liter was correlated with the number of outcrossed pollen grains per stigma lobe of a spikelet (r = 0.968**) as well as seed set percentage (r = 0.978*). Namai (1987) further concluded that wind velocity of 2-3 m/s and more than 15 airborne pollen grains per liter per day must be sufficient for economical hybrid rice seed production.

Floral traits of the seed parent which have been found to enhance out­crossing potential in rice include:

- .higher stigma exsertion rate (Table 3.6); - longer duration of floret opening (Namai, 1987); - longer period of stigma receptivity; - large and feathery stigma.

Natural Outcrossing Mechanisms in Rice 93

t-.1.---230mm ------i.!

9

Fig. 3.4. Durham's pollen sampler

A higher stigma exsertion rate was found to be significantly correlated with stigma length and pistil length (Table 3.7). The latter were negatively correlated with internal angle of the stigma lobes. Therefore, it is assumed that a wider internal angle of the stigma lobes reduces stigma and pistil length and also reduces the percent of spikelets with exserted stigmas and the outcrossing rate. The internal angle of the stigma lobes was 73.60 on average and varied from 50.80 to 108S in the 32 samples of 16 temperate Japonica rice cultivars (Namai 1987).

94 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Table 3.7. Correlation coefficients among floral traits for seed parent in the 32 samples of 16 temperate Japonica cultivars. (Namai and Kato 1987a)

Stigma Pistil Internal angle of Angle of floret length length- the stigma lobes opening at 1 h

after flowering

Spikelet 0.655** 0.679** - 0.381* 0.365* with exserted stigma, % Stigma length 0.953** - 0.523** 0.458** Pistil length - 0.519** 0.553** Internal angle of - 0.315

the stigma lobes

• Pistil length here = stigma + style.

Fig. 3.5. Relationship of relative humidity to seed set and seed yield on a cytoplasmic male-sterile li~e (lR62829A) of rice

Some morphological traits of rice plants also affect outcrossing potential in rice. For example, incomplete panicle exsertion hinders outcrossing, while a small and horizontal flagleaf enhances the rate of outcrossing on seed parents. The number of panicles per square meter and the number of spikelets per panicle are the main seed yield components. Therefore. high tillering capacity and larger number of panicles with many spikelets are the basic traits of the seed parent (Namai 1987).

The synchronization of flowering day and time in pollen and seed parents is the key factor to enhance outcrossing rate and seed set percentage in a hybrid rice seed production plot. Therefore, the pollen parent, viz., maintainer and

Natural Outcrossing Mechanisms in Rice

Seed yield (kglha) 1200

1000

800

600

400

200

95

0~~~ __ 1--L~ __ ~~ __ L--L~ __ L--L~~

Seed set ('l6)

~r J. No. of airborne pollen grains/liter

::[u muum mom o ! I I "

Wind velocity (m/s)

mmu uuuu __ mmuuJ Threshold level

, , , ! , , ,

5~---------------------------------------,

4

3

2

1 O~~~--L--L~--~~~~~~~~~~~

Relative humidity (%) 80

60

40

20

o AG28 527 N13 JA3 F5 Ml A4 514 016 028 JA24 F17 M19

I--- Wet season --I 1001.>------- Dry season -----.

Date of 50% flowering

Fig. 3.6. Number of airborne pollen grains ofIR62829B, wind velocity, relative humidity, seed setting (%), and seed yield of IR62829A as affected by time of flowering

96 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

restorer plants, should be genetically well synchronized in floret opening with the seed parent, independently of environmental factors (Namai 1987).

The environmental factors influencing natural outcrossing in rice include temperature, relative humidity, light intensity and wind velocity. Studies con­ducted at IRRI (Fig. 3.5) indicated that the seed set percentage and seed yield of a CMS line were negatively correlated to relative humidity. The highest seed yield was observed at IRRI when the seed and pollen parents flowered during the end of February to early March, when the relative humidity was 50-60% and wind velocity was above 2.5 m/s (Fig. 3.6).

Xu and Li (1988) specified favorable environmental conditions for hybrid rice seed production in China to include a daily temperature of 24-28°C, a relative humidity of 70-80%, a difference between day and night temperature of 8-lO oC and sunny days with a breeze. Generally, the spikelet opening percent­age is reduced when the temperature is high, humidity is low or when temper­ature is low and humidity is high. Such conditions also reduce pollen viability of the pollen parent and stigma receptivity ofthe seed parent. Flowering of the seed production plots at a location should be planned when unfavorable weather conditions are over.

3.3 Guidelines for Hybrid Rice Seed Production

Extensive research in China, IRRI and Japan have led to the identification of the following guidelines for successful hybrid rice seed production:

1. Selection of seed and pollen parents with synchronized time of anthesis. 2. Selection of seed parents with long, exserted stigma, longer duration and

wider angle of floret opening. 3. Selection of the pollen parent with larger anthers and with a high percentage

of residual pollen per anther after anther exsertion. 4. Synchronization of flowering time of the two parents by seeding them at

different dates depending on their growth duration or estimated accumulated temperature requirements for initiation of flowering. Extent of synchroniz­-ation of flowering of seed and pollen parents affected seed yield considerably (Fig. 3.7).

5. Use of optimum seed parent: pollen parent row ratio. Depending on the floral and morphological traits of the parental lines and environmental conditions at the site of seed production, the row ratio can vary from 6: 2 to 14: 2.

6. Use of seed and pollen parents with small and horizontal flagleaves, or cutting long and erect flagleaves.

7. Use of gibberellic acid (GA3) to improve panicle exsertion and prolong duration of floret opening.

8. Planting of seed parent: pollen parent rows across the prevailing wind direction and use of supplementary pollination with a rope or stick when wind velocity is below 2.5 m/s.

Guidelines for Hybrid Rice Seed Production

Yield (kgfha)

800

97

Synchronization of flowering

Fig. 3.7. Effect of synchronization (difference in days to flowering) of CMS line, IR62829A with pollen parents on hybrid seed yield, IRRI

Table 3.8. Effect of time of sowing on seed setting, Tamil Nadu Agricultural University, Coimbatore, India, 1990

Time of planting Seed set (%)

V20A V20B

1 February 1990 16.2 87.3 15 February 1990 12.4 87.8 1 March 1990 19.2 79.4 15 March 1990 22.4 82.5 1 April 1990 13.2 79.8

9. Selection of optimum time of flowering of parental lines in seed production plots. For example, seed yields at IRRI are higher during the dry season than during the wet season. Studies conducted at Tamil Nadu Agricultural University at Coimbatore also indicated seed set ranging from 12.4 to 22.4%, depending on time of planting of the CMS line V20A (Table 3.8).

Xu and Li (1988) listed the factors to obtain high seed yields in China. These include:

- Selection of prime field plots possessing fertile soil with good physical and chemical characteristics, adequate irrigation and drainage, sufficient sun­shine, even topography and regular plot shape; the incidence of disease and insect problems, especially those forbidden by quarantine regulations, should be minimal.

- Optimum weather conditions.

98 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

- Synchronized flowering attained by: • staggering seeding dates of pollen and seed parents; • sowing the pollen parent three times to extend the period during which

pollen is available, and • predicting and adusting flowering dates.

- Regulating growth to establish a population with large (more than 100 spikelets) and uniform panicles (more than 0.9 million productive panicles per hectare for the pollen parent and more than 3 million productive panicles per hectare for the seed parent).

- Planting of seed and pollen parents in such a way that they receive good aeration and equal amounts of sunlight and their row direction is perpendicu­lar to the prevailing wind direction at flowering.

- Clipping offlagleaves (1/2 to 1/3 from the top) 1-3 days before initial heading. - Spraying of GA3 two or three times (75 g GA3/ha) to increase panicle

exsertion. - Supplementary pollination by pulling a long nylon rope (5 mm diameter)

back and forth every 30 min during the blooming period. In hilly, uneven topography with small, irregular plots, a bamboo pole is also used for this purpose.

Namai (1987) listed the most important environmental and cultivating conditions for enhancing cross-pollination efficiency in hybrid rice seed produc­tion fields. These included:

1. Wind velocity 2-3 m/s. Suitable wind velocity increased the number of airborne pollen grains per liter (r = 0.868**) and the latter assured a higher outcrossing rate (r = 0.918**).

2. 1-1.5 m pollen flow distance from the pollen parent to seed parent. 3. High pollen shedding potential attained by getting 2000-3000 spikelets/m2

to bloom per hour during the peak flowering period.

3.3.1 Practices for Hybrid Rice Seed Production

Based on the above-mentioned guidelines, practices for hybrid rice seed produc­tion have been developed in China. These are being adapted and/or modified at IRRI and elsewhere to suit countries outside China.

Hybrid seed production involving cytoplasmic - genetic male-sterility and fertility restoration systems involves two steps, viz., multiplication of a CMS (A) line and production of hybrid (A/R) seed. However, production of hybrid seeds involving photo- or thermosensitive genetic male sterility involves one step (Fig. 3.8), i.e., production of hybrid seed; the ms line multiplication is done by selfing, which is achieved by growing the ms line under suitable day length and/or temperature conditions. In order to ensure purity of the hybrid seed and avoid pollination by unwanted rice varieties, the hybrid rice seed production field must be strictly isolated. An isolation distance of 100 m has been found to

Guidelines for Hybrid Rice Seed Production

Seed parent (Maintainer)

Three-line method

Pollinator (Restorer)

99

Seed parent Pollinator

r-­EGMS :

1- _____ "

Two-line method

Fig. 3.8. Comparison ofF 1 seed production systems. Three-line method: F 1 seed produc­tion system by cytoplasmic male sterility (eMS) and restorer gene (Rf). Two-line method: F 1 seed production system by environment-sensitive genetic male sterility (EGMS)

be satisfactory. Within this range no other rice varieties should be grown except for the pollen parent. Alternatively, a 3-week isolation is also recommended, which means that the flowering stage of rice varieties grown within 100 m around the seed production field should be 3 weeks earlier or later than that of the seed parent of the hybrid seed production plot. In some places topographical surface features and artificial obstacles may also be used as a means of isolation (Yuan 1985). The planting of two-three rows of Sesbania rostrata (which attains a height of about 2 m in about 6-8 weeks) around seed production plots has also been found to be a useful barrier to further strengthen isolation at IRRI.

As mentioned earlier, synchronization of flowering of the pollen and seed parent is the key to obtain high seed yields. Synchronized flowering can be obtained by using the criteria shown in Fig. 3.2.

Xu and Li (1988) also reported that the period from initial to full heading of a CMS line is 4-6 days longer than for a restorer line. The first sowing of the male parent establishes the dates for the second and third sowing. The second sowing is done when the leaf emergence on the first sowing is 1.1; the third sowing is done when the leaf emergence is 2.1. Plants from the second sowing provide the maximum pollen load, therefore the planting ratio of the seedlings of first, second and third sowing of the pollen parent is 1: 2: 1. The planting patterns commonly used in China are shown in Fig. 3.9.

Beginning about 30 days before heading, three or four random samples of the main culm of both parents are taken every 3 days. Young panicle develop­ment of the pollen and seed parents is compared under magnification. Based on the morphological features (Fig. 3.10), the young panicles are classified into eight developmental stages. The criteria for synchronization include (Yuan 1985):

1. The male parent should be one stage earlier than the female during the first three stages of young panicle differentiation.

100 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

R1 A A A A A A A A R2

R1 A A A A A A A A R2

R2 A A A A A A A A R3

R2 AAAAAAAA R3

R3 A A A A A A A A R2

R3 A A A A A A A A R2

R2 A A A A A A A A R1

R2 A A A A A A A A R1

Single row of male parent

R2R1AAAAAAAAR2R1

~~ A A A A A A A A ~~

R2R3 A A A A A A A A R2R3

R2R3AAAAAAAAR2R3

R2R1 A A A A A A A A R2R1

~~ A A A A A A A A ~~

R2R3 A A A A A A A A R2R3

R2R3 A A A A A A A A R2R3

Two rows of male parent

Fig. 3.9. Planting patterns of A and R lines used in hybrid rice seed production plots in China. (Xu and Li 1988)

2. Both parents should be in the same stages during the fourth, fifth and sixth stage of development.

3. The female parent should be slightly earlier than the male parent during the seventh and eighth stage of panicle development.

If during the first three stages of panicle development synchronization does not appear to be attainable, the earlier developing parent should be supplied with quick releasing nitrogen fertilizer and the later developing parent should be sprayed with a 1 % solution of potassium dihydrogen phosphate. This can adjust development differences of 4-5 days. During later stages of panicle differenti­ation, a difference of 3-4 days may be adjusted by drainage or irrigation, because the p~llen parents are more sensitive to water than the seed parent. For instance, if the pollen parent is found to be earlier, draining water from the field will delay panicle development. On the other hand, if the pollen parent is found to be late, higher standing water would facilitate rapid panicle development.

If the differ,ence in flowering period between the two parents reaches 10 days or more, it is necessary to remove the panicles from the early developing parent and apply nitrogen fertilizer subsequently, thus making its late emerging tillers or unproductive tillers bear panicles and subsequently achieve synchronization of flowering (Yuan 1985).

In order to regulate growth of the pollen and seed parents, their seedlings should be standardized (Xu and Li 1988). Seedlings with healthy tillers give larger panicles. For short duration hybrids, seedlings of the male parent should be 20-30 days old with 5.5-7 leaves and 2-3 tillers at the time of transplanting. For hybrids with a longer growth duration, seedlings of the male parent should be 30-35 days old with 7-8 leaves and 3-4 tillers. Seedlings of the female parent should be 20-30 days old with 5.5-7 leaves. Experience has shown that higher

Guidelines for Hybrid Rice Seed Production

Fig. 3.10. Morphological features of the growing panicle of different stages of development. (Yuan 1985)

1

Differentiation of the first bract primordium

~. - --......

A

A

c

2

Differentiation of primary branch primordium

3

B

Differentiation of secondary branch primordium

B

A 4 B

Differentiation of stamen and pistil primordia

101

102 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Table 3.9. Effects of row ratios on panicle number of female parent, seed setting percentage and yields. (Xu and Li 1988)

Row ratio Productive Spikelets Seed set Yield panicles panicle (%) (tfha) (x 103 )

1:15 2619 65.9 33.2 1.6 1: 14 2642 68.6 30.4 1.6 1: 13 2583 64.8 38.7 1.9 1:12 2458 67.9 37.1 1.8 1: 11 2562 74.5 39.9 2.3 1: 10 2354 72.4 33.7 1.8 1:9 2151 72.0 40.2 1.8 1: 8 2005 67.0 36.4 1.4

row ratios, narrower row spacing and more seedlings per hill will increase seedlings, panicles and grain yields (Tables 3.9, 3.10, 3.11). In general, the pollen parent is transplanted with a single seedling per hill and separated by a spacing of 13 cm from plant to plant and 165-200 em from one row of pollen parent to another row. About 45000 hills per hectare are needed. The seed parent is transplanted with two seedlings per hill with a spacing of 13 x 16 cm, and approximately 300 000 hills per hectare are needed.

After the transplanting of seed and pollen parents good field management is essential to obtain high seed yield. Adequate fertilizer is applied during the early stage of growth to produce more effective tillers. Lesser fertilization and irrigation during the middle and late stages restricts the growth offlagleaves and this provides good aeration and sunlight penetration to the canopy, which favors pollen spread (Yuan 1985). Appropriate measures should be taken to control weeds, diseases and insects in the seed production plot.

3.3.2 Flagleaf Clipping, Gibberellin Application and Supplementary Pollination

These are the specialized techniques for hybrid rice seed production and have been found very effective for increasing the outcrossing rate.

Flagleaves taller than the panicles are the main obstacles to cross-pollina­tion. Their cutting helps to remove this barrier to pollen spread and provides the seed parent an opportunity to receive more pollen from the pollen parent. Generally, flagleaf clipping is carried out 1-2 days before the initiation of heading. One-half to two-thirds of the flagleaf is cut to promote uniform pollen movement; it may also improve the microclimate to help synchronization of blooming of the two parents.

Gibberellic acid (GA3 ) spraying of 60 ppm at 5-10% heading, followed by another spraying of 30-60 ppm at 30% heading, helps to enhance panicle

Tab

le 3

.10.

Eff

ect

of p

lant

spa

cing

on

pan

icle

num

ber

of

fem

ale

pare

nt a

nd

gra

in y

ield

. (X

u an

d L

i 19

88)

0 =

a: P

lant

spa

cing

(cm

) S

eedl

ings

P

rodu

ctiv

e P

anic

le

Spi

kele

ts/

Fil

led

Seed

10

00-g

rain

Y

ield

!!

. (x

103

) pa

nicl

es

bear

ing

pani

cle

spik

elet

s/

sett

ing

rate

w

t. (t

/ha)

er (1

)

(x 1

03)

till

ers

(No.

) pa

nicl

e (%

) (g

) <

Il 0'

(%)

(No.

) .... ::c

I"

6.7

x 13

.3

2985

37

35

78.0

88

.7

43.3

48

.8

30.0

3.

4 '<

0

-

6.7

x 16

.7

2385

31

35

79.2

92

.6

43.2

46

.7

30.2

3.

4 :I

. Q

..

lOx

10

2655

38

10

82.0

82

.6

39.8

48

.2

30.1

3.

4 :;c

lIb

10

x 1

3.3

3984

34

20

76.0

76

.1

o·

35.8

47

.0

29.8

3.

4 (1

)

10 x

16.

7 31

80

3465

73

.0

78.0

41

.4

53.1

29

.3

3.9

til

(1)

(1)

13.3

x 1

3.3

2985

34

50

85.0

76

.8

39.5

51

.4

30.7

3.

8 Q

..

13.3

x 1

6.7

2385

31

05

72.2

74

.2

31.8

42

.8

30.5

3.

8 '"

tj .... 0

b 2

Pla

nts/

hill

. C

ombi

nati

on V

20A

jlR

26.

Row

rat

io =

1:

9.

Q..

a 1

Pla

nt/h

ill.

fi ::to

0 ::s

Tab

le 3

.11.

Eff

ect

of s

eedl

ings

pla

nted

on

pan

icle

num

ber

of

fem

ale

pare

nt a

nd g

rain

yie

ld

See

dlin

gs

See

dlin

gs

Pro

duct

ive

Pan

icle

S

pike

lets

/ F

ille

d Se

ed

1000

-gra

in

Yie

ld

per

hill

(x 1

03)

pani

cles

be

arin

g pa

nicl

e sp

ikel

ets/

se

ttin

g w

t. (t

/ha)

(x

103

) ti

ller

s (%

) (N

o.)

pani

cle

rate

(g

) (N

o.)

(%)

1 46

8 25

20

77.0

99

.9

20.1

20

.1

30

1.02

2

933

2985

78

.0

67.7

25

.6

37.8

30

2.

23

3 14

00

3570

82

.0

75.3

28

.8

38.3

31

2.

66

4 18

66

3360

85

.0

100.

2 24

.7

24.7

32

2.

67

2 93

3 31

35

89.9

87

.2

25.4

29

.1

32.3

2.

43

4 18

66

3540

83

.5

69.9

26

.3

37.6

31

.5

2.81

6

2799

42

90

82.0

75

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104 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Table 3.12. Effect of ftagleaf clipping and GA3 application on outcrossing rate. (Xu and Li 1988)

Treatment Productive Panicle Spikelets/ Seed panicles/ha bearing panicle setting (x 103) tillers (No.) rate

(%) (%)

Flagleaf clipping and 3187 82.5 68.0 27.2 GA3 application

Check 2625 71.4 65.2 17.2

exsertion in some CMS lines. It also increases the rate of stigma exsertion, increases the duration of opening of florets and causes the secondary and tertiary tillers to grow faster. The effect of leaf clipping and GA3 application on the outcrossing rate on a CMS line in China is given in Table 3.12. Recently, Chinese scientists have recommended higher dosages of GA3 (IOO-IS0 g/ha) to increase seed yield even without practicing flagleaf clipping (L. P. Yuan, pers. comm.).

Shaking the panicles of the pollen parent by rope pulling or rod driving during anthesis (called supplementary pollination) can make the anthers dehisce and spread the pollen widely and uniformly. It is generally carried out in the morning when blooming occurs in the seed and pollen parent. When only the pollen parent is blooming, supplementary pollination should not be done. In the afternoon, when the pollen parent is still blooming, supplementary pollination should be continued even if the seed parent has closed its glumes (Yuan 1985). Generally, supplementary pollination is carried out at 30-min intervals, five times daily, until no pollen remains in the pollen parent; however, it is not needed when the wind velocity is greater than a moderate breeze (Yuan 1985).

3.3.3 Roguing

The performance of a heterotic F 1 rice hybrid also depends on the purity of its seed. Hence production of pure seeds for meeting a specified standard is essential to the success of a hybrid seed production program. In China, certified seed of an F 1 rice hybrid must be 98% pure (Yuan 1985). In order to meet these requirements, it is necessary to thoroughly rogue the seed production plot in addition to observing strict isolation standards. Rogues to be removed are: the off-type plants mixed in with both seed and pollen parents, and maintainer and semisterile plants which appear in the seed parent rows. Off-type plants can be identified based on their morphological characteristics, viz., color of leaf sheath, leaf collar, size of leaf blades, growth stage, plant type, plant height and growth duration. Maintainer plants generally flower 3-S days earlier than the ms line, the basal part of their panicles normally exsert out of the flagleaf sheath and

Guidelines for Hybrid Rice Seed Production 105

their anthers are yellow, plump and completely dehiscent after flowering. Semisterile plants can be identified by their yellowish and partially dehiscent anthers which are slightly larger than those of the male-sterile line and which become dark yellow several hours after flowering (Yuan 1985).

Roguing is practiced from seedling to harvesting stages. The final inspection of the seed production field is done just before harvesting to remove any off-type plant.

3.3.4 Harvesting and Threshing

Harvesting of the hybrid rice seed production field differs from the harvesting of the inbred rice seed production field as it involves the seed and the pollen parent, which should be harvested and processed separately so as to maintain the high standard of physical and genetical purity. The timely harvest of the seed production plot provides higher yield and seed of good quality, as it is less prone to bad weather conditions, shattering, lodging, damage by rats, birds and other insect pests.

As harvest time approaches, the crop should be observed daily and panicles on the main tiller of the seed parent inspected. The crop is ready to be harvested when the filled grains on the upper portion of the panicles are clear and firm and some in the base are in the hard dough stage. At this time, 90% of the grains are straw colored. The moisture content of the grain is 20% or below when the crop is ready for harvest, otherwise the quality of the seed harvested is affected.

The pollen parent is harvested first and taken out of the field, ensuring that not a single panicle of the pollen parent is left there. The seed parent is harvested by sickles, scythes or by a mechanical device such as a combine. The mechanical device should have adjustable speed and clearance so that the quality of seed is not affected by crushing, brushing, or cracking.

Although the pollen parent is harvested first, threshing of the seed parent should be done first, either manually by a hand flail on a threshing floor or by manual or power-driven threshers. The pollen parent should be kept on a dried threshing floor suitably separating it from the seed parent and taking all precautions that the two parents do not get mixed by wind or any other means. Care should be taken that the threshing floor, the thresher, etc. are thoroughly cleaned before the threshing of the seed parent is undertaken. After cleaning the floor and the thresher, the R line is threshed.

3.3.5 Seed Processing

Seed received after threshing often has a high moisture content and contains trash, inert matter, deteriorated and damaged seeds, weed seeds, etc. Seed processing is done in order to dry the seeds to a safe moisture level, remove or reduce various undesirable materials to the greatest extent possible, and to carry

106 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

out uniform size grading so as to improve the quality of the seed received from the fIeld. Thus, seed processing actually prepares the threshed seed for marketing.

Drying of the seed can be done either in the sun or by forced air-drying (generally by heated air, at 40-50 DC, in a batch-type drier) to bring the moisture level to safer storage limits. Rice seeds are generally stored at 13 % moisture content. Seed can only be dried to the greatest extent when the moisture­retaining tendency of the seed and moisture-withdrawing tendency of the air do not come into equilibrium; when they do come into equilibrium, seeds no longer lose moisture to the air. Seed should be dried generally, so as to keep the seed viability intact. After the seed has been dried to 13% moisture content or lower, it should be cleaned properly and graded.

The basic purposes of cleaning and grading are: (1) to remove impurities (such as trash, leaf, weed seeds, inert matter, broken seeds, etc.); (2) to remove seeds of wild or other plant species; (3) to remove immature, shrivelled, unfilled and empty grains; (4) to remove seeds which do not conform to the average size, and (5) to obtain the maximum amount of clean seeds having intact viability.

Basic seed cleaning and grading is done by an air screen machine, commonly referred to as an air screen cleaner. The air screen machine uses three cleaning elements, viz., aspiration (to remove light seed and chaffy materials), scalping (to separate good seeds from oversize materials) and grading (to separate good seeds of uniform size from smaller particles and undersized, broken and shrivelled seeds).

3.3.6 Seed Storage

Seeds should be stored properly after threshing until the next sowing so that seed germination and vigor are maintained. Seed storage is more important where surplus quantities of seeds (carry-over seeds) are to be stored. Temper­ature and relative humidity during storage are the most important factors during the storage life of the seeds. Generally, dry, cool conditions are best for seed storage. As a general rule:

- A decrease of 1 % in the seed moisture content nearly doubles the storage life of seeds.

- A 5.5 °C decrease in temperature nearly doubles the storage life of seeds. - Good seed storage is achieved when the percentage of relative humidity in the

storage environment and the storage temperature in degrees Fahrenheit add up to 100.

- Temperatures below 20°C are considered unconducive for insect activity in the seed storage area.

3.3.7 Cost of Hybrid Rice Seed Production

Studies conducted in China (He et al. 1987b) indicated that total labor input to multiply a hectare of a CMS line or produce a hectare of a hybrid seed

Guidelines for Hybrid Rice Seed Production

Table 3.13. Means and coefficient of variability (CV) of labor inputs for male­sterile (A-line) seed multiplication and for F I hybrid seed production. (He et al. 1987b)

Item Man-days/ha

A line FI hybrid (n = 8) (n = 21)

Land preparation 31 18 (45) (28)

Seed selection and preparation 3 2 (33) (21)

Seed preparation and care 15 14 (25) (40)

Transplanting 67 59 (16) (19)

Organic fertilizer application 32 18 (85) (70)

Chemical fertilizer application 13 14 (44) (28)

Water control 28 25 (48) (23)

Disease and insect control 15 20 (34) (43)

Weed control 17 21 (54) (40)

Special activities 69 85 (40) (26)

Harvest and post-harvest 70 70 (27) (17)

Others 39 30 (48) (30)

Total labor days 397 375 (8) (7)

Figures in parenthesis are CV (%).

107

production plot used nearly 400 man-days in contrast to 300 man-days/ha to grow a crop of inbred rice in Jiangsu province of China (Table 3.13). Thus, it takes about 100 more man-days/ha to produce a crop of A-line than to grow a rice crop. The coefficient of variability (CV) of total labor, at 8% was small, although the CV of individual operations varied somewhat more. The largest labor inputs were for transplanting (67 days/ha), special activities (69 man­days/ha) and h~rvesting and threshing (70 days/ha). The high labor input for transplanting is a result of the fact that the pollen parent is often transplanted two-three times to ensure that the flowering of pollen and seed parents is synchronized. In the hybrid seed production plot, pollen and seed parents are transplanted at two different times, depending on their growth duration. The special activities include flagleaf clipping, GA3 application and supplementary pollination.

Nonlabor inputs used for the multiplication of CMS lines and production of hybrid seed in Jiangsu province, China, are given in Table 3.14.

108 Outcrossing Mechanisms and Hybrid Seed Production Practices in Rice

Table 3.14. Means and CV of non-labor and recurrent inputs for male-sterile A-line seed multiplication and for F 1 hybrid seed production. (He et al. 1987b )

Item A-line product F 1 hybrid seed

Users Mean value CV Users Mean value CV (%) (n = 8) (%) (%) (n = 21) (%)

(/ha) (/ha)

Purchased inputs Pesticides 100 15 kg 58 100 46 kg 82 Organic fertilizer 88 45 t 128 100 33 t 79 Inorganic fertilizer: N 100 99 kg 52 100 150 kg 37

P 75 39 kg 25 81 53 kg 40 K 38 3kg 30 29 47 kg 87

Total N 100 268 kg 103 100 288 kg 46 Seed female parent 100 30 kg 26 100 35 kg 13

(A-line) Seed male,parent (BIR) 100 11 kg 40 100 9kg 26

Machinery inputs Power tiller 100 32 $" 47 100 24$ 41 Machine thresher 63 9$ 121 86 13$ 20 Irrigation cost 100 23 $ 35 100 2lkg 39

a Exchange rate in 1984 $1.00 = Yuan 2.70.

Total labor and nonlabor costs per hectare of seed production were found to be US$ 725 per hectare for CMS line multiplication and hybrid seed production excluding the cost of GA3 , which is about US$ 90 per hectare (150 g or $ 0.60 per g). Thus, a hectare of hybrid rice seed production costs about US$ 815. This does not include the cost of land use or the cost of money (interest) invested in the seed production of hybrids (He et al. 1987b).

The study of He et al. (1987b) also showed that seed growers were getting seed yields of 1.4 t/ha from the seed parent and 1.2 t/ha from the pollen parent (maintainer line) in the CMS line multiplication plots, and 1.7 t/ha from the seed parent and 1.8 t;ha from the pollen parent (restorer line) in the hybrid seed preduction plot. These yields fetched gross returns of US$ 1940 from CMS line multiplication and US$ 2717 from the hybrid seed production plot.

A line/hybrid seed was procured by the national seed company @ US$ 1.3 per kg. Rice growers in China also price rice straw fetching about US$ 50-70/ha; however, this has not been considered for the estimation of gross returns because outside China straw is not priced. The net returns for seed production are estimated to be US$ 1135/ha for CMS line multiplication and US$ 1602/ha for hybrid seed production.

Outside China, the cost of hybrid seed production has been estimated in the Philippines (IRRI, unpubl.). The preliminary assessment indicates that labor inputs are about 270 man-days/ha costing US$ 505/ha and nonlabor inputs are US$615/ha, thus total cost of seed production is US$ 1120/ha.

Guidelines for Hybrid Rice Seed Production lO9

While the labor inputs are less, their cost is higher than in China. Nonlabor inputs are also higher than in China, primarily because of the high price of GA3 (US$ 8 per g compared with US$ 0.60 per g in China). The total cost of GA3 is estimated to be US$ 400 for 50 g in comparison to US$90 for 150 g in China.

Seed yield in the Philippines has been found to range from 0.7 to 2.2 t/ha (mean 1.3 t/ha) for the seed parent and 1.1 to 2.9 tjha (mean 2.1 tjha) for the pollen parent. At a rate of US$ 200 per ton of paddy rice, the produce from the pollen parent is worth US$ 420/ha. Thus, the cost of production of 1.3 t/ha of hybrid seed is estimated to be US$ 700/ha or US$ 0.54 per kg, which is comparable with the cost of hybrid seed production in China. If the ex-farm price of hybrid seed is US$ 1 per kg before processing and cleaning, the seed grower will have a net profit of about US$ 6oo/ha.

To the ex-farm price of US$ 1 per kg of hybrid seed, the cost of seed processing, cleaning, grading, packing, storage, and marketing are to be added; this may be provided by a public or private sector seed company. Post-harvest processing ana handling costs plus profits may bring the sale price of hybrid rice seed for the hybrid rice cultivator to about US$ 1.75 to 2.00 per kg.