This document provides information about self-incompatibility in cole crops. It begins with background information on the student, K. Modunshim Maring, and their major and minor guides. It then discusses the definition and general features of self-incompatibility, different classifications of self-incompatibility systems, and mechanisms and molecular basis of self-incompatibility in cole crops specifically. It also covers the history, importance, assessment, and exploitation of self-incompatibility for hybrid breeding in cole crops.

2. K. Modunshim Maring
Reg. No.:- 2010118055
Dept. Of Genetics & Plant Breeding
N. M.College Of Agriculture, N.A.U.
Navsari-396 450
GP.591 (1+0)
Major Guide
Dr. Madhu Bala
Assistant Professor,
Dept.of Genetics and Plant Breeding
N. M. College of Agriculture,
N.A.U., Navsari - 396 450.
Minor Guide
Dr. V. B. Parekh
Assistant Professor (Plant Biotechnology),
Department of Basic science and Humanities,
ASPEE College of Horticulture and Forestry,
N.A.U., Navsari- 396 450.
2

3.  It refers to the inability of a plant with functional pollen to set seed upon
self pollination. OR
It is defined as the prevention of fusion of fertile (=functional) male and
female gametes after self-fertilization. It was first described about the
middle of eighteenth century by Koelreuter.
 The process of pollen germination, pollen
tube growth, ovule fertilization or embryo
development is halted at one of its stages
and consequently no seeds are produced.
Self-incompatibility
3

4. General features of self-incompatibility
 Prevent self-fertilization and thus encourage out-crossing or allogamy.
 Increases the probability of new gene combinations.
 Normal seed set on cross pollination.
 May operate at any stage between
pollination and fertilization.
 Reduces homozygosity (aa/AA) and
increases heterozygosity (Aa).
 In plants, self-incompatibility is often inherited by a single gene (S) with
different alleles (e.g.S1,S2,S3 etc.).
AA x aa
Aa
4

5. Classification of self-incompatibility
Distyly
Heteromorphic SI Homomorphic SI
Self-incompatibility
Tristyly Gametophytic SI Sporophytic SI
Lewis, 19945

6. Table.1 Heteromorphic system of self-incompatibility
Phenotype Genotype Genotype Phenotype
Pin x Pin ss x ss Incompatible mating
Pin xThrum ss x Ss 1 Ss : 1 ss 1 Thrum : 1 Pin
Thrum x Pin Ss x ss 1 Ss : 1 ss 1 Thrum : 1 Pin
Thrum x Thrum Ss xSs Incompatible mating
Heteromorphic self-incompatibility system
 Flowers of different incompatibility group are
different in morphology.
Distyly
• Pin type (ss) – long styles, short stamens
• Thrum type(Ss) – short styles, long stamens
6

7. Short Style Medium Style Long Style
(S) (S and M) (s and m)
Tristyly
• In tristyly, styles and stamens have three different positions.
• It is determined by two genes S and M, each with two alleles.
7

8. Gametophytic SI Sporophytic SI
The stigma is smooth and wet The stigma is papilate ( hairy) and
dry
Pollen tube inhibition in style Pollen tube inhibition take place on
the stigmatic surface itself
The pollen-pistil interaction governed
by haploid genome of male gametes and
diploid genome of pistil tissue
The pollen-pistil interaction governed
by genome of the plant on which the
male and female gamete are produced
S alleles show co-dominance S alleles show dominance
Homomorphic self-incompatibility system
S1S3
8

9. It involves the following interactions
Pollen-stigma interactions
Pollen tube-style interactions
Pollen tube-ovule interactions
Mechanism of self-incompatibility
Singh, B.D., 20169

10. Pollen-stigma Interactions
• These interactions occur just after pollen reaches stigma and
generally prevent pollen germination. It is a common feature of
the sporophytic system where stigma is covered with a
hydrated layer of proteins known as pellicle. Pellicle appears to
be involved in SI reaciton.
• Within few minutes of reaching the stigma, pollen releases an
exine exudate. This exudate induces immediate callose
deposition in the stigma cells that are in direct contact of the
pollen; this occurs only if the pollen and the stigma cells have
the same SI reaction (incompatible mating).
10

11. Pollen Tube-style Interaction
• In most cases of gametophytic system, pollen grains
germinate and pollen tubes enters the stigma. But in
incompatible combinations, growth of the pollen tube is
retarded within the stigma or in the style. In the latter
case, there is cessation of protein and polysaccharide
syntheses in the pollen tubes, which ultimately leads to
the bursting of pollen tubes.
11

12. Pollen tube-ovule interaction
• In some cases, pollen tubes reach ovules and fertilization
takes place. But in incompatible combinations, embryos
degenerate at an early stage of development.
• Generally, species showing this
type of interaction have
hollow styles.
12

13. Elimination of self-incompatibility
• Doubling the chromosome number is effective in case
of single locus gametophytic system.
• Isolation of self-fertile(Sf) mutants. For this, flower
buds are generally irradiated at the PMC stage, and
pollen from these buds is used to pollinate flowers
having known S alleles. It is more successful in the
gametophytic than in the sporophytic system.
• Transfer of self-compatibility alleles from related
species.
Singh, B.D., 201613

14. Temporary suppression of self-incompatibility
During production of inbreds, it is sometimes desirable to
temporarily suppress the SI reactions of pistils to enable the
desired crosses to be affected. A temporary suppression of the
SI reaction is often referred to as pseudofertility, it may
achieved by following approaches.
 Bud pollination
 Surgical techniques
 Irradation
 High temperature
 High CO2 concentration
Sodium chloride spray
End- of-season pollination
Double pollination
In vitro pollination
Singh, B.D., 201614

15. Order-Brassicales (Cruciales)
Family- Brassicaceae (Cruciferae)
Tribe-Brassiceae
Genus -Brassica
Species -oleracea
Taxonomy of cole crops
Singh et al., 201615

16. Cole crops
Cole crops (Brassica oleracea) are a group of highly differentiated
plants having 18 numbers of diploid and somatic chromosomes (2n=2x=18),
and these are grown all over the world from arctic to tropical climatic
conditions. The word ‘cole’ seems to have been derived from the abbreviation
of the word ‘caulis’ meaning stem/cabbage/stalk.
It is known with different names as Kale (English), Kohl (German), Chou
(French), Cal (Irish), Col (Spanish), Cavolo (Italian) and Couve (Portuguese)
but generally, the word cole is more recognized in the literature worldwide.
The cole crops like, broccoli, brussels sprout, cauliflower, cabbage, kale and
kohlrabi at present have evolved after a long time of mutation, natural/artificial
hybridization, selection and domestication due the presence of variation within
and between subspecies of B. oleracea.
Singh et al., 201616

17. Important Cole Crops
Sprouting broccoli Cabbage
Kale
Knol KohlCauliflower
Brussels sprouts
17

18. Crops Scientific name
Chromosome
no.(2n)
Type of
pollination
Plant part used
Cabbage
Brassica oleraceae L.
var. capitata
18
Cross pollination
(73%)
Head
Cauliflower
Brassica oleraceae L.
var. botrytis
18
Cross pollination
(70%) Curd
Knol khol
Brassica oleraceae L.
var. gongylodes
18
Cross pollination
(91%) Swollen stem
Brussels
Brassica oleraceae L.
var. gemnifera
18
Cross pollination
(72%) Sprouts
Sprouting
broccoli
Brassica oleraceae L.
var. italica
18
Cross pollination
(95%) Flower heads
Kale
Brassica oleraceae L.
var. acephala
18
Cross pollination
(83%) Top leaves
18

19.  Low in carbohydrates, fats, calories.
 Good source of protein (balanced), minerals, vitamin A,
vitamin C and vitamin D.
 Known for anticancer properties (Indole-3-carbinol).
 Cabbage juice: against poisonous mushroom.
 Flavor compound : sinigrin.
 Includes antioxidants – ascorbic acid, tocopherols,
carotenoids, isothiocyanates, indoles, flavanoids.
Importance
19

20. Inflorescence
 Florescence of cole crops is racemose except cauliflower having
cymose type.
 Numerous small yellow/white flowers with 4 sepals, 4 petals, 6
stamens, of which 2 are short and other 4 stamens are longer than the
style and single superior ovary.
 The stigma receptive even 5 days before and 4 days after anthesis.
 Flowers are protogynous in nature.
 Major pollinators are honey bees and blowflies.
Singh et al., 2016
20

21. Self-incompatibility in cole crops
Plants have evolved many systems to prevent undesirable
fertilization. Among these, incompatibility is a well-organized system in
which pollen germination or pollen tube growth is inhibited in pistils. To
enable F1 hybrid breeding, an efficient, reliable and stable method of F1
seed production without contamination by self-fertilized seeds from each
parent is vital. Because of the size and structure of Brassicaceae flowers, it
is impossible to implement commercial hybrid seed production based on
manual emasculation and pollination.
The self-incompatibility system, was developed mainly by
Japanese breeders and has been effective in F1 breeding of vegetable
Brassicaceae.
Kitashiba & Nasrallah, 2014.
21

22.  First discussion on self-incompatibility by Darwin (1877)
 The term self incompatibilitywas given by Stout (1917)
 Bateman (1952, 1954, 1955) gave explanation on
incompatibility in three Brassica plants namely, Iberis amara
L., Raphanus sativus L. And Brassica campestris L.
 This system was confirmed :-
(In Cole Crops)
 Kale ( Thompson 1957)
 Broccoli ( Sampson 1957 and Odland 1962)
 Cabbage ( Adamson 1965)
 Cauliflower ( Hoser-krauze 1979)
History of self-incompatibility in cole crops
22

23. Why self incompatibility for development
of hybrids?
Reduced the cost of hybrid seed production
Production of large scale of F1 seeds
Speedup the hybridization programme
Commercial exploitation of hybrid vigour
It avoids enormous manual work of emasculation and
pollination
23

24.  Bateman (1955) described the control of SI in family Brassicaceae by
a single Mendelian locus, the S (Sterility) locus, which exists as
multiple alleles.
 The number of S-locus alleles is usually large:-
 22 in Iberis (Bateman,1955)
 34 in Raphanus (Sampson,1957)
 30 in B. rapa (Nou et al.,1993)
 50 in Brassica oleracea (Brace et al.,1994)
Genetics of self incompatibility
In Brassica sps , the exploitation of heterosis is mainly
through the sporophytic self incompatibility.
24

25. Molecular mechanism is SSI in cole crops
• SI recognition is controlled by a multiallelic gene complex at a
single locus, termed the S-locus.
• The S-locus consists of three genes
1. SRK (S-locus receptor kinase); (Stein et al.,1991).
2. SP11(S-locus protein 11)/SCR (S-locus cysteine rich);
(Suzuki et al.,1999; Takayama et al., 2000).
3. SLG (S-locus glycoproteins).
25

26. • SLG and SRK exhibit a number of characteristics that would be
expected for the female determinant of SI.
• They are predominantly produced in the stigma papilla cells, which
come into direct contact with pollen.
• Their expression occurs just prior to flower opening.
• SP11/ SCR is the male determinant and is predominantly expressed in
the anther tapetum.
• Immunohistochemical studies suggest that the SP11/SC protein is
secreted in a cluster from the tapetal cells into the anther locule and
translocated to the pollen surface.
Female Determinant
Male Determinant
26

27. (Takayama and Isogai 2005)27

28.  Upon pollination, SP11 penetrates the papilla cell wall and
binds SRK in an S-haplotype-specific manner.
 This binding induces the autophosphorylation of SRK,
triggering a signaling cascade that results in the rejection of
self-pollen.
 MLPK(M-locus protein kinase) localizes papilla cell membrane
and may form a signaling complex with SRK.
 ARC1(Armadillo repeat-containing 1) identified through
protein interaction with SRK.
 The proteasomal degradation of these substrates could result in
pollen rejection.
28

29. Assessment of self incompatibility
The plants are categorized as self-incompatible/compatible based on
average number of seeds in natural pollination per pod/ average number
of seeds in self pollination by bagging per pod. (Watts, 1963). It is also
known as Fertility index.
Values-
 Value (>2) Self Incompatible line
 Value (<1) Self Compatible line
 Value (>1 but <2) Pseudo incompatible line
Disadvantages :
One has to wait for 60 days till seed reaches to maturity after
pollination.
The no. of seeds reaching to maturity may also be reduced by any
biotic or abiotic stresses.
Chittora and Singh, 2016
29

30.  Fluorescence microscopy:-
The fluorescent microscopic observations on pollen ability to
penetrate style, has provided a more reliable method to directly
measure incompatibility/ compatibility reaction within 12-15
hours after pollination.
Penetration of style by none or few tubes indicates
Incompatibility
Penetration by many tubes indicate compatibility
Penetration by intermediate number indicates intermediate
level of Incompatibility
Chittora and Singh, 2016
30

31. Characteristics of superior SI lines
Stable self-incompatibility
High seed-set of self pollination at bud stage
Favorable and uniform economic characters
Desirable combinating ability
Continuous inbreeding in many cole crops may lead to complete loss
of inbred lines due to high rate of inbreeding depression.
Pseudo incompatibility may lead to self seed instead of hybrid seed
and deteriorate the genetic purity of the hybrids.
The degree of SI varies in different cole crops.
SI reaction is dependent upon specific temperature and photoperiod.
Hybrid seed will be expensive when SI lines are difficult to maintain.
Problems in exploiting SI lines
31

32. Basic steps in the use of SSI
Identification of self-incompatible plants in diverse
population/genotypes
Development of homozygous self-incompatible lines
Maintenance of parental self-incompatible
lines
Commercial hybrid seed production
Detection and establishment of inter-allelic relationships
among the S-alleles
Identification of S-alleles in the homozygous self-
incompatible lines
32

33. A plant is selected on the basis of its superiority in phenotype of
characters and selfed by bud pollination (A0) and the selfed seeds are
sown in rows (A1) and at the time of flowering a few buds are self
pollinated. In the next year, single plant progenies are grown (A2) and
about 10-12 plants in each progeny are maintained. These homozygous
families can be identified by pollinating each plant of a family with a
mixture of pollen taken from each plant of the family. If there is not or
very poor seed set, the line is homozygous for self incompatibility.
Development of homozygous SI line
Tripathi et al., 2008
33

34. Use of SI line for hybrid seed production
LINE A X LINE B
S11 S22
Propagation through bud-pollination
S11 X S22
COMMERCIAL HYBRID
A X B
S1234

35. A self-incompatible line may be inter planted
with a self compatible line
o Used in all the Brassica vegetables crops in Europe anda Japan
Seed from only the self-incompatible line would be hybrid seed,
while that from self-compatible line will be mixture of hybrid and
selfed seed.
Self-incompatible
line
Self-compatible
line
X
35

36. Maintenance of homozygous SI inbreds
• Bud pollination / Sibmating
• Treatment with CO2 gas (CO2 enrichment) (Jirik,1985) or
sodium chloride ( Kucera,1990)
 Other methods:-
• Electronic aided pollination (EAP); (Roggen et al., 1972)
• Steel brush method ( Roggen and Dijik,1972 )
• The pollen washing ( Roggen,1974)
• Thermally aided pollination (TAP); (Roggen and Dijik, 976)
36

37. Crossing methods involved in self-incompatibility
Line A x Line B
S1S1 S2S2
Line C x Line D
S3S3 S4S4
S1S2
(A x B)
Single-cross hybrid
S2S4
(C x D)
Single-cross hybrid
x Any Line
(A x B) x (C x D)
Double-cross hybrid
Three-way hybrid
x
Line A x OPV
F1
Top Cross
37

38. 38

39. Evaluation of self incompatibility in Indian cauliflower
(Brassica oleracea L. var.botrytis)
 Objective
To investigate in 14 genotypes for potential hybrid and synthetic development
in cauliflower of Indian origin.
Varanasi, India Singh et al.,2002
 Materials and Methods
The seedlings were transplanted at the distance of 45 x 13 cm2. The
inflorescence of bud stage was partitioned into four parts of each plants of
genotype. The first part was used for selfing by bud-pollination, second part
was used for sib-mating, third for natural selfing by covering with butter bag
and fourth part was allow for open pollination. At pollination time, 20 buds
were taken and at harvesting time all the matured slilqua were collected from
each plant , then siliqua setting percent and seed set ratio was calculated.
Case study 1
39

40. Genotypes
Number of siliqua set per
plant
Siliqua setting upon different
pollination systems(%)
OP SM BP NSP OP SM BP NSP
IIVR-1 14.0 13.0 10.0 0 70.00 65.00 50.00 00
IIVR-50 16.0 12.0 8.00 0 80.00 60.00 40.00 00
Kataki Early-29 19.0 15.0 5.00 0 95.00 75.00 25.00 00
Aghani JBT 23/60 15.0 9.0 3.00 0 75.00 95.00 15.00 00
Hazipur-4 17.0 15.0 7.00 0 85.00 75.00 35.00 00
Late Aghani 12.0 10.0 4.00 0 60.00 50.00 20.00 00
Pusa Hazipur 18.0 19.0 11.0 0 90.00 95.00 55.00 00
Pusi-4 15.0 15.0 12.0 0 75.00 75.00 60.00 00
Agahani 8 15.0 16.0 5.00 0 75.00 80.00 25.00 00
Agahani-31 17.0 14.0 8.00 4.00 85.00 70.00 40.00 20.0
Agahani Long Leaf 15.0 13.0 10.0 0 75.00 65.00 50.00 00
Agahani Small Leaf 14.0 12.0 7.00 0 70.00 60.00 35.00 00
Kuwari-1 16.0 10.0 10.0 0 80.00 50.00 50.00 00
Kataki-12 14.0 14.0 11.0 0 100.00 70.00 55.00 00
CD at 5% 0.67 0.70 1.33 0 11.86 3.58 4.07 -
Table 1: Number of siliqua upon Open Pollination(OP), Sib-Mating(SM), Bud Pollination(BP),
and from Natural Self Pollination(NSP).
40

41. Genotypes
Self pollination
OP
Seed set in different pollination
systems as compared to open
pollination(%)
SM BP NSP SM BP NSP
IIVR-1 2.61 1.75 0 6.07 43.01 28.83 0
IIVR-50 1.57 0.65 0 5.02 31.28 12.95 0
Kataki Early-29 2.43 3.38 0 6.72 36.16 50.30 0
Aghani JBT 23/60 1.45 1.20 0 6.84 21.20 17.54 0
Hazipur-4 3.99 2.43 0 8.36 47.73 29.07 0
Late Aghani 8.82 3.40 0 11.31 77.98 30.06 0
Pusa Hazipur 6.69 4.50 0 10.97 60.99 41.02 0
Pusi-4 3.92 2.10 0 8.95 43.80 23.46 0
Agahani 8 2.81 1.67 0 5.00 56.20 34.40 0
Agahani-31 3.82 1.66 1.04 3.88 98.45 42.78 26.8
Agahani Long Leaf 3.68 1.92 0 5.98 61.54 32.11 0
Agahani Small Leaf 3.99 1.94 0 13.64 29.25 12.02 0
Kuwari-1 2.77 2.02 0 5.81 47.68 34.77 0
Kataki-12 8.00 2.90 0 9.84 81.31 29.47 0
Table 2: Seed set per siliqua upon Sib-Mating(SM), Bud Pollination(BP), Natural Self
Pollination(SP) and Open Pollination(OP).
41

42. Genetic analysis of cabbage (Brassica oleracea
var.capitata) using self incompatible lines
 Materials and Methods
The material comprised of 4 stable self-incompatible lines, viz., 83-1, 83-
2, 83-5 and Sel-8 maintained by bud pollination and standard variety,
‘Golden Ace’ of cabbage. All this 5 lines were crossed in full diallel
mating design. The resulting 20 F1 hybrids along with their 5 parents were
evaluated in randomized block design with 2 replications. All 25 entries
were planted in 3 rows each in a plot of size 3m x 3m and a spacing of 45
cm x 45 cm. Data were recorded on 10 plants selected random in each
replications for 7 characters.
Parkash et al., 2003Himachal Pradesh, India
Case study 2
42

43. Parents or crosses
Days to
first
harvest
Frame
size
Stalk
size
index
Head
size
index
Unwr-
apped
leaves
Gross
weight/
plant
Net
weight/
plant
GCA status
83-1 11.45* 1.05 0.92* -1.24 1.48* 0.16* 0.02 Moderate(M)
83-2 -0.35 1.72 0.49* 13.54* 0 0.17* 0.08* High (H)
83-5 9.60* -1.22* 0.87* 15.92* 1.42* -0.08 -0.04 Low (L)
Sel-8 -9.60* 0.45 -1.83* 4.05 -1.36* 0 0.04 Moderate(M)
Golden Ace -11.10* -2.01 -0.45* -0.44 -1.52* -0.25* -0.10* Low (L)
SE 2.26 0.85 0.22 5.65 0.29 0.06 0.03
Best crosses
83-1 x 83-2 -1.20 3.49* -0.80* 45.69* -0.46 0.34* 0.20* M x H
83-1 x 83-5 -4.40 -3.42* 2.87* -38.54* -0.68 -0.38* -0.25* M x L
83-1 x Sel-8 -7.95* 3.31* 0.23 12.58 0.10 0.12 0.07 M x M
83-1 x Golden Ace -11.45* 1.52 0.80* 11.44 -0.34 -0.03 0.01 M x L
83-5 x Sel-8 -8.60* 4.13* -0.99* 3.14 -0.89 0.21 0.03 L x M
83-5 x Golden Acre -9.60* 4.14* 0.71 17.46* 1.02 0.30* 0.15* L x L
Sel-8 x Golden Ace 9.60* 1.32 0.06 32.59* 0.20 0.23* 0.23* M x L
SE 3.68 1.57 0.36 7.60 0.61 0.11 0.07
Table : Estimates of gca effects of parents & sca effects of hybrids in cabbage
Parents
43

44.  Results and conclusions
Negative sca effects for days to harvest shown by majority of the
crosses indicated. The cross showing significant positive sca effects for
net weight per plant involved the parents with low x low, moderate x
high or moderate x low gca effects, which indicated the presence of of
non-additive effects. Therefore the heterosis breeding would be
desirable for improvement towards higher yield and earliness in
cabbage. It is evident from the study for stable self incompatible lines
showing good general combining ability and may economize the
production of hybrid seed.
44

45. Environmental factors and genotypic variation of
self-incompatibility in Brassica oleracea L. var. capitata
 Materials and methods
Lines of Brassica oleracea L. var. capitata (P 1116, P 703, P 601,
7023b, 7024b/8, 15425, 15428/14, 378/4, 378/8, 1344, 705, 263, K-48, 26/2)
were examined in an experimental sequence lasting three years. The material was
over-wintered in a frostproof glasshouse, and in mid April, it is set into plastic
pots and placed in a glasshouse at ~20°C. About a week before flowering, the
plants were distributed in different temperature and humidity. After a week of
pollination, the plants were transferred to open-air conditions where they
remained to harvest time. Samples of the styles were gathered 48 h after
pollination (at least 10 flowers from 3 plants) and examined for pollen tube
growth using fluorescence microscopy or left on the plants to set seed, and for the
seeds to ripen. Data are presented as percentages of styles containing pollen tubes
and percentages of siliques containing seeds (fruit set).
Cracow, Poland Zur et al. 2003
Case study 3
45

46.  Temperature effect
Fig. 2. Effect of temperature on self-incompatibility among 14 Polish cabbage lines.
(a) Open flower pollination, (b) Green bud pollination.
Analyses of green bud pollination data revealed that higher temperature halved
the percentage of fertile siliques as compared with lower temperature. Such
effect was noted for almost all examined lines (Fig. 2b).
46

47. The humidity effects on the frequency of seed production were less than the temperature
effects. The results indicate that the best condition for open flower pollination for most lines
was 12°C and 60% relative humidity.
Temperature
Line
12oC 23oC
Relative humidity Relative humidity
60% 90% 60% 90%
378/4 0 13 6 0
P1116 20 5 0 5
P703 14 15 0 1
705 9 2 1 1
263 82 48 3 66
P601 20 45 0 1
7023B 76 54 0 0
7024B 69 17 0 0
15425 50 21 80 53
Mean 38 27 10 14
Table 1: Fruit set ability after open flower pollination of 9 lines of Brassica oleracea L.var.
capitata under various temperature/humidity conditions.
47

48. Similarly, after green bud pollination at the more favorable lower temperature
(12°C), no clear effect of humidity was noted.
Temperature
Line
12oC 23oC
Relative humidity Relative humidity
60% 90% 60% 90%
378/4 48 76 22 23
P1116 22 36 28 25
P703 73 100 9 11
705 90 39 49 62
263 99 84 30 41
P601 100 98 41 61
7023B 98 77 2 1
7024B 81 65 17 52
15425 91 78 64 82
Mean 87 81 29 40
Table 2: Fruit set ability after green bud pollination of 9 lines of Brassica oleracea var.capitata
under various temperature/humidity conditions.
48

49. Hybrid breeding of cauliflower using self-incompatibility
and cytoplasmic male sterility
Objective
The aim of this work was to verify the possibility of utilizing created self-
incompatible and male sterile lines for practical cauliflower hybrid breeding.
Material and methods
 Plant material
Self-incompatible S homozygous sublines were derived from the
cauliflower hybrid cv. Montano (MT) by means of inbreeding and subsequent
selection of SI plants.
The male sterile analogues of two cauliflower cultivars Brilant and Fortuna
were produced using repeated backcrossing with the original CMS material
that was characterized by an inferior agronomic quality. The open pollinating
stocks of cv. Brilant (BR) and Fortuna (FT) were used as maintainers of CMS
lines.
Olomouc, Czech Republic Kucera et al., 2006
Case study 4
49

50.  Reproduction of SI lines
In the first trial six SI MT sublines were planted into isolation cages of 5 × 3 m size
(plant spacing 50 × 50 cm2) and grown until flowering. Honeybees were used as
pollinators and the lines were treated during the flowering period with 3% NaCl solution
spray every morning before bees flies out.
The second trial was arranged in the same way but the NaCl
treatment was applied every other evening and bumblebees
were used as pollinators. After harvesting, the mean weight of
seeds per mature plant was counted for every subline.
 Production of CMS lines
The trials were situated into two isolation cages and were arranged under the same
conditions as mentioned above. CMS lines of Brilant and Fortuna were grown with
their fertile analogues in alternate rows (2 rows of CMS and 1 row of fertile lines).
Honeybees were used as pollinators. After harvesting, the mean weight of seeds per
one CMS and fertile plant was counted.
50

51.  Production of hybrid seed based on SI and CMS lines
The possibility of hybrid seed production in isolation cages with insect
pollinators were verified using SI (MT) and CMS (BR) lines as mother parents
and the fertile stock from cv. Fortuna as a donor of pollen. SI and CMS lines were
grown in separate isolation cages in alternate rows at the rate of two rows of SI or
CMS line and one row of the fertile parent FT 13.
 Preliminary evaluation of F1 hybrids
F1 hybrids based on SI and CMS were evaluated in the preliminary field trial
with 30 plants per plot in two replicates. Seeds were sown during the last week of
May. The seedlings were planted in the field at the end of June in plant spacing 50
× 50 cm2. Treatment and harvesting of the trial were arranged in accordance with
the state variety trials.
51

52.  Results and discussion
The results in second trial (Nacl treatment every other evening, bumblebees as
pollinators), were much better than the first trial. Mean weight of seeds in all SI
MT sublines reached from 4.0 to 6.5 g per harvested plant (Table 1). The mean
weight of seeds per plant from all sublines was 5.1g.
SI subline Number of
grown plants
Number of
harvested seeds(g)
Weight of
harvested(g)
Mean weight seeds
per plant(g)
3/1 6 6 39 6.5
4/1 6 5 30 6.0
4/2 6 3 15 5.0
5/1 6 4 18 4.5
6/1 6 4 16 4.0
8/1 6 4 19 4.8
Table 1: Reproduction of SI MT lines by means of NaCl spraying
52

53. The low seed set in Brilant CMS line might have been caused by the fact that
fertile parent plants finished flowering earlier than the sterile analogue. The CMS
plants were also weaker in growth and some of them were partly damaged by
fungal diseases. Flowering of both parental components from Fortuna were better
time-balanced. The results confirm the possibility to reproduce CMS cauliflower
lines in isolation cages by pollination with their fertile analogues using insect
pollinators.
CMS/fertile
line
Number of
grown plants
Number of
harvested plants
Weight of
harvested seeds(g)
Mean weight of
seeds per plant
(g)
BR CMS 24 20 15 0.8
BR 19 16 16 90 5.6
FT CMS 24 22 43 2.0
FT 13 16 16 157 9.8
Table 2 : Reproduction of CMS lines by means of pollination with fertile analoques
53

54. In the hybridization experiment based on self-incompatibility, the homozygous SI
MT line was crossed with fertile line FT 13. The mean seed weight per SI plant was
1.8 g and in the fertile parent FT 13 line 6.5 g. In the experiment based on CMS, the
maternal sterile line BR CMS was also pollinated with the line FT 13. The mean seed
weight per plant was 2.3 g in the CMS line and 18.3 g in the FT 13 male line. Lower
seed set, particularly in earlier female components could be due to curd reduction
caused by putrescence. It was caused by humid and cold weather, which made
chemical disease control ineffective.
Parental
genotype
Number of
grown plants
Number of
harvested plants
Weight of
harvested seeds (g)
Mean weight of
seeds per plant (g)
SI MT 24 22 40 1.8
FT 13 16 14 91 6.5
BR CMS 24 20 45 2.3
FT 13 16 15 275 18.3
Table 3 : Hybridization trial based on SI and CMS
54

55. The F1 hybrid of SI line Montano × self-pollinating line FT 13 turned out to be the
best combination in a preliminary field trial. The other hybrid BR CMS × FT
showed to be less uniform forming smaller and lighter curds.
SI line Montano × self-pollinating
line FT 13
BR CMS × FT
 Conclusion
In general, it can be concluded that both SI and CMS lines are suitable for
hybrid breeding of cauliflower. As regards for seed production, self-
incompatibility appears to be more effective than cytoplasmic male sterility.
On the contrary, CMS system provides much more reliable sterility than self-
incompatibility.
55

56. Combining ability, gene action and heterosis studies
involving SI and CMS lines and testers in cabbage
 Materials and Methods
Seven lines viz.,SI 1-4-6 (L1), SI 2008-09-03-01(L2) and II- 12-4-10 (L7)
(self incompatible) and CMS I (L3 ), CMS II (L4 ), CMS III (L5 ), CMS
GAP (L6 ) (cytoplasmic male sterile) were crossed to each of the four
testers viz., Glory-1 (T1), E-1-3 (T2), E-1-10 (T3) and SC 2008-09 (T4)
(pollen fertile) by line × tester mating design to develop 28 hybrids. The
hybrids were evaluated along with parents and two standard checks i.e,
standard check 1 (Varun) and standard check 2 (KGMR-1) in RBD with 3
replications. The plot size and spacing were 3.15 m x 0.90 m and 45cm x
45cm, respectively (14 plants/treatment). Observations were recorded on
5 randomly selected plants in each replication.
Palampur, India Thakur & Vidyasagar, 2016
Case study 5
56

57. Sr
No.
Sources of variation
df
Rep.
2
MSS
Crosses
27
MSS
Lines
6
MSS
Testers
3
MSS
L x T
18
MSS
Error
54
MSS
1. Plant spread (cm) 225.42* 20.99* 55.20@ 37.16@ 6.90 7.32
2. Non-wrapper leaves 6.36* 5.86* 15.65@ 8.72@ 2.12 1.59
3. Gross head weight (g) 92296.8* 53725.7* 139529.@ 108223.5@ 16041.4* 5025.8
4. Net head weight (g) 8322.47* 27319.2* 56131.9@ 58598.7@ 12501.8* 1834.1
5. Polar diameter (cm) 7.80* 2.45* 2.54@ 12.44@ 0.75 0.44
6. Equatorial diameter (cm) 9.06* 3.98* 10.82@ 4.28 1.65 1.14
7. Days to harvest 68.06* 5.43 4.70 9.47 5.00 8.42
8. Head shape index 0.001 0.03* 0.07@ 0.06@ 0.01* 0.01
9. Compactness of head
(g/cm3)
13.61 11.59 2.71 67.61@ 5.21 10.70
10. No. Of marketable head
per plot
8.94* 2.12 4.99@ 0.52 1.43 2.19
11. Heading percentage (%) 253.12* 58.25 139.37@ 6.04 39.91 53.56
12. Marketable head yield(kg) 0.19 3.69* 6.92@ 9.11@ 1.71* 0.28
Table 1: Analysis of variance for combining ability
Traits
57
‘*’ Significant when tested against MSS due to error and ‘@’ significant when tested against MSS due
to L x T interactions.

58. 58

59. 59

60. The proportional contribution of lines, testers and line tester interaction (in per cent) varied
from 5.20 (compactness of head) to 60.33 (equatorial diameter), 1.15 (heading percentage)
to 64.80 (compactness of head) and 19.90 (gross head weight) to 61.37 (days to harvest),
respectively. For marketable head yield the contribution of lines, testers and lines tester
interaction was 41.71, 27.42 and 30.85 per cent, respectively. In general, the contribution of
lines was the highest for most of the traits followed by line x tester interactions and testers.
These results conclude that there are good prospects of heterosis breeding in cabbage.
Table 5. Estimates of genetic components of variance and proportional (%)
contribution of lines, testers and their Interactions
60

61. x
x
61

62. Crop Name of Hybrid
Type of Genetic
Mechanism (Parentage)
Developing
Institution
Cabbage
Pusa Cabbage
Hybrid-1
Self-Incompatibility
(PCH-1=83-1-621 x GA-111)
IARI regional
station,
Katrain
Cabbage H-43, H-44
Self-Incompatibility
(H-43=S2S2 x PusaMukta)
(H-44=S2S2 x Cornell 83-6)
IARI regional
station,
Katrain
Cauliflower
Pusa Hybrid-2
Pusa Kartik Sankar
Self-Incompatibility
(Pusa Hybrid-2=CC x 18-19)
(Pusa Kartik Sankar= CC 14 x
41-5)
IARI, New Delhi
Cauliflower
Xiahua 6
( heat-resistant ) Self-incompatibility
Xiamen Agricultural
Research Institute of
Sciences, China.
Public sector hybrids developed by utilizing SI in cole crops
62

63. • Hybrid technology offers tremendous potential for the
much needed second Green Revolution.
• Expansion of area is not possible thus need is to
increase production/unit areas.
• Cost effective hybrid seed production.
• To fulfil the demand of vegetables in our country.
• To identify the stable SI lines.
• There is need to acquire deeper knowledge about SI
mechanisms for development of location
/environmental specified SI lines.
Conclusion and future prospects
63

64. 64