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Module 3 Overview of Plant Breeding

I. Introduction II. Setting Breeding Goals III. Mega Environments IV. Choosing Parents and Types of Crosses

a. Single cross b. Top and double cross c. Backcross d. Single backcross e. Synthetics and Wide Crosses f. Recording Pedigrees

V. Making the Cross VI. Selection Methods Following Hybridization

a. Breeding populations b. Heritability c. Selection procedures d. Genetic advance

VII. Genotype by Environment Interaction

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I. Introduction Plant Breeding is the science, art, and business of improving plants for human benefit (Bernardo, 2002; Breeding for Quantitative Traits in Plants). The science of plant breeding is based on genetics and the understanding of genes and heredity; but knowledge of other sciences surrounding plants (i.e. anatomy, physiology, biochemistry, soil science, pathology, and statistics) is necessary in order to be a successful plant breeder. The art of plant breeding revolves around the breeder’s savvy and keenness in observation, their intuition and judgment. The business of plant breeding is the management of resources (money, people, land, and time) in an effort to maximize returns. These three facets of plant breeding: science, art and business; must be used with an integrative approach to create and recognize that which forms the basis of plant breeding: variation.

Variation, among individuals of a population, is due to genetic and environmental effects. The genetic component of variation is provided by hybridization and recombination. The genetic variation observed in a population throughout a breeding program will depend on the parents used for hybridization, the selection procedure and the selection intensity, and on the interaction with the environment. In some cases, it will be desirable to limit the environmental effects on variation, but in some cases it will be beneficial to exaggerate an environmental effect in order to better select for superior genotypes. The goals of the breeding program will dictate the degree to which genetic and environmental effects should be manipulated. It is the job of the plant breeder, in

“The diversity of the phenomena of nature is so great, and the treasures hidden in the heavens so rich, precisely in order that the human mind shall never be lacking in fresh nourishment.”

Johannes Kepler

“The secret of improved plant breeding, apart from scientific knowledge, is love.”

Luther Burbank

Successful plant breeding depends on recognition of the natural diversity that can be found within the population. A plant breeder must attain an intimate knowledge of her crop. Without love, one wheat is the same as the next; with love, the nuance among individuals is made obvious.

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order to reach the goal, to explain and exploit these genetic and environmental effects that account for observable variation.

II. Setting Breeding Goals Before a single seed is sown and before a single cross is made a set of objectives must be established. In order to properly develop a set of breeding goals the breeder must understand the needs of his clients: the farmers, the millers, and the bakers. The breeder must understand the target environment, and the levels of biotic and abiotic stress that occur in that environment over each year and over multiple years. The breeder must understand the production practices of the farmers. The breeder must have knowledge of the economic infrastructure; are inputs such as fertilizers and pesticides readily available to the producer, and what product the grain will be processed into. The breeder must understand all these things and more if he is to properly develop an improved cultivar that will be adopted. The CIMMYT Wheat Program currently distributes advanced bread wheat lines to more than 60 countries. Primary clients are the national agricultural research systems, both private and public. CIMMYT breeding objectives attempt to address specific problems and limitations associated with wheat production in these countries. The primary goal of CIMMYT is to develop broadly adapted, high yielding germplasm that has high yield stability, durable disease resistance, and acceptable end-use quality. Soil type, temperature, and moisture levels greatly influence crop stability and productivity. In order to accomplish this primary objective over many diverse environments, CIMMYT has divided the wheat growing areas of the world into 12 distinct mega-environments (ME), delineated based on water availability, soil type, temperature regime, production system, and associated biotic and abiotic stresses. Breeding objectives have been identified for each mega-environment:

ME1: Irrigated, low rainfall environment (Spring Wheat) – ME1 represents the optimally irrigated, low rainfall areas of the world. The climate during the wheat growing period ranges from temperate to conditions of late heat stress. This ME encompasses about 36 million hectares spread primarily over Asia

• Spring Wheat - No vernalization, no photoperiod requirements (except ME6) • Facultative Wheat - Some vernalization/photoperiod requirements for optimal

growth • Winter Wheat - Vernalization and photoperiod requirements + cold tolerance

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and Africa between 35°S - 35°N latitudes. Breeding Objectives include high yield potential, yield stability, semi-dwarf stature, input responsiveness, lodging tolerance, improved industrial and end-use quality, and durable resistance to the three rusts (leaf rust, stripe rust, and stem rust). Certain areas require resistance to Karnal Bunt, powdery mildew, and fusarium head blight. Most areas require photoperiod insensitivity, but with a range of flowering dates. Some tolerance for late heat is also needed for certain locations. Greater emphasis will be given to tolerance to saline soils. White (amber)-grained types predominate in the vast majority of areas.

ME2: High rainfall environment (Spring Wheat) – ME2 is defined as those areas with average rainfall in excess of 500 mm during the cropping cycle. Total area exceeds 8 million hectares. Stripe rust, leaf rust, Septoria tritici, and pre-harvest sprouting are major production constraints. Fusarium head blight is becoming more widespread and is a serious problem in many areas. Resistance to barley yellow dwarf virus, bacteria, powdery mildew and the root rots must also be considered in certain regions of ME2. Tolerance to lodging, shattering, and soil micronutrient imbalances are becoming more important. For high yield potential, semi-dwarf stature is essential. Photoperiod insensitivity is preferred but with a wide range of flowering dates. Red-grained wheat provides better sprouting tolerance than white grained wheat, and is therefore generally preferred. Demands for better industrial quality are increasing, and emphasis is given to the development of germplasm with high quality.

ME1:

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ME3: High rainfall, acid soil environment (Spring Wheat) – The total estimated area is close to 2 million hectares. Disease and stress problems are similar to ME2 along with rice blast. Also, aluminum and manganese toxicities, plus phosphorus deficiency, are major constraints to production. Red grain is generally preferred, except in the Himalayas. High-level quality is demanded especially in Latin America.

ME4: Semi-arid, drought environment (Spring Wheat) – Three distinct types of drought have been identified based on the stage of plant development at which drought is most severe. These are: ME4A: Winter rain or Mediterranean-type drought associated with post-

flowering moisture and heat stress typical of the West Asia and North Africa region. Representative locations include Aleppo (Syria) and Settat (Morocco). Total estimated area: 6 million hectares. Stripe Rust, leaf rust, stem rust, Septoria blotch, root rots,

ME2:

ME3:

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nematodes, and bunts are the key biotic constraints. Also late frosts may occur. Photoperiod insensitivity is preferred but with a wide range in flowering dates. White-grained wheat with good quality is preferred.

ME4B: Winter drought or Southern Cone-type rainfall associated with pre-flowering moisture stress. Marcos Juarez (Argentina) is a representative location. Total estimated area is 3 million hectares. Resistance to leaf rust, stem rust, stripe rust, Septoria spp, fusarium spp. and tan spot are requirements. Pre-harvest sprouting is also a common problem, due to late rains; hence red seeded varieties are preferred. Photoperiod insensitivity is preferred but with a wide range in flowering dates. Good bread-making quality is demanded.

ME4C: Stored moisture after monsoon rains results in continuous or Subcontinent-type drought under receding moisture conditions. A representative location is Dharwar (India). Total estimated area is 2-3 million hectares, and probably decreasing, as irrigation facilities spread and/or other crop options are explored. Leaf rust occurs occasionally. Photoperiod insensitivity is preferred but with a wide range in flowering dates. Seed must be large, bold and amber in color to fetch a premium at the market place. Medium dough strength is required.

The breeding approach of CIMMYT attempts to combine high yield potential with drought resistance for ME4. The combination of water-use efficiency and water responsive traits plus yield potential is important in drought environments where rainfall is frequently erratic across years. When rains are significantly above average in certain years, the crop must respond appropriately (water responsive) with higher yields, while expressing resistance to the wider suite of diseases that appear under more favorable conditions.

ME5: Warm humid tropical environment (Spring Wheat) – These are generally located at low altitude (<1000 m), with a mean minimum temperature during the coolest month greater than 17°C. As a result relative humidity tends to be

ME4:

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high. The estimated area is about 9 million hectares. In these warm, humid locations, resistance to common root rot, and leaf rust, and Helminthosporium blight, along with tolerance to sprouting are major objectives. Photoperiod insensitivity is preferred with an emphasis on medium to early maturity. Input responsiveness is also an objective. Quality demands vary from moderate to high primarily using white grains.

ME6: High latitude environment (Spring Wheat) – The higher latitude (>45°N or

45°S) requires materials that have a degree of photoperiod sensitivity, which is different to other spring wheat MEs. Wheat is spring-sown in this ME because winters are too severe for survival. Typically this ME experiences pre-anthesis drought followed by rainfall during flowering and grain-filling. Resistance to Fusarium spp., tan spot, leaf rust, stem rust, and stripe rust along with tolerance to sprouting are breeding objectives in this environment. Very dry representatives of ME6 also occur in central and northern Kazakhstan (10 million hectares) and the southern Siberian wheat belt (8 million hectares). The major diseases are leaf rust and root rots. In addition, drought tolerance specific to the rainfall pattern in the region is needed. Taller wheat generally does better under these conditions. The total estimated ME6 area in developing countries is 20 million hectares. Wheat with high protein and strong dough are required.

ME7: Irrigated, moderately cold environment (Facultative Wheat) – In these irrigated regions temperatures vary from 0°C to 5°C. Yield potential, semi-

ME5:

ME6:

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dwarf stature, input responsiveness, and disease resistance are the main breeding objectives. ME7A: Fully irrigated. The major diseases are stripe rust, leaf rust, and

powdery mildew. A representative location is Zhenzhou, Henan (China).

ME7B: Supplementary irrigation. Representative countries are certain regions in Turkey, and Iran with stripe rust, and bunt as the major diseases; and in the Central Asian Republics with leaf rust, common bunts, stripe rust and loose smut as the main diseases.

ME8: High rainfall, moderately cold environment (Facultative Wheat) – Rainfall on average exceeds 600 mm per crop cycle, with temperatures varying between 0°C and 5°C. Objectives include input responsiveness and semi-dwarf stature. ME8A: Photoperiod-sensitive varieties are cultivated. The major diseases

are stripe rust, leaf rust Septoria spp., powdery mildew, Fusarium spp., and root rot. A representative location is Temuco (Chile).

ME8B: Photoperiod –insensitive varieties are cultivated. Representative regions are the transitional spring/winter wheat zones in Turkey with stripe rust and common bunt being the major diseases; and Thrace (Turkey) with leaf rust, root rot, powdery mildew ad common bunts as the main diseases. Other representative areas include certain regions on the Central Asian Republics where stripe rust, leaf rust, root rot, powdery mildew and common bunt are the main diseases.

ME7:

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ME9: Semi-arid, moderately cold environment (Facultative Wheat) – Rainfall is moderate to low (<400 mm), and temperatures are in the range of 0°C to 5°C. Drought tolerance is required and input efficiency an objective. ME9A: Heat stress at grain-filling. Mainly non-semidwarf varieties

cultivated. Areas in West Asia and North Africa and the Central Asian Republics are representative with stripe rust; common bunt and leaf rust the major diseases.

ME9B: Mainly semidwarf varieties cultivated. The major disease is leaf rust. Representative locations are certain regions in China.

ME9C: Less cold tolerance required than in 9A and 9B. Mainly semidwarf varieties cultivated. Areas in South America and South Africa are representative with leaf rust and Russian wheat aphid the major biotic stresses.

ME10: Irrigated, severely cold environment (Winter Wheat) –These irrigated areas experience very cold conditions with temperatures dropping to between 0°C

ME8:

ME9:

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and -10°C. High yield potential, semi-dwarf stature, cold tolerance, input responsiveness and disease resistance are requirements. ME10A: Fully irrigated. The major diseases are stripe rust, leaf rust,

powdery mildew, barley yellow dwarf virus and nematodes. A representative location is Beijing (China).

ME10B: Supplemental irrigation. Specific locations in Turkey and Iran are representative. Stripe rust, common bunt, root rot, and nematodes are the major diseases. Areas in Western and North-Western Iran where stripe rust and common bunts predominate; and in the Central Asian Republics where leaf rust, common bunt, nematodes, and powdery mildew predominate, are also representative.

ME11: High rainfall, severely cold environment (Winter Wheat) – Rainfall on

average in excess of 600 mm is common, while temperatures are low, between 0°C and -10°C. Semi-dwarf stature and cold tolerance are desirable traits. ME11A: Photoperiod-sensitive varieties cultivated. The major disease are

stripe rust, leaf rust, powdery mildew, Septoria spp., Fusarium spp., barley yellow dwarf virus, and root rot. No representative locations occur among the developing countries. However, other example locations are Martonvasar (Hungary), Krasnodar (Russia), and Odessa (Ukraine).

ME11B: Photoperiod-insensitive varieties cultivated. The major diseases are leaf rust, Fusarium spp., Septoria spp., root rot, Barley yellow dwarf virus, and stripe rust. A representative country is North Korea.

ME12: Semi-arid, severely cold environment (Winter wheat) – Conditions are harsh, with little rainfall (300-450 mm); temperatures vary from 0°C to -10°C. Drought tolerance is needed, cold tolerance, and zinc deficiency tolerance are objectives.

ME10:

ME11:

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ME12A: Heat stress at grain-filling. Mainly non-semidwarf varieties are cultivated in certain representative regions in Turkey, Iran, and Afghanistan. In these areas stripe rust, common bunt, root rot, nematodes, and zinc deficiency are the major biotic and abiotic stresses. In the Central Asian Republics stripe rust, common bunt, and leaf rust are the main diseases. In representative areas of China, semidwarf varieties predominate, with stripe rust and powdery mildew the key diseases.

ME12B: Medium heat stress at grain-filling. Mainly non-semidwarf varieties are cultivated in certain representative regions in Turkey, and Iran with stripe rust and common bunt the major diseases. In representative regions of China semidwarf varieties are generally cultivated, and stripe rust and common bunt are the main diseases.

Breeding goals will almost always revolve around yield and yield stability, resistance to one or more environmental stresses (winter hardiness, drought tolerance, nutrient deficiency or toxicity, disease), and quality. As every wheat breeding program has its financial limits, it is important to stay within the means of the program when deciding on the objectives. The numbers of individuals of a segregating population necessary to select from in order to improve a population can quickly get out of hand if too many traits are placed in the list of breeding objectives. Consider a trait controlled by a single locus with alleles A1 and A2, and the desired phenotype has the genotype A1A1. The genotypic ratio among inbreds will be 50% A1A1 and 50%A2A2. The probability of an inbred with the ideal genotype for this

simple trait will be: 1

( )

10.50

2Ideal GenotypeP = =

, and the population size (N) required to

have the ideal genotype will be 12 2N = = . However, in considering a trait such as yield, that is controlled by multiple traits, let us say 20, the probability of having the ideal

genotype for this complex trait will be: 20

( )

10.00000095

2Ideal GenotypeP =

, and the

population size required to have the ideal genotype will be 202 1,048,576N = = ; not an easy number of individuals to either maintain or screen.

ME12:

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III. Choosing Parents and Types of Crosses After the goals of the program are established, the next step will be to create the variability for the traits that are to be improved. Sources of germplasm for the crosses will need to be identified but locally adapted material, CIMMYT developed cultivars for the target ME, and exotic sources should be considered. Also, a decision on the best type of cross to carry out must be made: • Single Crosses (Figure 1) Single crosses are the cross of a cultivar or line with another cultivar or line. In some cases, a promising segregating line may be used as one or both parents. Procedure Select the female parents utilizing your objectives and your knowledge of the materials for the various crosses. It is necessary to maintain and enhance the variability to have an effective crossing program. Therefore, select several different lines as females for each one of your projected crosses, i.e. several with good rust resistance, others with short straw, others with excellent yield, etc. After the females have been selected choose the male parents. When selecting male parents you should try to complement the female parent, i.e. the male parent should have the good characteristics that are lacking in the female parent. For example, a line has been chosen for its resistance to Septoria but it is tall, susceptible to stem rust, and has poor yield potential. The male parent can be susceptible to Septoria but, it must be a dwarf, must have resistance to stem rust, and should have excellent yield potential. Making crosses in this manner will increase the chance of obtaining segregates with all of the desirable agronomic characteristics. Do not cross close sisters! Every cross should try to obtain lines which are better than the existing cultivars. To accomplish this, the parents of each cross must be analyzed (mentally) for the complete group of desirable agronomic characteristics. Never make a cross with only one objective, for example:

To obtain a line resistant to Septoria regardless of the other characteristics as this may in fact produce progeny that are resistant to Septoria, but the progeny have a poor chance of being agronomically acceptable. A cross may be made for Septoria resistance but the selected progeny must also have resistance to the other prevalent diseases and good yield potential etc. A line with Septoria resistance is of no benefit to the farmer if it will not yield.

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• Top and Double Crosses (Figure 2) Top or three-way cross is the cross of an F1 to a variety or line. Double cross is the mating of two Fl's. Procedures

In addition to the knowledge of the various varieties and lines, it is also necessary to have agronomic notes on the various Fls. A general idea of the maturity, height, straw strength and head characteristics must be noted before crossing begins.

The purpose of using top and double crosses is to increase the chance of obtaining a desirable gene or genes from exotic or difficult materials. Exotic refers to lines from other countries which are generally poorly adapted to local conditions. Difficult material refers to varieties or lines which are tall, poor combiners, or dominant susceptibles, etc. i.e. lines which have given poor results (progeny) from single crosses in previous crossing cycles. As in the case of single crosses, parents should be chosen to increase variability and to complement agronomic traits.

The F1 which will be the female parent should be chosen first, again following your crossing objectives. After emasculation, the male must be chosen. In making top and double crosses, only single cross Fl's are utilized because they are uniform. The top and double cross Fl's will be segregating and it is impossible to identify superior plants at crossing; therefore, they are not used. The Fl's are selected for desirable agronomic characteristics or for desirable parentage. For example:

P1 P2

Parent 1:

Good Yield Resistance to Fusarium Moderately Susceptible to Root Rot Moderate Gluten Strength

Parent 2: Excellent Yield Susceptible to Fusarium Resistance to Root Rot Strong Gluten Strength

Breeding Goal: Excellent Yield, Resistance to Fusarium and Root Rot, and Strong Gluten Strength

X

self

self

self { Segregating Population

IC

Improved Cultivar: Excellent Yield Resistance to Fusarium Resistance to Root Rot Strong Gluten Strength

Fig. 1 - Single Cross

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An F1 made between a tall, poor yielding line with excellent leaf rust resistance and a semi-dwarf line with excellent yield but poor resistance is chosen. To enhance the probability of obtaining all of the desirable characteristics, top or double cross this F1 to a line or F1 with the desirable characteristics but of different parentage to increase variability. Another example could be the objective of broadening the base of stem rust

resistance: Line A, tall, poor yield, poor quality and excellent stem rust resistance was crossed to line B short, good yield, good quality and good stem rust resistance. Since line A had many negative values, it probably would be desirable to top or double cross this F1. Therefore, select a line of F1 which is short, has good yield, good quality and good resistance.

The Choosing for Top or Double Crosses

The question of whether or not to top or double cross an F1 can frequently be answered by the amount of labor available. Top crosses require 5 heads per cross and double crosses require 8-10 heads per cross. This number is necessary because these crosses will segregate in the next F1 generation and at least 80 and 150 plants are required from top and double crosses respectively, to facilitate the selection of desirable plants in the F1. Therefore, if labor is expensive top crosses should be utilized more than double crosses.

Another criterion in solving the question of whether to top or double cross is the amount of variability that you think will be necessary to obtain desirable phenotypes. Double crosses segregate more widely than the top cross in the F2. Double crosses

PA PB PC PD

X

F1AB

X

F1CD X

self

self

self

(In the case of a top cross this circle would represent parent “C”)

Parent A: Tall Poor Yield LR ResistanceB

Parent B: Semidwarf Excellent Yield Poor LR Resistance

Parent C:

Tall Poor Yield LR resistanceA Parent D: Semidwarf

Excellent Yield Moderate LR ResistanceC

IC

Improved Cultivar: Semidwarf Excellent Yield Durable LR ResistanceABC

Fig. 2 - Double Cross

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involve 4 different parents, generally 3 desirable and one less desirable, while the top cross involves 3 parents, 2 desirable and one less desirable parent. Therefore, the decision to top or double cross depends on the number of poor features in the exotic lines. Frequently, an exotic line has only one desirable trait. If this is the case, single crosses will be ineffective and top or double crosses must be utilized to increase the proportion of desirable segregates.

• Backcross (Figure 3) It may occur that a certain cultivar is both adapted to and productive in a certain region, but lacks a desirable allele for a single trait. This desired trait is found in other cultivars, but these cultivars perform very poorly overall in the target environment. The goal of the back cross is to introduce the exotic allele or alleles into the adapted varieties background. Beginning in the F1 and continuing for several generations, hybrid plants containing the desired allele are selected and successively crossed back to the adapted parent cultivar. The adapted cultivar, to which the successive crosses are being made to, is known as the recurrent parent. The other parent, containing the desired allele that is used in the initial cross is called the donor parent. The number of backcrosses used depends on how completely the breeder wishes to recover the recurrent parent’s genetic background. This varies from between two to five, or more; but in theory, after the fifth cross 96.875% of the recurrent parent’s genes will be recovered in the backcross progeny. The backcross procedure is most easily carried out if the trait being added is simply inherited, dominant, and easily recognized in the hybrid plants (Pohlmen and Sleper, 1995). If the desired phenotype occurs with the homozygous recessive genotype (aa), the trait will not be identifiable after the first cross as the progeny will be segregating with genotypes (AA) and (Aa). It is necessary to self the progeny one generation in order to identify the desired genotype (aa) before making the next backcross. Backcross procedures are well established and straight forward, and results are predictable. It can be very efficient as small numbers of plants can be grown in the first generations of backcrossing, so it can be the least expensive way of improving a cultivar, or developing improved parent material. Recurrent parents are often identified in locally adapted material, where a single weakness is frequently recognized. This method is a rapid and easy response to the weakness. In a dynamic and progressive breeding program backcross methods may not be a way to realize the breeding objectives as by the time the backcross generations have been completed (perhaps 5 or 6 years), the recurrent parent of the backcross may no longer be looked at as an elite cultivar, and the product of the backcross procedure may likely perform worse than the cultivars that have been produced using the single cross or double cross methods. Figure 3 details the backcrossing method to introduce a dominant allele (RR) into a recurrent parent’s genetic background.

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.

75% genes

from A

50% genes

from A

Selfing

Cultivar B

(RR)

Cultivar A

(rr)

F1

(Rr)

Cultivar A

(rr)

Cultivar A

(rr)

Cultivar A

(rr)

Cultivar A

(rr)

X

X

X

X

X

Discard (rr)

Discard (rr)

Discard (rr)

Discard (rr)

(Rr) (rr)

(Rr) (rr)

(Rr) (rr)

(Rr) (rr)

1st Backcross

2nd Backcross

3rd Backcross

4th Backcross

F1 of BC4

87.5 % genes

from A

93.75 % genes

from A

96.875 % genes

from A

Discard (Rr) on basis of progeny test

F2 of BC4 Discard (rr)

(RR)

(Rr)

(rr)

Fig. 3 - Diagram of the backcross method. The average proportion of alleles from cultivar B (the donor parent) in the hybrid population is reduced by one-half with each backcross. Because R is dominant to r, plants of genotype Rr can be recognized after each backcross and used for the next backcross. Plants of genotype rr are discarded. At the end of the process the selected progeny will be over 96% genetically similar to cultivar A (the recurrent parent) and carry also the donor parents RR alleles

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• CIMMYT and the Single Backcross Strategy CIMMYT breeders typically use a single backcross strategy. This strategy allows the breeder to create a breeding population that maintains most characteristics of certain cultivar (the recurrent parent), but still allows selection for several new genes. In effect the single backcross strategy enables three things to occur:

1. Increase the probability of maintaining and reselecting desirable genes of the recurrent parent.

2. Allows for the transfer of multiple genes or characters simultaneously. 3. Allows additional genes or characters from the donor parent to be selected for.

The basic method of a single backcross at CIMMYT is to cross an adapted cultivar with 6-10 donor parents that are selected based on the needs identified in the breeding objective. F1 plants are, from each of the 6-10 crosses, then backcrossed to the adapted cultivar with the goal of developing 400-500 BC1 seeds. It has been shown that this crossing method has been successful in creating breeding populations with a higher mean value for yield than through traditional simple or three way crosses (Fig. 4).

0

5

10

15

20

25

<6060

-65

65-7

070

-75

75-8

080

-85

85-9

090

-95

95-1

00

100-

105

105-

110

110-

115

Grain yield (% Kambara )

% L

ines

Single back cross Traditional

N = 4088N = 726

0.8% > Check

10.7% > Check

Fig. 4 - Grain yields of wheat lines developed through traditional (Simple and 3-way crosses) and single-backcross approach

Cd. Obregon 2004-2005

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Synthetics and wide crosses CIMMYT scientists have been crossing durum wheat with wild relatives of wheat, such as goat grass (Aegelops tauschii), since the early 1990s. This cross creates synthetic wheats, which can easily be crossed with improved varieties to incorporate new, useful genes. The resulting wheats are improved varieties that also have desirable traits from the wild parents. CIMMYT has produced synthetic wheats and their derivatives with traits such as resistance to Septoria spp. and Fusarium spp. head blight and also tolerance to drought, heat, salt, or waterlogging. Researchers in China recently crossed CIMMYT synthetic wheats with local wheats and released the results to farmers in 2003. Breeders in Sichuan province have been using the CIMMYT-developed synthetic hexaploid wheat since 1995 to improve quality, yield potential, and disease resistance. After crossing and backcrossing this wheat with high-yielding local varieties, they have developed several lines and are currently testing five more. The synthetic wheats pass on beneficial traits such as large kernels, heavy spikes, and resistance to new races of Chinese stripe rust. During two years of yield trials, the two varieties derived from synthetic wheats had 20% to 35% higher yields than the commercial check variety. One of these varieties, named Chuanmai42, had the highest average yields – more than six tons per hectare – in the trials. Since it was released in Sichuan in 2003, Chuanmai42 has been recommended by the government to farmers and has been delivered to most wheat breeding research programs in China. In 2003, Spain registered a CIMMYT synthetic wheat derivative under the name Carmona. This fast-growing variety matures and provides seed in a shorter period than most commercial cultivars, which is valuable for wheat growers who often plant late in the year in southern Spain. Carmona has better grain quality and is suited to zero-tillage systems, where it resists foliar diseases and produces higher yields. Wide crosses involve transferring alien genes into wheat. An alien gene can be considered to be any gene transferred to wheat from a related species (Knott D., Wheat and Wheat Improvement). The genus Triticum contains a broad range of species some of which cross readily and some of which cross with great difficulty to bread and durum wheats. Wheat will also cross with species in a number if different genera including: Agropyron, Elymus, Elytrigia, Haynaldia, Hordeum, Leymus, and Secale. These relatives of wheat are adapted to a large range of environments and are potential sources for useful genes such as: disease resistance, cold tolerance, salt tolerance, drought tolerance, lodging resistance, early maturity, and yield. Wheat 1B/1R translocation, a common alien transfer, is the wheat translocation line in which the short arm of 1B chromosome in wheat is replaced by the short arm of 1R chromosome in rye. Because of the characteristic of disease resistance, high and stable yield and broad adaptation, it has been widely planted around the world. In China the proportion of 1B/1R translocation lines has reached 45% in major extended cultivars of main wheat growing areas in the past thirty years and partial recently bred lines (Chai J et al (2006) Plant Genomics in China VII). Synthetics and Wide Crosses along with Hybrid Wheat are discussed further in the module: Biotechnology and Wheat Breeding.

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• Recording Pedigrees Pedigrees provide the parentage or the sequence through which a cultivar was obtained and are important sources of information. The pedigree for each line is generally entered into field books, crossing books, and on crossing tags. At present, CIMMYT uses the United States Department of Agriculture (USDA) system of designating pedigrees. This system is widely used around the world, making it easier to trade data. This system consists of just a few basic rules:

1. The female parent is written first and is separated from the male parent by: a. A single slash “/” in the first cross

Example: Logan / Heart b. A double slash “//” in the second cross

Example: Logan / Heart // 3270-A / Rusalka c. Two slashes with the number of the cross between them, /3/, /4/, /5/, etc…

in the subsequent crosses. Example: Logan / Heart // 3270-A / Rusalka /3/ Tn-1685 / IA-22 // 6767 / 216-6-3

2. The parental material involved in any particular cross includes all that is listed on either side of the highest number of crosses in the pedigree.

Parent 1 pedigree: Logan / Heart // 3270-A / Ruskalka Parent 2 pedigree: Tn-1685 / IA-22 // 6767 / 216-6-3

3. Backcrosses are indicated with an asterisk (*) and a number indicating the number of times the recurrent parent was used. The asterisk and the number are placed next to the crossing symbol that divides the recurrent parent and donor parents.

Examples: LR 64 / 3* Son64 Son 64 / *2 TZPP

IV. Making the Crosses There are some important things to consider before carrying out the crossing of two cultivars. The plants to be used as the females should be emasculated slightly (3 to 4 days) before dehiscence of the male plants. This can become complicated if the cultivars to be used have different heading dates. If the wheat to be crossed is a winter wheat, it will need to vernalized for usually 8 weeks. Emasculation is a time consuming process that takes patience, and concentration; if too many emasculations need to be done in a single day, anthers will be left behind and self fertilization will occur. A way to avoid

HOPEWELL

LOGAN HART 3270-A RUSALKA TN-1685 IA-22 6767 216-6-3

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some of these potential problems is to plant more than you will need of the selected parents over a number of days, many of these plants will then not be used but it will ensure that both the female parent and the male parent will be at the proper flowering stage when you are ready. Figure 5 shows a wheat spikelet and may be useful to those unfamiliar with emasculation.

In order to make a cross the breeder will need some basic materials: Small, sharp, fine quality scissors; forceps or pincers with somewhat blunted ends; 3x5 cm tags with an attached string; 5x15 cm glassine bags; a wax pencil; a permanent marker; staples; paper clips; rubber bands; and a crossing block book.

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Preparing Crossing Tags

Figure 6

1. Assembly and Preparation of Tags a) Ready tags before crossing starts b) On side 1, identify year and location (MV-07) c) On side 1, write or stamp date of emasculation (VII-31), and date of pollination

(VIII-2) d) The initials of the individual doing the crossing are sometimes added as well.

2. Identify female Parent a) On the top half of side 2, write the nursery plot number (CB HARI, 1), the name

and pedigree of the female parent (Glennson). 3. Identify the male parent

a) Usually it is easier to prepare all of the needed tags for each female first and then to add the name and pedigree of the male. Select the male parents from the nurseries for each cross (CB HARI 15). Place the name and pedigree of the male parent on the lower half of the tag (BOW “S”; CM 33203…).

b) The complete pedigree must be made on at least one tag. At CIMMYT, one complete tag is made for single crosses, two for top crosses and three for double crosses. You can abbreviate the name on additional tags, but be sure you can positively identify the parents.

Steps of Emasculation: 1. Select Lines and Varieties to Be Emasculate

a) The number of spikes to be emasculated will depend upon the genetic stability, (more for segregating materials in top and double crosses) seed set, and the desired size of the F1 population.

b) Specify the number of spikes to be emasculated for each cross i) For each single cross: 3-6 ii) For each top or double cross: 8-13

2. Select Individual Plants and Spikes for Emasculation a) Individual plants should be healthy and representative of the variety or line. b) Do not emasculate more than one spike per plant. c) Select spikes which are almost fully exerted from the flag leaf. d) Do not select spikes which have begun anthesis. Emasculation should be done 3

to 4 days before expected dehiscence.

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e) Carefully separate the selected spike from those surrounding it in order to make work easier, being careful not to break the stem.

3. Remove Upper and Lower Spikelets a) With your scissors or forceps, remove one or two spikelets from both the bottom

and the top of the spike. These spikelets are removed because they mature more slowly than the middle spikelets and very often aborted or produce smaller seeds.

b) Keep 15 to 20 spikelets per plant. 4. Remove the Center Florets

a) With your forceps pull out the center florets of each spikelet, leaving only florets 1 and 2.

b) This is done for two reasons: i) It makes the emasculation much easier. ii) The ovaries of these florets do not mature at the same time.

5. Cut Back the Lemma and Palea a) With your scissors cut the glumes, lemma, and palea just above the top of the

stigma b) Do not injure the stigma or the anthers by cutting too low. c) If the cut is too high, emasculation is more difficult and pollen may not reach the

stigma as the palea and lemma will remain folded over the stigma. ** Steps 4 and 5 may be reversed if it is more convenient for the emasculator **

6. Remove the Anthers a) Carefully remove the three anthers from each floret with the forceps. b) With practice, you will be able to remove all three florets at one time. c) Remove the anthers carefully. Do not break them, as pollen may be shed causing

self pollination. d) The extracted anthers may stick to the forceps. An easy clean way of removing

the anthers from the forceps is to eat them. e) Do not injure the stigma with forceps. f) At first, you will want to check each floret after you have completed the spike;

with experience, double checking may not be necessary. 7. Cover the Spike with Glassine Bag

a) Mark the date and your initials on the glassine bag b) Place bag over spike. c) Fold bottom edge over and staple or place a paper clip on fold so that it is tight

against the peduncle or stem to prevent wind from blowing off the bag. 8. Keep record of which plants have been emasculated in your crossing block book.

This way you will know when it is time for pollination. Pollination by the Twirl Method 1. When to Begin Pollination

a) Depending on temperature and humidity, the female parent will be ready for pollination two to four days after emasculation. The female is receptive when the stigma is enlarged and feathery.

b) A crossing tag, representing the cross that is to be made, should be made the day before pollination will take place.

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2. Gather Pollen a) Take one spike, as a pollen source, for each tag. b) Select spikes from healthy plants. c) Spikes should just be ready to flower – one or two anthers may show. d) Pull the spike from the stem; if it is ready to flower, it will separate easily.

3. Tie the Spikes Together a) Group all spikes of the same male parent, and fasten with a rubber band. b) Attach appropriate tags to each group of spikes. c) Clip peduncles to uniform length if desired.

4. Place Spikes in Water a) Place the spikes in water and put the container in the shade. b) Spikes may be kept for four to five hours. c) Be sure to maintain the identity of each pollen parent.

5. Clip the lemma and palea a) Clip the lemma and palea just above the anthers. This cut will be somewhat

higher than the cut for emasculation. b) Clip the number of spikes needed for each cross to a particular female.

6. Place the Spike in the Sun a) Place the peduncles in the ground in a sunny place protected from the wind; or if

it is cold, hold the spike in your cupped hands and blow on it gently until anthers emerge.

b) While the other spikes are clipped the anthers of the first spikes will extrude and are then ready to be used for pollination.

7. Cut Open the Top of the Glassine Bag a) With the scissors cut open the top of the glassine bag of the selected female

parent. b) Blow into the bag to open it. c) Keep open only long enough to pollinate the enclosed spike.

8. Pollinate the Emasculated Spike

a) Carefully place the pollen spike in the bag. Avoid dusting the pollen outside the bag.

b) Twirl the spike around the female several times, and then discard the pollen spike. c) One male spike is enough to pollinate the female with just two or three good

anthers extruded 9. Close the Bag

a) Fold the top of the glassine bag over. b) Place a paper clip on top, or staple to hold in place.

10. Attach Crossing Tag a) Attach the tag specific to each cross to peduncle. b) Place the tag under the paper clip, at the bottom of the glassine bag with side

showing dates of emasculation and pollination out.

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V. Selection Methods Following Hybridization Following hybridization, the breeder must establish and maintain the segregating population (that ideally has a high mean and a large genetic variance for the desired traits) from which he will make selections. Selection occurs when some individuals produce more progeny than others due to a superior genotype. In nature, this is due to either superior fertility or superior viability; but in a crop, such as wheat, the decision of which individuals will or will not produce progeny is largely in the hands of the plant breeder. The job of the breeder is to, with the breeding objectives in mind, select individual or groups of genotypes such that the population contains all the alleles that confer the superior phenotype, and none of the alleles that confer the inferior phenotype. Before selection can be further discussed, it will be important to understand some things about population genetics, gene action and interaction and heritability. The presentations, of the above concepts, that are to follow are based on mathematical models of probability developed by Godfrey Hardy and Wilhelm Weinberg. Although, the assumptions that are made in order to justify these mathematical models are rarely seen in the practice of plant breeding (random mating, large population size, no selection, no mutation, and no immigration or emigration) the concepts are useful in understanding sources of variation within the crop and will aid the breeder in making sound decisions during the selection process. Breeding Populations The population of plants created by crossing two or more parents is known as a breeding population, and a population can be described in terms of its genotypic or allele frequencies. Considering one locus, a population of 900 individuals is a population of 1800 alleles. For example, at a single locus A, an F1 population derived from two homozygotes A1A1 × A2A2 will have genotype frequencies (P), and allele frequencies (p the dominant allele and q the recessive allele) of:

Genotype Frequency (P) A1A1 P11 = 0.0 p = P11 + ½ P12 = 0.5 A1A2 P12 = 1.0

A2A2 P22 = 0.0 q = P22 + ½ P12 = 0.5 The sum of allele frequencies at any locus is p + q = 1 (two alleles); and a locus with three alleles: (p1 + p2 + p3) = 1. A population, after one generation of random mating will have the allele frequencies of p2 + 2pq + q2. As an example if a population with the following genotypes were to randomly mate:

Genotype Number Pij A1A1 20 0.2 p = P11 + ½ P12 = 0.4 A1A2 40 0.4 A2A2 40 0.4 q = P22 + ½ P12 = 0.6 total 100 1.0 0.1

The next generation’s genotype frequency would be:

( ) ( ) ( )2 2 21 1 1 2 2 2( ) 2 0.16 0.48 0.36 1.0p q p pq q A A A A A A+ = + + = + + = .

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Likewise for the three allele locus: Genotype Number Pij A1A1 20 0.2 A1A2 10 0.1 p1 = P11 + ½ P12 + ½ P13 = 0.275 A1A3 5 0.05 A2A2 15 0.15 p2 = P22 + ½ P12 + ½ P23 = 0.35 A2A3 30 .30 A3A3 20 0.2 p3 = P33 + ½ P31 + ½ P32 = 0.375 Total 100 1.0 1.0 and the next generation’s genotype frequency will be:

( )( ) ( ) ( ) ( ) ( ) ( )

2 2 2 21 2 3 1 1 2 1 3 2 2 3 3

1 1 1 2 1 3 2 2 2 3 3 3

2 2 2

0.076 0.193 0.206 0.123 0.263 0.141

1.0

p p p p p p p p p p p p

A A A A A A A A A A A A

+ + = + + + + +

= + + + + +=

.

A superior phenotype is a result of both its genotype and the environment which it is in. The phenotypic value of individual k with the AiA j genotype can be modeled as

( ) ( )ijij k ij kP G e= +

and as a deviation from the population mean

( ) ( )ijij k ij kP g eµ= + +

where ( )ij kP is the phenotypic value of individual k with the AiAj genotype, µ is the

population mean, Gij is the value of the AiAj genotype (gij being Gij-µ), and ( )ij ke

represents the value for the non genetic effects (the environment) that individual k with the AiAj genotype is exposed to. Value, refers to the measure of the effect on the quantitative trait. Genotypic values cannot be measured directly but can be estimated from phenotypic values. The value of a genotype, AiAj, can be modeled as a deviation from the midparent value,P , the average of the two parental values. The value a is given to the A1A1

genotype, the value –a is given to the A2A2 genotype, and the value d is given to the A1A2 genotype (see fig.6). The value of d can very depending on the degree of dominance exhibited by one allele over another. Partial dominance exists when 0<d<a; no dominance exists when d/a = 0; whereas d/a = 1 indicates complete dominance. Over dominance exists when d>a.

- 26 -

Figure 6

At a single locus, the mean (µ) of a population is equal to the sum of genotypic values multiplied by their allele frequencies. As an example let us consider the following population of 100 individuals with single locus and no dominance of the A1 allele:

Genotype Number Pij Trait Value A1A1 63 0.63 20 p = P11 + ½ P12 = 0.705 A1A2 15 0.15 18 A2A2 22 0.22 10 q = P22 + ½ P12 = 0.295 Total 100 1.00

Using the coded genotypic values, at a single locus the mean (µ) of the population following random mating will be:

( ) 2

15

5

3

5

15 5(0.705 0.295) 2(0.705)(0.295)(3)

18.30

P a p q pqd

P

a

d

a

µ

µ

= + − +===

− = −= + − +=

When the trait is controlled by two or more loci that do not interact with each other, the population mean is the sum of the contributions from each locus:

( ) 2l l l l l l lP a p q p q dµ = + − +∑ ∑ ∑

where l refers to the lth locus. Coded genotypic values are reflections of genotypes, not alleles, and it will be more useful to know the average effect of an allele that is passed on. Ronald Fisher, a founder of today’s statistical science, formulated the average effect of an allele. The average effect of allele A1 expressed as a deviation from the population mean, is denoted by α1.

- 27 -

( )( )

1 ( ) 2P pa qd P a p q pqd

q a d q p

α = + + − + − +

= + −

Likewise, the average effect of the A2 expressed as a deviation from the population mean, can be calculated as:

( ) ( )[ ]

2 2

(

P pd qa P a p q pqd

p a d q p

α = + − − + − +

= − + −

These equations show that the average effect of an allele is not a property of the allele itself, but it is a joint property of the allele and the population in which the allele is found. During the selection process one allele will be favored over another and this implies an allele substitution. The average effect of an allele substitution (A1 for A2) is equal to:

1 2

( )a d q p

α α α= −= + −

.

Given the above, the genotypic value of AiAj can be expanded from:

ij ijG gµ= + to ij i j ijG µ α α δ= + + + ,

where δij is the dominance deviation. Dominance deviations imply that the heterozygote is more like one parent or the other. Dominance deviations are zero in the absence of dominance, and are modeled by:

( )

11 11 1 1

2

12 12 1 2

22 22 2 2

2

2( )

2

( )

2

2( )

2

G

a pa qd

q d

G

d pa qd pd qa

pqd

G

a pd qa

p d

δ µ α αµ µ

δ µ α αµ µ µ

δ µ α αµ µ

= − − −= − − + −

= −= − − −= − − + − − − −== − − −= − − − − −= −

Epistatic effects are the result of interactions of genes affecting a single trait but are found at different loci. Epistatic effects exist when the total of the effect is not equal to the sum of the individual allele effects. With that in mind the two-locus model can be introduced with individual AiAjBkBl, and Iijkl representing the epistatic effect:

( ) ( )ijkl i j ij k l kl ijklG Iµ α α δ α α δ= + + + + + + + .

As stated before, variation is the key to plant breeding; and variation within the population (assuming equal environments) is a function of alleles. It is the goal of the plant breeder to shift the population’s distribution of the trait value by selecting favorable alleles, and in a sense altering the variation within the breeding population. As the previous models showed a phenotype is a function of both genotype and environment;

- 28 -

In the same manner phenotypic variance of a population can be explained by genetic variance and environmental variance. The variance is defined as the mean of the squared deviations of a random variable from the population mean. The population variance (σ2) for variable X is equal to the expectation 2( )i XE X µ− , but would be

difficult if not impossible to calculate so the sample variance is generally used: 2

2 ( )

1X

x xs

n

−= −

∑ .

And just as genotypic values could be broken down into a mean along with additive effects, dominance effects, and epistatic effects; so too can the genotypic variance be broken down into different variance components and can be useful in the selection process:

Variance Component Symbol Phenotypic VP

Genetic VG

Additive VA

Dominance VD

Epistatic VI Genotype x Environment VGE

Error VЄ

Further:

Fig. 7 Selection for a quantitative trait shifts the population distribution.

- 29 -

[ ]

[ ]

( )

( ) ( ) ( ) ( ) ( )

( )

( ) ( ) ( ) ( ) ( )

P G e ij k

G A D I i j ij k l kl Iijkl

GEe ij k

P i j ij k l kl Iijkl GE

V V V

V V V V V V V V V

V V V

therefore

V V V V V V V V

α α δ α α δ

ε

α α δ α α δ ε

+ +

+ +

= +

= + + = + + + +

= +

= + + + + + +

This states that phenotypic variance is a function of (in a two loci system) variance for the additive and dominant effects for alleles A1, A2, B1, B2, their epistatic relationship with each other, and how they interact in the environment, along with unexplained variance. Next we will take the theoretical models introduced in the previous pages and put them towards a few useful applications (For a more inclusive look at population genetics see Rex Bernardo’s text: Breeding for Quantitative Traits in Plants from where much of the information was gathered. Heritability Individual plants in a breeding population will ideally vary for the traits identified in the breeding goals. These will include height, heading and maturity dates, disease resistance, quality parameters, and yield. Two randomly selected plants will show these variations but it will not be known whether the differences are due to genotype or the environment. The plant breeder is interested in identifying how much of the observed phenotypic variability is due to genetics and to do this she must estimate heritability . Heritability is defined and estimated in two distinct ways: Broad-sense heritability and Narrow-sense heritability. Broad-sense heritability (H) estimates the proportion of the phenotypic variance that is explained on the basis of all genetic effects:

G PH V V= .

Calculation of broad-sense heritability can be done by using data to estimate the variance components (ANOVA). These procedures will not be detailed here but can be found in statistics and some quantitative genetics books. However, in a cross between two pure lines an estimate of the broad-sense heritability can be calculated by using the variance of the F2 population as VP. Environmental variance, or error variance (VE), can roughly be estimated by using a uniform population grown in proximity to the segregating F2

population since:

P G E

G P E

V V V

V V V

= += −

Narrow-sense heritability (h2) estimates the proportion of the phenotypic variance that is explained by only the additive effects:

- 30 -

2A Ph V V= .

Narrow-sense heritability can be estimated by finding the slope of a line from regressing progeny values on parent values:

2 2( )( ) / ( )h b x x y y x x= = − − −∑ ∑

with x representing the parent values and y representing the progeny values. The x and y data are taken from measurements of a quantitative character. Quantitatively inherited traits differ in heritability. A polygenic trait such as yield, influenced greatly by the environment, will have low heritability; while monogenic and oligogenic traits unaffected by the environment will have high heritability. Selection from a population in the F2 generation will not be very effective for traits with low heritabilty; while selection for traits, within the same F2 population, that have high heritability estimates can be very effective. Examples of traits with relatively high heritability are: heading date and kernel size; while traits generally having low heritability are yield, lodging resistance, and protein content. Selection procedures that may be used to identify favorable genotypes from segregating populations following hybridization for wheat will be explained in the following figures and are: Fig. 8 Pedigree Selection, Fig.9 Bulk-Population Selection, Fig.10 Single Seed Descent, Fig.11 Modified Pedigree Selection, and CIMMYT’s current preferred selection method called the selected bulk method shown in figure 12. It is implied that each subsequent generation after the initial hybridization is a product of self pollination. Bear in mind that the following examples are basic ideas and may vary according to specific breading goals and resources.

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Fig. 8 Pedigree-Selection Method

Modifications of the pedigree-selection method can be made, such as introducing yield trials as early as the F3 and F4 generations. Keep in mind that progeny from a single plant are 50% more identical than the previous generation’s progeny. The pedigree method is labor intensive and requires detailed record keeping. The advantage is that only progeny with desirable characteristics are carried forward to the next generation. This method also permits collection of genetic information that can be used as variation among and within families can be partitioned allowing for a closer examination of narrow sense heritability and selection efficiency.

Cross cultivar A by cultivar B according to the breading goals that have been previously identified PA PB X

F1- Grow 50 to 100 plants.

F2- Grow 2000 to 3000 plants. Harvest seed from individually selected plants, keeping seed from

each separate.

F3- Grow head rows with seed harvested from

superior plants. Identify superior rows, and

select the best 3 to 5 of the best plants within

rows. Maintain identity of plant and row while

recording phenotypic data. Continue selection

between and within rows through the F5

generation.

F4

F5- Normally, 25 to 50 families are harvested at the

end of this generation.

F6- Grow families of head rows. Rouge off types.

Uniform related families can be harvested

together and the seed bulked. Harvested Seed

lots are designated experimental lines.

F7- Experimental lines are grown in preliminary yield

trials in comparison with local checks.

F8 to F10 – Yield trials of superior experimental lines

are tested at multiple locations with local checks. Only the lines identified as

having all the traits desired, are

maintained for testing in subsequent generations.

F11- A cultivar found to be superior to local checks is named, certified, and released for increase and

distribution

- 32 -

Fig. 9 Bulk-Population Selection Method

The bulk-population method of breeding is simple, convenient, requires less labor, and is less expensive to conduct during the early segregating generations than the pedigree-selection method. It is necessary to grow a large population of the spaced plants (F5 above) to have a reasonable chance of finding desirable segregates. During the early segregating generations there is a good chance that some desirable genotypes will be lost as they are suppressed by neighboring plants that are taller or have a more spreading habit.

PA PB X Cross cultivar A by cultivar B according to the

breading goals that have been previously identified.

F1- Grow 50 to 100 plants. Harvest en masse and bulk seed

F2- Grow 2000 to 3000 plants. Harvest en masse

and bulk seed from all plants.

F3 to F4- Grow 100m2 plots with bulked seed from

previous generation. Plots can be

subjected to biotic and/or abiotic stresses in order to foster natural selection.

F5- Space plant 3000 to 5000 seeds. Select and harvest 300 to 500 superior plants keeping

seed separate from each plant.

F6- Grow head rows of selected plants. Select 30 to 50 head rows that show desirable

characteristics. Bulk seed from selected rows.

F7- Grow plots made of selected rows from F6 in

preliminary yield trials.

F8 to F10 – Yield trials of superior experimental lines

are tested at multiple locations with local checks. Only the lines identified as

having all the traits desired, are

maintained for testing in subsequent generations.

F11- A cultivar found to be superior to local checks is

named, certified, and released for increase

and distribution

- 33 -

Fig. 10 Single-Seed-Descent Method

The single-seed-descent method was developed as a way to maintain a maximum of diversity in the F2 generation, and reduce the loss of genotypes during the segregating generations. The single-seed-descent method can also reduce the time necessary to grow segregating generations. There is also no need for record keeping in the early segregating generations. As only one seed is harvested from each plant careful development of the F2 to F4 generations is not necessary. In a greenhouse; by thickly planting seeds, with low soil fertility, and using light regimes to force early flowering, two or three generations can be harvested in a 1-year period. Undesired genotypes, however, will not be eliminated in the early generations, and as no family structure is maintained, superior segregate families can not be identified.

PA PB X Cross cultivar A by cultivar B according to the

breading goals that have been previously identified.

F1- Grow 50 to 100 plants. Harvest seed en masse and bulk.

F2- Grow 2000 to 3000 plants. Harvest a single

seed from each plant.

F3 and F4 - Grow seed harvested from previous

generation harvest a single seed from

each plant.

F5- Space plants in field with single seeds harvested from F4. Select plants according to breeding

goals and harvest seeds from selected plants.

F6- Grow head rows with seed harvested from F5.

Harvest rows superior for desired traits.

F7- Grow plots made of selected rows from F6 in

preliminary yield trials. F8 to F10 – Yield trials of superior experimental lines

are tested at multiple locations with local checks. Only the lines identified as

having all the traits desired, are

maintained for testing in subsequent generations.

F11- A cultivar found to be superior to local checks is

named, certified, and released for increase and

distribution

- 34 -

Fig. 11 Modified Pedigree-Bulk Selection Method

With the globalization of CIMMYT’s Bread Wheat Breeding Program in the 1980s and the evolution of the concept of 12 MEs, the number of crosses made annually increased dramatically from 2000 in the early 1970s to 10 000 in the 1980s. The total number of segregating populations (F2 to F7) grew from 20 000 lines to 150 000. Similarly, the number of entries in yield trials increased from 1 000 to 5000 annually. The total area in breeding and testing expanded from 30 ha to 100 ha in the same period. To accommodate this increase in breeding populations, the methodology of selection was changed from a pedigree system to a modified pedigree-bulk selection approach. The new method allows one experienced CIMMYT breeder to evaluate all segregating populations, in a timely fashion, except for the F2. Simultaneously, total mechanization of planting and harvesting and the computerization of field books have allowed a limited group of support staff and technicians to carry out all responsibilities. These three major changes introduced in the CIMMYT operation have increased the ability to introgress variability by significantly increasing the number of crosses directed for specific MEs, while keeping the selection program highly efficient and without sacrificing population size per cross.

PA PB X Cross cultivar A by cultivar B according to the

breading goals that have been previously identified

F1- Grow 50 to 100 plants. Back-cross or top cross if desired.

F2- Grow 2000 spaced plants. Individual plants are

selected based on agronomic type, disease

resistance, seed health and grain size.

F3 to F5- Selected F2 are grown at a commercial

seeding rate in 3 rows of 2m in order to

observe competitive ability within the line. Selection is based on agronomic

performance and disease resistance in

the plot. 10 to 15 heads are harvested

and bulked for each selected plot and

promoted to the next generation.

F4

F5

F6- Five to ten heads from selected plots are

threshed individually.

F7- Heads selected from F6 are planted individually in

head-rows. Selected rows are harvested in bulk.

F8 to F10- Selections continue based on preliminary

yield trials and subsequent replicated yield trials, along with industrial quality

testing.

F11- Superior lines are included in one of CIMMYT’s

international Nurseries or Yield Trials.

- 35 -

Fig. 12 Selected Bulk Method

The main difference between modified pedigree/bulk and the bulk selection method is how selected plants from the F2 populations are handled. In the modified pedigree-bulk selection method plants are kept separate in the F2; using the selected bulk method all seeds are bulked in the F2. The advantage of bulking the F2 is that it requires less land and much less bookkeeping. The advantage of keeping the F2 plants separate is that selection is more efficient at an early stage. However, computer simulations have shown that the genetic gain is similar for the two methods. A more detailed look at how CIMMYT conducts their Selected Bulk method will be found in the module: Managing a Breeding Program.

Selected bulk method

F1 F2 F3 F4 F5 F6 F7 F8 to F10

Parent A x Parent B

Bulk plot Space planted Bulk plot with selection Bulk plot with selection Bulk plot with selection Plant rows Preliminary yield trials

Replicated yield trials

- 36 -

Genetic advance is the expected gain in the mean of a population for a particular trait by one generation of selection. Genetic advance can be both predicted and calculated, and therefore can be used to both determine breeding objectives and track progress while selecting from populations. For predicting genetic advance the following formula can be used:

( ) ( )( )2s PG i V h= .

Where Gs is the predicted genetic advance, i is a constant based on selection intensity (table 1), VP is the phenotypic variance, and h2 is the narrow-sense heritability. In order to use this formula narrow-sense heritability must be known. For calculating genetic advance during the selection process the following formula can be used:

( )( )( )2p PR k V h= .

Where R is the response to selection, and kp is the standardized selection

differential p

P

Sk

V

=

. The selection differential is denoted by S and is equal to the

difference of the selected individuals mean minus the mean of the population: selected populationS µ µ= − . The wheat breeder will need to improve not only

one trait, but several traits at one time. This can be done by establishing minimum performance for each trait and excluding the others in the following generations. This is known as independent culling. Independent culling levels, along with amount of resources available to the breeder will determine the selection intensity. For example, 60% of the population might meet the height requirements, 30% might meet the disease resistance requirements, and 40% might meet the quality requirements. In this case, 60% 30% 40% 7.2%× × = of the current population will be advanced to the following selection cycle. If the breeder has 10,000 head rows and the resources to manage 1000 preliminary yield trial plots, these independent culling levels are appropriate. A more efficient form of multiple trait selection is by using a selection index with separate traits being given different weights depending on their level of importance. The Smith-Hazel index, or optimum index, is a good example of a selection index; however an understanding of covariance and matrix algebra is needed and cannot be covered here.

Table1 Commonly Practiced Selection Intensities and Their Corresponding Constants Selection Intensity (%) i

1 2.665 5 2.063 10 1.755 20 1.400

VI. Genotype X Environment (GxE) Interactions As stated previously, phenotypic variance can be attributed to both genetics and genetic by environment interaction. Genetically identical wheat populations grown in

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various environments will perform quite differently. Soil fertility, temperature, precipitation (amount and timing of), tillage practices, crop-rotation practices, day length, and disease pressure are all environmental variables among many others that can have an effect on wheat performance. Environmental variability can be accounted for by both diverse positions (locations, and replications) and time (years, or seasons). There are three approaches a plant breeder can take when addressing GxE. First, the breeder can ignore it. With this approach potential cultivars are tested over a wide range of environments. The performance of cultivars, resulting from selection, is superior when averaged over all the target environments; but is probably not the best in each environment. The second approach is to reduce GxE interaction. This can be done by partitioning the target environments into smaller more homogeneous subgroups. This can be done on a grand scale such as CIMMYT has done by defining the 12 Mega-Environments, or it could be done on a much region. Cultivar recommendations are then made for each distinct subgroup. Cluster analysis and principal component analysis are useful when partitioning environments. The third approach is to exploit GxE interaction (reduce the GxE variance component). This approach is used to identify cultivars best suited for a specific, niche environment in an effort to maximize productivity in that one environment. This third approach is a way to exploit positive GxE interaction and might be at first thought the best approach of all, until the amount of resources necessary to breed cultivars for every niche environment is considered. Stability analysis and multiplicative models can be useful tools when the third approach to addressing GXE is desired. As stated and implied previously the wheat breeder’s job is to select genotypes that will outperform the local check or standard cultivar. Also expressed earlier is that a pure line population’s performance is based not only on its genotype but also how the genotype interacts with the environment: P G GEV V V Vε= + + . In order to more accurately

select the superior genotypes the breeder must be able to explain a maximum of VP by VG. This can be done by increasing the number of environments that the genotypes are grown in and the number of replications in each environment. By increasing the number of environments a line is tested in; the unexplained variance VЄ, or the error variance, is reduced and more of the variance can be explained by VG and VGE. In turn an increase in variance explained by genotype will increase heritability estimates, leading to an increase

in selection response (remember: ( )( )( )2p PR k V h= ).

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Questions to Test Understanding: 1. In what way is plant breeding an art?

2. What are the advantages to the CIMMYT breeders of dividing the world

in Mega Environments (ME)?

3. What is a single backcross breeding strategy?

4. What is the purpose of the single-seed-descent method?

5. What is the definition of Genetic Advance?

6. What are the different approaches that a breeder can take to addressing

GxE?

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References: John Milton Poehlman and David Allen Sleper. Breeding Field Crops (4th Ed) 1995 Iowa State University Press, Ames Iowa E.G. Heyne editor. Wheat and Wheat Improvement. 2nd edition. American Society of Agronomy, Inc. Crop Science Society of America, Inc., Soil Science Society of America, Inc. Publishers Madison, Wisconsin USA 1987 Rex Bernardo. Breeding For Quantitative Traits In Plants. 2002 Stemma Press