Euphytica 100: 207–217, 1998.

207
 1998 Kluwer Academic Publishers. Printed in the Netherlands.

Golden calves or white elephants? Biotechnologies for wheat improvement

J.W. Snape
John Innes Centre, Norwich Research Park, Colney, Norwich, UK

Summary

The 1990s have seen an acceleration in the development of new biotechnologies which can increase the effi-
ciency of wheat breeding by providing new and novel sources of variation, speeding up the breeding cycle, and
increasing the efficiency of selection. This paper reviews the most significant technologies and their probable
impact on wheat breeding into the next millennium. Amongst techniques developed from the application of
tissue culture methods, doubled haploid systems are at last making a contribution through the development of
the maize pollination system. By the introduction of various improvements, this is now efficient enough to
produce material from a range of adapted genotypes in large numbers, and varieties are entering national list
trials from this system. Developments in tissue culture have also led to the realistic possibility of genetically
engineering wheat, based on biolistic methods of gene delivery into immature embryos. Some problems relat-
ing to gene stability and expression remain to be resolved, but targets, particularly with respect to disease and
pest resistance and end-use quality, are now being actively pursued. The development of the genetic wheat
map using molecular marker systems has revolutionalised the power of genetical analysis in wheat, enabling
agronomic trait loci, whether major genes or QTL, to be identified, located and ‘tagged’. Additionally, strate-
gies for the molecular cloning of loci are being developed, particularly by exploiting a comparative mapping
approach which combines the genetical information from all cereals in a common framework. This will lead to
tools for modifying crop phenotype in a directed fashion to produce improved and novel phenotypes.

Introduction have their major roots in ground-work laid over the

Scientific approaches to wheat breeding have their
history in the development of the pedigree system
and the rediscovery of Mendel’s laws at the begin-
ning of this century. Consistently since then, wheat
breeders have been amongst the most receptive to
scientific developments and their potential applica-
tion to crop improvement. There has been a con-
stant search for technologies that can convert plant
breeding from an ‘art’ to a ‘science’ – a search stim-
ulated by the need to increase food production, the
necessity of counteracting the evolution of pests
and diseases, the introduction of changes in agron-
omic practise, environmental perturbations, and in
the developed world, the commercial aspirations of
competitors! Most of the current developments in
biotechnology applied to wheat breeding however,
last 40 years through developments in, and the evo-
lution of, technologies for cytogenetic manipula-
tion, tissue culture, genetical analysis and more re-
cently, molecular biology. This paper discusses
some of these developments and their potential to
contribute to scientific approaches to wheat im-
provement in the 1990s and into the next millenni-
um.
Historical developments in wheat biotechnology
During the 1950s and 1960s, the use of new tech-
niques for modifying crop phenotypes were intro-
duced into wheat breeding. Developments in radi-
ation biology encouraged the application of muta-
tion techniques. Gamma irradiation and chemical

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208
mutagens were applied with some success, to devel-
op new morphological mutants, particularly, e.g.,
dwarfing genes (Maluszynski, 1990). These metho-
dologies continue to be used and are becoming
more widely used as developments in molecular
biology require mutants for techniques of gene iso-
lation. The work of Sears (1954) and Riley and col-
leagues, led to cytological methodologies for the
transfer of genes from wild species through a more
detailed cytogenetic understanding of the wheat ge-
nome and the refinement of embryo culture tech-
niques. The transfer of genes for disease resistance,
in particular (Riley et al., 1968), has been a great
success, is now widely practised, and continues to
make a major contribution to wheat breeding
worldwide (Gale & Miller, 1984). These innova-
tions can be classified as biotechnological applica-
tions, although perhaps not always recognised as
such. They have been refined even further in the
1990s, particularly by the application of molecular
cytogenetics and molecular marker techniques.
During the 1970s, the development of methodol-
ogies for the regeneration of plants from embryo-
genic callus from various explants, particularly im-
mature embryos, encouraged the use of tissue cul-
ture as a tool for plant breeders. The phenomenon
of somaclonal variation was recognised from these
investigations (Scowcroft & Larkin, 1981) and
raised the expectation of a new method of wheat
breeding. However, even though large resources in
many laboratories were devoted to generating vari-
ants and their assessment, very little agronomic ad-
vancement was achieved using this technique, and,
in wheat and other crops, it was less successful than
hoped. Indeed, it is now recognised that the tech-
nique is probably just another method of mutagen-
esis, with few unique features of the variants pro-
duced relative to other mutagen treatments. Never-
theless, an important feature was that it established
the basis of tissue culture protocols in wheat which
are now essential for methods of genetic engineer-
ing.
During the 1970s, tissue culture methodologies
for the regeneration of plants from gametocytes
were also developed, and their potential for pro-
ducing doubled haploid lines was recognised. Tech-
niques for anther culture were developed, particu-
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154800 EUPH Ordernummer 208284
larly by Chinese workers, and were successful in
producing new, better adapted varieties in a shorter
time scale (Hu & Yang, 1986). However, genotypic
difference in response frequencies limited the uni-
versal applicability of the technology. Alternative
systems were sought, and the recognition in barley
of the phenomenon of chromosome elimination in
the interspecific cross of barley with Hordeum bul-
bosum by Kasha & Kao (1970) promoted similar
studies in wheat (Barclay, 1975). Although the use
of H. bulbosum as a pollinator was restricted be-
cause of crossability problems (Snape et al., 1979),
the search for alternative pollinators proved a fruit-
ful line of investigation. This culminated in the
identification and use of the intergeneric cross of
wheat with maize (Laurie & Bennett, 1988), which
has greatly increased the potential for the wide-
spread adoption of doubled haploid systems in
wheat breeding programmes.
Genetic analysis has always made a contribution
to wheat breeding since the demonstration by Bif-
fin (1907) that yellow rust resistance in wheat was
inherited as a Mendelian character. In the 1960s and
1970s, both conventional and aneuploid methods of
analysis enabled the identification and establish-
ment of the chromosomal locations of many genes
controlling economically important traits (Law et
al., 1981; Worland et al., 1987). These techniques
were very successfully applied to major genes, and
the locations of genes controlling aspects of adapta-
tion (vernalisation requirement, photoperiod re-
sponse), dwarfing, spike morphology, grain and
glume colour, aspects of quality such as grain hard-
ness, storage proteins, and disease resistance, were
established.
The analysis of the genetical basis of quantitative
variation and the locations of specific genes (now
known as Quantitative Trait Loci, QTL) was more
problematic. There were few reports of detailed
analysis of individual genes, with perhaps a few ex-
ceptions such as the work of Law (1966, 1967) in lo-
cating several genes on chromosome 7B. However,
there were major problems in providing a compre-
hensive description of the complete genetical con-
trol of any particular trait. The major limitation was
that there was an insufficient number of morpho-
logical variants to enable the development of com-

prehensive genetic maps to provide landmarks for
the intrachromosomal locations of genes. This sit-
uation improved in the 1970s and 1980s with the de-
velopment of electrophoretic systems to visualise
differences in isozymes and grain storage proteins;
but again, the levels of polymorphism and the num-
bers of available marker loci were too low to devel-
op detailed maps. Only in the 1990s, due to the de-
velopment of molecular marker systems, has this
situation changed. It has now been revolutionised
by the availability of detailed genetic maps based
primarily on RFLP (restriction fragment length po-
lymorphism) markers. This is resulting in a dramat-
ic increase in the power of genetic analysis so that
the totality of the genetical variation for any trait in
a cross can be described and attributed to specific
locations in the genome. There are still methodo-
logical and statistical problems to be overcome with
this approach, but notwithstanding, it is now lead-
ing to a much more detailed understanding of ge-
netical variation for economically important traits.
Importantly, this information can be translated into
selection tools for plant breeders.
The contribution of doubled haploid systems to
wheat breeding
The production of a new wheat variety using the
normally practised pedigree system is still a time-
consuming process. It requires many generations of
self-pollination and selection to achieve the levels
of homozygosity and hence genotypic stability re-
quired for a variety to be released. One of the major
contributions of tissue culture techniques to wheat
breeding has been the development of doubled ha-
ploid (DH) systems which can short-circuit this pro-
cess. Generally, a minimum of two years can be
saved in the release of a new cultivar by the devel-
opment of recombinant DH populations from in-
ter-varietal F1s. An added and significant advantage
is that these systems not only speed up the advance
to homozygosity, but can also increase selection ef-
ficiency (Snape, 1989). This is obtained because of
the greater proportion of additive genetic variation
available for selection for quantitative traits, and
the absence of dominance effects for major genes.
Please indicate author’s corrections in blue, setting errors in red
154800 EUPH Ordernummer 208284
209
This allows better discrimination between geno-
types within crosses, better discrimination between
crosses, and greater selection response across gen-
erations (Snape, 1982).
The widespread use of DH technology in wheat
has been impeded by the lack of a technique which
can satisfy all of the expected criteria for a success-
ful system (Snape at al., 1986), namely: 1) easy, con-
sistent production of large numbers of DHs of all
genotypes in the breeding programme, 2) DHs
should be genetically normal and phenotypically
stable, and 3) recombinant DH populations should
contain an adequate sample of the genetical varia-
tion in the parents. The discovery by Laurie & Ben-
nett (1988) that chromosome elimination could be
exploited by using the intergeneric cross with maize
pollen, has at last led to a commercially exploitable,
genotype-independent system. This system is now
enabling large populations of homozygous recom-
binant lines to be produced at reasonable cost. This
has been possible by the modification of a number
of features of the original published technique.
Firstly, crossing techniques have been improved to
allow large scale emasculation and high fertilisation
frequencies, such as by the use of chemical hybrid-
isation agents, with improved pollinators. Although
the original maize sweetcorn hybrid used by Laurie
& Bennett (1989), Seneca 60, is still amongst the
most successful, other commercial F1 sweetcorn hy-
brids, better adapted to local conditions, are also
very successful pollinators (Snape, Ellerbrook &
Fish, unpublished). Other pollinators such as teo-
sinte, sorghum, millet and Tripsacum have also
proved highly successful. Secondly, there have been
improvements in the post-pollination application of
plant hormones which allow high frequency em-
bryo survival and good quality embryos with a high
frequency of germination to be formed. Typically,
2-4 D is used, but other chemicals have also been
used successfully, and cocktails of auxins have
proved equally, if not more, successful. Thirdly, im-
proved media have been developed, mainly arising
from developments in tissue culture techniques for
transformation systems, which ensure the success-
ful development of healthy haploid seedlings. Fi-
nally, chromosome doubling techniques have been
refined to ensure high survival yet efficient dou-
210
bling. Colchicine is still the most widely used chemi-
cal because of its efficiency. However a safer alter-
native is required, and ongoing experiments with al-
ternatives such as the herbicide Oryzalin look
promising.
Several commercial wheat breeding companies
worldwide are exploiting these developments and
very efficient methods have emerged in-house. In
Europe, DH lines produced from this system are
now entering national list trials. For example, Table
1 shows typical frequencies from work at the John
Innes Centre and unpublished results from a lead-
ing UK plant breeding company. Clearly, a frequen-
cy of over four doubled haploid plants per spike
pollinated can be achieved with adapted, commer-
cial varieties, opening up the possibilities for the
production of several thousand lines/year for
agronomic assessment. The system is also applica-
ble to durum wheats (O’Donoughue & Bennett,
1994), and although success frequencies at present
appear lower than with bread wheat (Savaskan et
al., 1996), it is the best technique currently available
for this species.
The maize system is now being widely practised,
but nevertheless, it has the disadvantage of being
more expensive than conventional breeding tech-
niques and only a limited number of crosses can
usually be handled. There is a need to move the
technologies on further. Current research in cereal
microspore culture may provide the next break-
through in DH technologies. Several thousand mi-
crospores can be obtained from an individual spike,
and, if these can be induced to undergo sporophytic
development, then the gate will be open for the
large scale production of DH lines with low inputs.
Table 1. Doubled haploid production frequencies in wheat
Parental No. of spikes
material pollinated
Portuguese wheat 64
F1s (JIC)
(average 4 crosses) % florets pollinated
Commercial UK 185
winter wheat F3s
(average 8 crosses) % florets pollinated
Please indicate author’s corrections in blue, setting errors in red
154800 EUPH Ordernummer 208284
Hence, many more crosses could be handled than at
present with the maize system. Studies in barley mi-
crospore culture suggest that this is possible (Har-
wood et al., 1995). Future developments in tissue
culture procedures should give further progress,
and if such successful microspore culture tech-
niques can be transferred to wheat, this technique
has great potential for producing large numbers of
doubled haploids with low technical inputs.
Exploiting transformation technologies in wheat
Recent developments in tissue culture and transfor-
mation technologies are finally making the genetic
engineering of wheat for agronomic traits a possi-
bility (Jahne et al., 1995). Although a number of dif-
ferent techniques have been attempted over the
years, including protoplast electroporation and pol-
len tube microinjection, undoubtedly the most suc-
cessful approach at present is via biolistic methods
of gene delivery into proliferating scutellum tissue
of immature embryos (Weeks et al., 1993). This
technique is now established in many laboratories,
particularly in industry. This system relies on the in-
troduction of a gene of interest, either on the same
plasmid as a selectable marker and reporter gene,
or on a separate plasmid in co-transformation ex-
periments. The selectable marker which is proving
to be the most reliable is the Bar gene from the bac-
terium Streptomyces, conferring resistance to the
herbicide bialophos, under the control of the maize
ubiquitin promoter. The Gus gene is still the most
widely used reporter of transient and stable trans-
formation in wheat, although the use of the firefly
No. of florets Embryos Seedling Success
pollinated rescued survival (DH
(per spike) (per spike) plants-spike)
1188 (17) 284 (4) 88 1.4
23.7 7.4
6696 (36) 2249 (12) 753 4.1
33.6 11.2

luciferase gene and the jellyfish green fluorescent
protein are being tested as non-destructive reporter
systems. Several different ‘guns’ are available, in-
cluding gunpowder driven devices, but the most
widely used is the BioRad PDS1000/Helium gun.
Particle inflow devices (Finer et al., 1992) are also
proving successful in achieving stable transforma-
tion.
The success of these systems is now turning atten-
tion to what can be achieved by the modification of
wheat for agronomic traits. There are two ap-
proaches to the genetic engineering of wheat. The
first is to use the technologies to modify and reinsert
native genes back into wheat. This can be done
either to increase the expression of a native gene,
modify its product, or to switch a gene off using anti-
sense technologies. With respect to altering expres-
sion levels, probably the most intensively investi-
gated example at the present time is the modifica-
tion of wheat storage proteins, particularly the
high-molecular-weight glutenins. For example, the
Dy10 and Dx5 subunits encoded by chromosome
5D have been isolated from the variety Cheyenne,
since these are known to impart good bread-mak-
ing quality (Shewry et al., 1989). These are being
used to study functionality by being re-inserted into
wheat under the control of native promoters to look
at their individual effects on bread-making quality
(Blechl & Anderson, personal communication).
Targets for antisense technology are, e.g., genes that
promote susceptibility to disease. Worland & Law
(1991) have identified one such target. From studies
of wheat aneuploids, they have shown that reducing
the dosage of chromosome 5D, as in the monosom-
ic, of the susceptible variety Hobbit ‘sib’, increases
adult plant resistance to yellow rust and also to mil-
dew. Thus, if the particular gene involved could be
isolated, an antisense construct could be inserted in
this or other varieties to increase resistance. As our
knowledge of disease resistance mechanisms in-
creases, other targets for modifying or switching off
the product, or increasing the expression of resist-
ance genes will be identified.
The second use of transformation technologies
will be to generate novel germplasm through intro-
ducing genes from other biological sources into
wheat, be it viral, bacterial, plant or even animal in
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154800 EUPH Ordernummer 208284
211
origin. Primary targets will be the introduction of
novel genes to alleviate pest, disease and stress
problems, or to create novel products for new end-
uses, such as industrial raw materials. Table 2, for
example, lists current targets being sought by differ-
ent groups, particularly in commercial breeding
companies. The success of these approaches will
obviously depend on the availability of cloned
genes of interest, and different sources of plant
genes are becoming available for both crop and
model plant species, particularly Arabidopsis. For
example, sources of fungal resistance are now
emerging from studies of resistance genes such as
the Cf2 and Cf9 genes of tomato (Dixon et al., 1996;
Jones et al., 1994). The sequences of these genes
have been determined, and degenerate PCR prim-
ers can be designed to pull out homologues from
other species including wheat. This offers the op-
portunity to take resistance genes from other Trit-
iceae species, particularly wild species, and intro-
duce them directly into wheat, obviating the need
for cytogenetical manipulation. Other genes are be-
ing introduced from bacteria, for example for herbi-
cide resistance. Additionally, novel sources of genes
for modifying end-use come from studies in other
crop species, e.g., peas. In peas, several genes have
been identified that control starch synthesis and
Table 2. Realistic targets presently available for the transforma-
tion of wheat crops
Yield Pest resistance: B.thuringensis genes
limiting Anti-metabolism proteins
factors Protease inhibitors
Fungal resistance: Arabidopsis/tomato/rice
homologues of resistance genes
Anti-toxin genes
Virus resistance: Coat protein genes
Replicase antisense
Anti-viral proteins
Herbicide resistance: Glyphosate tolerance
Quality Bread-making: Wheat glutenin genes
characters quality Pea lipoxygenases
Starch composition: Starch branching enzyme
Novel Pharmaceuticals
products Novel starches
Pathogen antibodies
Others Male sterility
Female sterility
212
biochemistry including the gene controlling Men-
del’s ‘wrinkled’ gene, which, in fact, codes for a
starch branching enzyme (Smith & Martin, 1992).
This and other genes can be used to modify the
starch composition of wheat which could generate
products for industrial uses such as biodegradable
plastics. Pea lipoxygenases have also been cloned
and could be reinserted into wheat to increase
dough rheology and ameliorate the need to add soy-
bean flour during the baking process.
Clearly there are many opportunities opening for
the genetic modification of wheat, although it
should not be forgotten that there are still technical
limitations to what can be achieved at present. For
example, the mechanisms of transgene insertion
and gene expression are not understood, and gene
expression can be highly variable between different
transgenics developed using the same construct, un-
der the same conditions. Additionally, the phenom-
ena of transgene silencing and unstable inheritance
patterns can occur for reasons still not understood.
Thus, there are challenges still remaining in terms
of understanding gene expression, stability and dur-
ability. Additionally, farmer acceptance and con-
sumer concerns will need to be taken into account,
and these techniques are unlikely to be a universal
panacea. At present, they should be regarded as
complementary to conventional breeding technol-
ogies, and used in an integrated approach to crop
improvement.
Biotechnology and genetical analysis
Genetical analysis, combined with character analy-
sis, plays a pivotal role in providing plant breeders
with information about the characters and genes
that they wish to manipulate to produce high yield-
ing varieties with better adaptability to appropriate
environments, better disease, pest and stress toler-
ance, and good quality. Current estimates are that
wheat probably contains between 25-30,000 unique
genes, but only a fraction of these have been
mapped so that their primary and pleiotropic effect
can be studied, understood and manipulated. There
is an urgent need to use genetical analysis to identi-
fy, locate and then gene tag agronomically impor-
Please indicate author’s corrections in blue, setting errors in red
154800 EUPH Ordernummer 208284
tant loci, so that the available variation can be ma-
nipulated in a more directed manner than has been
hitherto possible. This would enable yield potential
to be maximised in a given environment. There is
evidence in the United Kingdom, for example, that
dramatic increases in yields of varieties over the last
30 years can be traced, at least in part, to the intro-
duction of a few major effects such as the introduc-
tion of the Rht1 and Rht2 dwarfing genes and the
1Bl/1RS translocation (Angus, Nickerson Plant
Breeders, personal communication). The identifi-
cation of other new and novel effects and their as-
sembly into adapted backgrounds can lead to
greater yields and yield stability.
Biotechnological advances in DNA marker tech-
nologies are now enabling detailed genetic analysis
of all traits by associating allelic variation at marker
loci with phenotypic variation for traits of interest.
Comprehensive genetic maps of the whole wheat
genome have now emerged (Devos & Gale, 1993),
so that precise methods of genetic analysis, en-
abling the accurate location and manipulation of
major genes, and QTL controlling important agron-
omic traits, are possible. In wheat, this can be car-
ried out at two levels of complexity. The first is at
the whole genome level, when partial or complete
genome marker coverage can be achieved (Hyne et
al., 1994) using recombinant doubled haploid or re-
combinant inbred populations. This is being prac-
tised in many other species, e.g., in barley (Klein-
hofs et al., 1993; Laurie et al., 1995). Secondly, and
specifically for wheat, it can be carried out at the
individual chromosome level since it is possible to
combine previously developed, sophisticated, chro-
mosome assay procedures from using cytogenetical
approaches (Law et al., 1981; 1987) with the newly
developed detailed genetic maps. By combining the
current molecular-marker-derived maps with re-
combinant substitution lines, wheat geneticists
have tools for genetical analysis which undoubtedly
meet or surpass any available in any other arable
crop species (Snape et al., 1994).
In particular crosses of interest, it is theoretically
possible to dissect the total phenotypic variation for
any trait into components attributable to each indi-
vidual gene. This then allows the possibility for indi-
vidual molecular marker ‘tags’ for genes of interest
 213

which can then be manipulated singly or pyramided
together in breeding programmes. In practice, it is
unlikely that all loci will be detected and located,
since it is probable that individual genes will vary in
the magnitudes of their effects, and only those that
reach a threshold level of effect greater than the ex-
perimental error will be detectable. Nevertheless, it
should be possible to tie down a considerable pro-
portion of the variation for any one trait, particular-
ly if the variation is mediated by a few loci of rela-
tively large effect. Indeed, this type of inheritance
has been shown to be the case for many of the traits
of interest to plant breeders in wheat such as flower-
ing time, height, yield and yield components, stress
responses, and quality characteristics. Thus, an effi-
cient tagging strategy could be to target only a small
number of major genes or QTL of large effect,and
to ignore other loci of small effect.
An example of the way in which genetic analysis
is contributing to directed wheat breeding can be
illustrated by recent studies on the adaptation of
wheat to specific eco-geographical environments
through the genetic control of flowering time and
cold tolerance (Worland, 1996; Galiba et al., 1995).
Cold tolerance is a complex character in wheat, in-
fluenced by a number of factors. However, chromo-
some substitution line analysis has implicated chro-
mosomes of homoeologous group 5 as carrying im-
portant determinants. The genetic control of flo-
wering time is also complex, and chromosome sub-
stitution line analysis has shown that chromosomes
of nearly all homoeologous groups are involved
(Table 3). Indeed, the existence of extensive allelic
variation for these genes make bread wheat (Trit-
icum aestivum L.) the most widely adapted major
cereal crop in the world. Broadly speaking, three
separate sets of genes are involved in the genetic
control of flowering in wheat. First are the genes
controlling sensitivity to vernalisation, which deter-
mine the control of the spring wheat/winter wheat
difference, the Vrn genes. The second major group
of genes are those controlling response to photope-
riod/daylength (Welsh et al., 1973; Law et al., 1978;
Scarth & Law, 1983). These play an important part
in accelerating or delaying flowering time in au-
tumn sown wheats in the spring after vernalisation
requirement has been satisfied. The third group of
loci involved in flowering time are termed ‘earliness
per se’ or ‘developmental rate’ genes. These are
genes that do not respond differentially to different
lengths of cold treatment or photoperiod, and seem
to be distributed throughout the genome. In genet-
ical analysis terms these ‘eps’ loci are generally lo-
cated as QTL effects rather than as major genes.
Historically, most of the genetical studies of flo-
wering time and cold tolerance could not verify the
precise intrachromosomal locations of these major
genes and QTL (Sutka & Snape, 1989). It is only

Table 3. Genetic analysis of flowering time in wheat : whole chro- Table 4. Locations of genes controlling flowering time and frost
mosome effects. tolerance in wheat and barley

Homoeologous group Genes located A genome B genome D genome H genome

Group 1 Genes for sensitivity to Group 1 Ppd-H2

vernalisation Group 2 Ppd 3 Ppd2 Ppd1 Ppd-H1
Group 2 Genes for sensitivity to Eps-2BS Eps-2DS Eps-2HS
photoperiod Group 3 Denso
Earliness per se genes Eps-3HL
Group 3 Earliness per se genes Group 4 Sh
Group 4 Eps-4HL
Group 5 Genes for sensitivity to Group 5 Vrn1 Vrn3 Sh2
vernalisation Eps-5HL
Group 6 Genes for sensitivity to Fr1 Fr2? Fr-H1
vernalisation Group 6 Eps-6HL.1
Group 7 Genes for sensitivity to Eps-6HL.2
vernalisation Group 7 Vrn5 Eps-7HS
Eps-7HL
Source: Law et al., 1991.

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214
with the advent of molecular marker systems that
precise locations have been established for these
major effects, and the loci can now be targeted for
genetic manipulation as for other major genes.
From a detailed genetic analysis in barley, (Laurie
et al., 1995), most chromosomes appear to carry eps
genes, and by homoeology, these would be expect-
ed to be present in wheat, although detailed map-
ping in wheat is currently restricted to the eps-2BS
locus on chromosome 2B (Scarth & Law, 1983). Ge-
netic analysis using RFLP maps, is ongoing to map
these loci in detail. Table 4 illustrates the current
status of the genetic analysis of flowering time loci
in barley and wheat. Genetic analysis also indicates
that the loci for these three genetic systems appear
to have multiple alleles (Snape et al., 1976). This in-
dicates that there must be enormous potential to
adjust and fine-tune the flowering time of wheat to
particular geographical regions and specific envi-
ronments within these. Studies also show that genes
of all three systems have pleiotropic effects on other
aspects of plant growth and development which has
important consequences for wheat breeding for
specific adaptation.
The types of genetical analyses described above
are at last revealing detailed information about the
genetical control of adaptation in wheat and the in-
fluence of specific genes on adaptation and yield
performance in different environments. Similar in-
formation is emerging on the genetical control of
pest and disease resistance, including adult and
non-specific forms of resistance and the many as-
pects of grain and end-use quality. The challenge
now is to translate this genetical information into
selection tools that can be used by the wheat breed-
ers on an efficient and low cost basis.
The potential of comparative mapping
The great advantage of molecular markers over
conventional markers is that many DNA probes hy-
bridise across crosses within the same species,
across genomes within polyploid species such as
wheat, and frequently across independent genomes
of taxonomically distant but related species. Com-
parative genetic analysis is an additional tool avail-
Please indicate author’s corrections in blue, setting errors in red
154800 EUPH Ordernummer 208284
able to the wheat geneticist towards gene location,
which also provides new insights into gene action,
and gives breeders access to a wider spectrum of
genes (Snape et al., 1995; 1996). The genetics of
wheat can now be clearly linked to the genetics of
other Triticeae species, particularly barley and rye,
since extensive collinearity has been shown be-
tween the wheat, barley and rye genetic maps, and
common markers are being used to combine the
available information on important agronomic
characters into a common framework. Since this
synteny now extends much further to maize, rice,
sorghum, millet and forage grasses (Moore et al.,
1995), it should be possible to carry out comparative
major gene and QTL analysis across all these spe-
cies. This can be used to link known genes into ho-
moeologous series or to search for previously un-
described genes. For example, we have exploited
comparative mapping to show that Vrn1, control-
ling vernalisation response, and Fr1, controlling
frost tolerance on the short arm of 5A in wheat, are
homoeologous to Sh2, the major vernalisation gene
on chromosome 7 in barley; also a QTL for frost
resistance, identified by Hayes et al. (1993). The rye
Sp1 gene for vernalisation response on chromo-
some 5R must also be homoeologous to Vrn1 and
Sh2 (Plaschke et al., 1995; Galiba et al., 1995; Laurie
et al., 1995). An exciting development is that this
analysis can be extended to rice and maize, where
homologous genes are expected on chromosomes 3
in rice and chromosomes 1 and 5 in the duplicated
maize genome (Figure 1), even though these species
do not have a vernalisation requirement. Do such
homoeologous genes even exist in these species?, If
so, what is their function – do they affect flowering
Figure 1. Homoeologous locations of genes controlling vernal-
isation requirement and frost tolerance in wheat and barley, and
likely locations of homologues in rice and maize.

time? This possibility can now be tested by compar-
ative mapping in rice populations segregating for
heading date. Additionally, such mapping offers the
possibility of using the excellent genetics and mole-
cular tools of rice (molecular maps, yeast artificial
chromosomes, bacterial artificial chromosomes, ex-
pressed sequence tags, cDNA characterisation) as a
route for gene isolation in wheat and barley (Moore
et al., 1995). These can also link to the major ge-
nome projects on Arabidopsis. These approaches
are currently being developed and tested.
Discussion
The biotechnologies now available to the wheat ge-
neticist allow the opportunity to elucidate and mod-
ify the genetical architectures of most characters in
terms of the numbers of loci involved, their relative
magnitudes, their dominance and epistatic relation-
ship, and their primary and pleiotropic effects. Us-
ing the information on the genetical control of a
particular character for directed genetic manipula-
tion at the plant breeding level is still, however,
problematical. To do so requires that each locus and
each ‘desirable’ allele can be followed with a unique
‘gene handle’. With respect to RFLP markers, this
requires that a unique band profile is associated
with each particular allele of the agronomic trait
that is to be selected, that the marker is very closely
linked to the trait locus, and that there is linkage
disequilibrium for both loci in different crosses of
the gene pool being used. However, because of the
low levels of RFLP in wheat, this is presently prov-
ing to be difficult and very few loci have been
tagged to date. Additionally, RFLP analysis is ex-
pensive and is unlikely to be routinely usable on the
large populations that plant breeders handle. Thus,
considerable technology development is still re-
quired to make the process ‘breeder friendly’. This
can involve using the RFLP probes as entry points
to clone regions of the genome as near as possible to
the locus/allele being targeted, then sequencing the
region, and then designing specific PCR primers to
target the particular allele in question. Such primers
may then be suitable for use in dot-blot or PCR as-
says of breeding populations. Alternatively, the de-
Please indicate author’s corrections in blue, setting errors in red
154800 EUPH Ordernummer 208284
215
velopment of comprehensive microsatellite-based
genetic maps may provide the tools for marker-me-
diated selection.
Genetic analysis is also the first requirement for
cloning economically important genes at the mole-
cular level, since in wheat, a major generic method
for gene isolation is likely to be via chromosome
walking. Once genes can be isolated it will be pos-
sible to develop a much greater understanding of
gene structure and function, and to relate this to
plant phenotype. This, in turn, will then lead to ge-
netic manipulation at the molecular level to pro-
duce new alleles and allelic combinations. It will al-
so be possible to reintroduce these into wheat,
either as native genes or in a modified form, using
transformation technologies, or indeed they could
be used for the genetic manipulation of other cereal
and non-cereal species. Clearly, the door is now
open to carry out genetic analysis and genetic ma-
nipulation in wheat with a sophistication that has
previously been possible in only model plant spe-
cies.
Overall, new biotechnologies are at last making a
major impact on wheat breeding, and the pace
should accelerate into the next millennium. How-
ever, it must be kept in mind that these methodol-
ogies are complementary to, not exclusive of, con-
ventional wheat breeding techniques, which will
continue to be the mainstay of wheat improvement
for the foreseeable future.
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