Session
2 Breeding Methodologies i
Mapping
Oral Presentation
Experiences with Marker-Assisted Selection for
Quantitative Traits in Barley
S.
E. Ullrich1, A. Kleinhofs1,
1Department
of Crop and Soil Sciences, Washington State University, Pullman, WA 99164-6420,
USA,
E-mail: ullrich@wsu.edu; 2Centre UdL-IRTA, 25198
Molecular
marker-assisted selection (MMAS) is a proven breeding and genetics strategy for
simply inherited (13 gene) traits. MMAS for complexly inherited true
quantitative traits is less certain. Research through the North American Barley
Genome Project concentrated on malting quality and grain yield traits. These
traits are difficult to evaluate and select for due to number of genes,
genotype (G), environment (E), GXG, and GXE effects, and cost. For malting
quality, major QTL regions on chromosomes 1 (7H) (QTL1) and 4H (QTL2) with
multi-trait QTLs (malt extract, alpha-amylase, diastatic power, beta-glucan)
were MMAS targets in Steptoe/Morex (S/M) crosses. MMAS for QTL1 was effective
but not for QTL2. Fine mapping of QTL2 improved its selectability. For yield in
S/M, MMAS for QTLs on chromosomes 3H and 6H was more effective than for QTLs on
chromosomes 2H and 7 (5H). Another S/M MMAS study resulted in lines with
improved yield-related traits but not yield per se.
MMAS resulted in improved yield and malting quality in Harrington/Baronesse
backcross lines. Lessons learned include: (1) genotypic and/or tandem
genotypic-phenotypic selection was as good as or better than phenotypic
selection, (2) response of single QTLs can be complicated by epistasis and
cross-over GXE, (3) optimum QTL allele composition is difficult to predict, (4)
QTL fine mapping can improve MMAS.
A
Large Scale Mapping of ESTs on Barley Genome
K.
Sato1,
1Research
Institute for Bioresources, Okayama University,
E-mail: kazsato@rib.okayama-u.ac.jp; 2CREST,
ESTs are the most informative
sources of genetic markers on the linkage map in barley. We have generated ca.
60,000 ESTs from 3ends from nine different cDNA libraries. Each EST sequence
was basecalled by phred and trimmed by QV = 20. Unigenes were developed by
phrap (contig 8,753, singlet 6,686) to process primers by Primer3. Ca. 11,000
primer sets were generated. Our strategy of mapping ESTs are (1) polymorphisms
appear on agarose gel (2) SNP analysis after direct sequencing of parental PCR
amplicons either using PCR-RFLP or SNP typing system. The project aims to
localize several thousands of ESTs on the barley linkage map.
Genetic
Diversity of Barley Cultivars from
S.
1Institute
of Plant Physiology, Genetics & Bioengineering, Almaty, 480090, Kazakhstan;
2Department of Agroenvironmental Sciences and Technology,
40127-Bologna, Italy;
3enter for Crop Science and Farming, Almalybak, 483133,
Kazakhstan,
E-mail: research@nursat.kz;
absaule@yahoo.com
Hordeins, isozymes, RAPD, SSR, and AFLP markers were
used for studying the genetic diversity in 34 barley cultivars widely
grown in
Differences
between South American H Haplome Diploids
and I Haplome Diploids, from the Perspective of the 5S rDNA
Gene in the Genus Hordeum
B.
R. Baum1 and D. A. Johnson2
1Agriculture
and Agri-Food Canada, Ottawa, Ontario, ON K1A 0C6, Canada, E-mail: baumbr@agr.gc.ca;
2Ottawa-Carleton
Twelve South American diploid Hordeum species
belonging to the H genome and three diploid species belonging to the I genome
(including cultivated barley), were investigated for their 5S rDNA sequence
diversity. The ca. 500 sequenced clones were assigned to classes which were
further assigned to haplomes. Two classes were found to be present in each
haplome. These were labelled accordingly long H1 and short I1 for the I haplome
diploids, and long H2 and long Y2 for the South American diploids. The aligned
sequences were subjected to a series of Maximum likelihood analyses and various
tests, including molecular clock, which will be presented and discussed.
Use of the Functional Map to Identify QTLs
and Explore the Genetics of Biometric Agronomic
Traits
in the
G.
H. Buck-Sorlin1,2, R. K. Varshney3,
M. Prasad3, R. Kota3, A. Graner3
and A. Boerner3
1Department
of Cytogenetics and 3Genebank Department, Institute of Plant Genetics and
Crop
Plant Research (IPK), D-06466 Gatersleben, Germany,
E-mail: buck@ipk-gatersleben.de;
2Department of Computer Science, Brandenburg University of
Technology Cottbus (BTU),
D-03013 Cottbus, Germany
The
Oregon Wolfe Barleys (OWBs) are a well-characterised, phenotypically
polymorphic mapping population that has been used for a whole range of mapping
and QTL studies. The population was grown in the greenhouse twice in 1999/2000
and 2003, and biometric (organ lengths and ratios), meristic (number of leaves
per main stem, number of tillers) and phenological traits (flowering time) were
established in a systematic fashion. Additionally, the mapping population was
used to prepare a functional map by employing ESTs in different marker assays
like RFLPs, SSRs and SNPs. The functional map (or transcript map) based on the
OWB population contains > 500 genes. This map has been used for QTL analysis
for the above traits. The combination of the functional map with information on
biometric morphological traits promises to give new insights into the nature
and quantitative inheritance of these traits and is hoped to provide a decisive
progress in the field of expression mapping.
A
Consensus Molecular Genetic Map of Barley
M.
Cakir1, R. Appels1,2,
A. Hunter3, D. Schibeci3,
C. D. Li2 and M. Bellgard3
1Molecular
Plant Breeding CRC and WA State Agricultural Biotechnology Centre, Murdoch
University, Murdoch, WA 6150, Australia, E-mail: mcakir@central.murdoch.edu.au;
2Crop Improvement Institute, Department of Agriculture,
Bentley Delivery Centre, WA 6983, Australia; 3Centre
for Bioinformatics
and Biological Computing, Murdoch University,
Murdoch, WA 6150, Australia
An
extensive international collaboration allowed us to compile a consensus map of
barley. The consensus map was formed using four barley maps produced in our
laboratory as well as a number of other Australian and international published
genetic maps and includes over 2000 markers. c-MAP software was used to view
the consensus map and validate marker order by comparison of the consensus map
to the individual maps that contributed to the consensus map. The options in
c-MAP such as the matrix were found useful for quickly assessing the occurrence
of duplicate loci. The alignment of different genetic maps was generally
unambiguous with respect to the order of loci and examples of alignments will
be presented together with an estimate of the error inherent in producing the
consensus map. QTLs were also inserted into map in a searchable format within
the software and this has enhanced the value of the map considerably. This map
serves the purpose of summarising extensive datasets in the area of molecular
genetic mapping as well as in the application of molecular markers in plant
breeding programs.
Characterization
and Mapping of a Wild Barley eibi1 Mutation Identifying a Gene Essential for Leaf Water
Conservation
G.
Chen1, M. Sagi2,
1Institute
of Evolution, University of Haifa, Haifa 31905, Israel,
E-mail: nevo@research.haifa.ac.il;
2Institute for Applied Research,
A
spontaneous wilty mutant (eibi1) hypersensitive to drought was identified in
wild barley, Hordeum
spontaneum Koch. Mutant
eibi1 had a highest relative water loss rate among the
known wilty mutants, indicating that eibi1 is one of the most drought sensitive mutants.
When compared with wild type, eibi1
had the same
Molecular
Marker Validation and Physiological Determinants
of QTL Effect on Grain Protein Concentration of
Two-Rowed Barley
L.
C. Emebiri and D. B. Moody
Department of Primary Industries, Victorian Institute
for Dryland Agriculture, Horsham Victoria 3401,
With the
advent of DNA markers, it has become possible to locate quantitative trait loci
(QTL) for numerous phenotypes in plants. Beyond QTL identification, however, a
number of issues still need to be addressed in order to integrate quantitative
genetic information into genome-based breeding programs. Among these include
validation of QTL effects and a determination of the possible physiological
basis of gene action. In previous studies, QTLs that influenced variations in
barley grain protein concentration (GPC) were identified on chromosome 5H and
7H. The objectives of the present studies were
(1) to verify marker-trait
associations across genetic backgrounds, and (2) to establish the morphological
and physiological basis for differences in GPC among doubled haploid lines
selected for allelic variation at identified QTLs. In the first study, QTL
effects were confirmed using 3 independent doubled haploid populations with
different combinations of low and high GPC parental lines. In the second study,
doubled haploid lines with allelic variation at these QTL were grown at
Horsham, Victoria, in 2001 and 2002, at 0 and 80 kg/ha of nitrogen application,
and assessed for phenotypic differences. On average, lines with low-protein
alleles were lower in GPC by 1.2% units. There were no significant differences
in tillering or dry matter accumulation during vegetative growth, and at
anthesis, there were also no significant differences in nitrogen content of the
straw or spikes. At maturity, however, highly significant (P
0.01) differences were observed for traits associated with spike morphology
(spike weight, spike length and kernels per spike), indicating that lower GPC
was achieved by the distribution of a similar amount of nitrogen to a larger
number of grains. The implications on germplasm utilisation were discussed.
Identifying
Genes Controlling Heading Date in Spring Barley
J.
D. Franckowiak, N. N. Krasheninnik and
G. T. Yu
Department of Plant Sciences,
E-mail: j.franckowiak@ndsu.nodak.edu
Heading
date determines in part the adaptation of barley to specific production areas.
Most barley breeding programs use some exotic germplasm; thus, knowledge of
expected photoperiod responses is desirable. Photoperiod sensitive genes in
barley are commonly called early maturity (Eam)
genes. Heading date is also affected by temperature and vernalization
requirements (the Sgh1, Sgh2, and sgh3 genes for spring growth habit). The Eam1
gene, which is expressed only under long-day conditions (13 hours or longer),
is frequently present in wild barley, winter cultivars, and spring cultivars
grown during the winter or at high latitudes. Many two-rowed spring cultivars
have the long-day gene Eam11
(effective at 12 to 13 hours only).
The Eam6 gene confers early heading under both long- and
short-day conditions. The Eam5 and eam9 genes confer earliness under short-day conditions.
The eam8 gene shows a day length-neutral response. The eam10,
mat-f, and mat-i genes produce early plants in most tests. The
long-day genes Eam1, Eam6, and Eam11 can cause breeding problems because they are located
in chromosome 2H near QTLs for resistance to Fusarium head blight. The Eam6
and Eam11 loci are near the six-rowed spike type 1 (vrs1)
locus. The Eam6 gene exhibits additive interactions with Eam1,
Eam5, and eam9. Three or more unidentified genes can enhance or
inhibit the expression of some, but not all, Eam genes.
Development
of Functional Genetic Markers by Combination of cDNA-AFLP Based Expression
Profiling and Marker
Assisted Genotype Pooling
M.
Herz1,
1Research
Group of Genome Analysis and 2Research Group of Barley Breeding ,
Institute for Crop Production and
Plant Breeding, Bavarian State Research Centre for Agriculture,
D-85354 Freising-Weihenstephan, Germany,
E-mail: markus.herz@lfl.bayern.de
Malting quality of barley represents
the manifestation of the well-adjusted interaction of several different genes.
Due to this complex genetic basis, quantitatively inherited traits and in
particular malting quality require sophisticated methods to be tagged by
genetic markers. Induction of gene expression by a standardised micromalting
process was used to identify differentially expressed genes both in varieties
and in segregating populations. Based on a DH-population which was previously
used to construct a QTL map for malting quality, phenotypic pools were
constructed utilising the information about markers which flank significant QTL
intervals for malting quality traits and the observations of the traits. cDNA-
AFLP analysis was performed with a selected subpopulation of the segregating
progeny and differential TDFs were integrated into the linkage map. Several
polymorphic fragments could be assigned to QTL intervals of the reference map
and their correlation to malting quality traits was calculated. The favourite
alleles of differential TDFs increased the means of malting quality parameters
significantly compared to the means of the entire subpopulation. Differential
TDFs which are located within significant QTL intervals represent candidates
for functional genetic markers for malting quality and are therefore well
suitable for efficient selection in early stages of plant breeding.
Flowering Time Markers for Barley Breeding
1Department
of Genetics and Plant Production, Aula Dei Experimental Station, CSIC, 50080
Zaragoza, Spain, E-mail: igartua@eead.csic.es;
2IInstituto de Tecnología Agraria de Castilla y
47071-Valladolid,
Adjustment
of crop phenology to the resources of the production environment is among the
most important traits for barley adaptation. Three genes, Sgh,
Sgh2
and Sgh3,
are responsible for winter/spring growth habit and vernalization response. Two
more genes, Ppd-H1 and Ppd-H2, control photoperiod response. In most studies, the
main photoperiod and vernalization QTL coincide with the putative location of
the corresponding major genes. There are also a number of QTLs with lesser
effects on flowering, known as earliness per se
loci, more abundant, but showing less consistency across studies. All these
loci have been detected in a number of studies involving biparental progenies.
But their use in MAS requires knowledge of their effects and interactions at
the germplasm-pool level. For this reason, 17 populations of up to 20 doubled
haploids each, made of combinations of 14 parents routinely used in a
Spanish breeding program, are being phenotyped for heading time, and genotyped
with markers located in the vicinity of major photoperiod and vernalization
response loci. The objective is to identify a set of markers that can be used
to make predictions of flowering time of genotypes and candidate crosses. These
predictions would also be useful to perform MAS towards favorable loci
combinations. A progress report on association between markers and field
flowering time determinations will be presented.
The
Distribution of Retrotransposon-Based (S-SAP) Markers
in European Barley Cultivars
F.
J. Leigh, J. R. Law, V. J. Lea, E. Chiapparino, P. Donini and
J. C. Reeves
Molecular Research Group, National Institute of
Agricultural Botany,
E-mail: fiona.leigh@niab.com
The
retrotransposon based S-SAP technique has been shown to be a highly informative
method for genetic analysis. We have evaluated primers designed from the LTRs
of six retrotransposon families in conjunction with a large number of selective
MseI primers to evaluate the most informative primer combinations that generate
high quality profiles. Following extensive screening, we selected six of these
primer combinations from four of the retrotransposon families for wider
application. This subset of primer combinations has been used to profile a set
of over 500 barley cultivars using the S-SAP technique. The barley cultivars
were chosen to represent the barley germplasm cultivated across
Giving
the Right Gene Barley will Germinate on Time
C.
D. Li, R. Lance, A. N. Tarr, R. Appeals and
M. Cakir
Western Australia Department of Agriculture, South
Perth, WA 6151, Australia,
E-mail: cli@agric.wa.gov.au
Pre-harvest
sprouting results in significant economic loss for grain industry around the
world. Lack of adequate dormancy is the major reason for pre-harvest sprouting
in field under wet weather conditions. On the other hand, too much dormancy
also has detrimental effect in the malting house. A doubled haploid population,
derived from a cross of an Australian barley Chebec and a Canadian Malting
barley Harrington, was used to search for gene(s) controlling seed dormancy and
pre-harvest sprouting. The population was tested for pre-harvest sprouting
under high rainfall condition and tested for seed dormancy under rain-shelter.
One major locus was identified on chromosome 5H
to control pre-harvest sprouting, which could explain over 70% of the
phenotypic variation. The same locus also controls seed dormancy. Comparative
genomics approaches were used to identify the candidate gene(s) controlling
seed dormancy and pre-harvest sprouting. The barley dormancy/pre-harvest
sprouting locus showed high micro-synteny with the terminal end of rice
chromosome 3. The rice DNA sequences were annotated and a gene encoding
GA20-oxidase was identified as the gene controlling seed dormancy and
pre-harvest sprouting. This gene is specifically expressed in the developing
and germinating seeds. Different alleles of this gene with various levels of
dormancy were cloned and sequenced. Diagnostic molecular markers were developed
to distinguish the different alleles of the seed dormancy gene.
Identification
and Mapping of Disease Resistance-Related DNA Sequences in Barley
Z.
Liu, R. M. Biyashev, J. A. Mammadov and
M. A. Saghai-Maroof
Department of Crop and Soil Environmental Sciences,
Virginia Tech,
The objective of this study was to
identify disease resistance gene analogs (RGAs) from barley based on the use of
conserved motifs among cloned disease resistance genes. Over 150 barley RGA
clones were generated by using two pairs of degenerate primers designed from
nucleotide binding domain (NBD) of several previously cloned disease resistance
genes. The resultant RGAs were characterized based on both DNA sequence data
and RFLP patterns after Southern hybridization. Representative clones were
mapped using NABGMP mapping populations and disease resistance near-isogenuc
Imes. Thirteen different barley RGA classes were identified. The encoding amino
acids indicated that all these 13 clones, except one, contained continuous
open reading frames and had characteristic regions of NBD. RGA-related clones
were mapped to all barley chromosomes except chromosome 7. One RGA mapped to
chromosome 4 and at least 2 markers mapped to each of the remaining barley
chromosomes. Almost all of the RGAs mapped to barley chromosomal regions which
previously had been reported to contain known disease resistance factors. DNA
sequence comparisons showed that a diverse group of RGAs has been identified.
Genetic Mapping of Genes Affecting Interactions
between Barley
and Pyrenophora
teres-Fungus
O.
Manninen1, M. Serenius1,
1Department
Crops and Biotechnology, MTT
Agrifood Research Finland, Plant Production Research Unit,
FIN-31600 Jokioinen, Finland, E-mail:
outi.manninen@mtt.fi; 2All-Russian Institute for Plant Protection,
St-Petersburgh-Pushkin, 196606
Net blotch is a common and
economically important barley disease worldwide, especially in the temperate
and humid production areas. Net blotch is caused by the fungal phytopathogen Pyrenophora teres Drechsler. The most cost-effective and
environment-friendly way to control the net blotch disease is by using
resistant cultivars. We have used a well known resistance source, the Ethiopian
two-row barley line CI 9819, as the donor of resistance in our net blotch
resistance mapping progeny. A major resistance gene against net type isolates
was located on chromosome 6H, explaining up to 88% of the phenotypic variation
of seedling resistance. In addition, several epistatic minor genes were
detected on chromosomes 1H, 2H, 3H, 5H and 7H. The effect of the major
resistance locus on chromosome 6H has been verified in advanced backcross lines
in both greenhouse and field conditions. We also located a major resistance
gene against the Finnish spot type isolates on chromosome 5H, explaining up to
84% of the variation in seedling resistance. We are currently aiming to map P. teres
avirulence genes corresponding the resistance genes we have earlier mapped in
barley. We have crossed P.
teres isolates differing in their
virulence against the resistance source CI 9819. The single ascospore progeny
isolates have been analysed with AFLP markers and a preliminary linkage map has
been constructed. Genetic mapping or tagging of avirulence genes with the aid
of bulked segregant analysis is ongoing in several net blotch crosses.
A
Technology for Ultra Rapid Mapping of Complex Genomes
A.
Masoudi-Nejad1, P. H. Dear2 and R. Waugh1
1Scottish
Crop Research Institute, Invergowrie, Dundee, DD2 5DA, UK,
E-mail: rwaugh@scri.sari.ac.uk;
2MRC Laboratory of Molecular Biology,
Uncovering
the location and order of genes in a genome has fundamentally altered the
framework within which biological research is conducted. In humans and model
species this has been largely achieved through advances in DNA sequencing
chemistry, instrumentation and computational analysis. Though possible in
theory now for all organisms, financial realities dictate that for non-models
this objective is simply a pipedream. In this project we are extending a
generic technology we have pioneered on various species to barley. The approach
reduces the cost of locating and ordering genes on genomes to an estimated
0.11% of the current cost of a whole genome shotgun project. This is possible
using our current technology, which is based on liquid-handling. However, an
attainable challenge is to make the transition to automated solid phase
technology, which will further reduce the cost by ca. 90% and slash individual
project completion times to a matter of a few weeks. The approach integrates
well with other genomic technologies it can be used to greatly accelerate the
creation of physical (BAC- or other clone-based) maps, to guide sequencing
efforts, or to greatly refine genetic maps and thereby provide, for the first
time, a link between high-resolution physical maps and genetically-mapped traits
the bridge between genomics and genetics.
Identification of Marker Trait Associations in a
Barley
Four Way Cross
G.
L. McMichael1, J. K. Eglinton1,
A. R. Barr2 and K. J. Chalmers3
1School
of Agriculture and Wine, University of Adelaide, Waite Campus, Glen Osmond, SA
5064, Australia, E-mail: gai.mcmichael@adelaide.edu.au;
2Australian Grain Technology, University of Adelaide, Waite
Campus, Glen Osmond, SA 5064, Australia; 3Molecular
Plant Breeding CRC, University of Adelaide, Waite Campus, Glen Osmond, SA 5064,
Australia
Genetic mapping in cereals has been exclusively applied to populations
derived from simple crosses.Genetic studies are now beginning to target broader
population structures to take advantage of association mapping and whole genome
analysis techniques.This paper presents the preliminary results from the
genetic analysis of a population of 841 DH lines derived from the complex cross
Chieftan/Barque//Manley/VB9104.This cross is a significant departure from
typical mapping population structures. The population has been extensively
phenotyped for malt quality and adaptation characteristics, through evaluation
as a mainstream breeding population within the SA Barley Improvement Program
(SABIP). Of the 841 doubled haploid (DH) lines, 837 lines were evaluated in
double row trials, with 350 individuals promoted to stage one, 70 individuals
to stage two and ten individuals to stage three.One line (WI3408) has
subsequently progressed to pilot scale malting and brewing trials, with
potential for commercial release.Due to the large size of this population, high
throughput technology will be required for the genotyping. There were 290
microsatellite markers (SSRs), 90 of which were EST derived that were selected
for the initial parental screening, based on their association with the traits
of interest for this population.Of these combined markers, 60% were polymorphic
between one or more parents. An initial skeletal map is to be constructed using
a minimum of 48 SSRs from the parental screens. The 350 stage one lines will be
screened more comprehensively as will the 70 stage two lines and the ten stage
three lines respectively. The Gene Flow database will be the method of
recording and analysing the data.
The
Multivariate Data Analysis Revolution in Genetics,
Plant Breeding and Biotechnology
L.
Munck and B. Moeller
Department of Dairy and Food Science, The Royal
Veterinary and
DK-1958
The main focus in biology since the
rediscovering of the laws of Mendel 100 years ago has been centered around the
gene. This is also reflected in the choice of mathematical equations in
genetics and biotechnology focusing on gene effects and sequences by
probability calculus. As the classic geneticist Waddington in 1969 pointed out:
They say nothing what so ever about the actual phenotypes concerned. Classic
statistics is set up for studying populations and has difficulties in
characterizing phenotypes as individuums. This is now possible thanks to the
computer by the pattern recognition methods of multivariate data analysis using
a graphic interface for classification (Principal Component Analysis PCA) and
for correlation (Partial Least Squares Regression PLSR). These methods are now
extensively used in near infrared screening analyses and in the food and
brewing industry. Based on literature and the latest results from our
international research group on applied multivariate data analysis and
spectroscopy, this review paper aims at proving that multivariate data
analysis, now still in an introductory stage, will be able to revolutionise the
mutual connections between the genome and phenome aspects of data in genetics,
plant breeding and biotechnology. Thus whole spectroscopic fingerprints of the
physics and chemistry of barley seeds classified by PCA are differentiating
between carbohydrate and regulatory mutants and alleles and can as well be used
as specific multivariate selection criteria for improving a multigene quality
complex in e.g. malting barley breeding.
Identification of RAPD Markers Linked to Salt
Tolerance
in Cultivated and Wild Barleys
H.
Pakniyat1, A. Namayandeh1 and
B. P. Forster2
1Department
of Crop Production and Plant Breeding, Shiraz University, Shiraz, Iran;
2Scottish Crop Research Institute, Invergowrie, Dundee, DD2
5DA, UK, E-mail: bforst@scri.sari.ac.uk
Randomly
amplified polymorphic DNAs (RAPDs) were used to search for markers associated
with salt tolerance in barley. Initial screens involved growing 63 cultivated
and wild barley genotypes in saline conditions and testing for shoot sodium
content along with other physiological traits. From these tests 5 tolerant and
5 non-tolerant genotypes were selected. DNA from the tolerant and non-tolerant
genotypes were formed into two contrasting bulks and interrogated using 30
different 10-mer RAPD primers. One primer (P15) produced a band found only in
tolerant lines, and additionally produced a smaller product found only in the
non-tolerant group. Primer P10 produced a band specific to the tolerant bulk
and P22 produced a band specific to the non-tolerant group.
Marker-Assisted
Selection and Resistance Gene Pyramiding
in Barley
J. Vacke3
and V. Sip3
1Experimental
Institute for Cereal Research, 29017 Fiorenzuola dArda (PC),
E-mail: pecchioni.nicola@unimore.it;
2Universit degli Studi di
3Research
The soil-borne barley yellow mosaic
virus complex (BaMMV-BaYMV), the aphid-borne barley yellow dwarf virus (BYDV)
and the seed-borne fungus Pyrenophora
graminea (leaf stripe) are the most
serious diseases for the barley crop in
A Gene Mapped to Chromosome 7H in Barley (Hordeum vulgare) Causes Necrotic Leaf Spots and Less Susceptibility to
the Pathogen Puccinia hordei
M.
Persson, A. Djurle and A. Falk
Department of Plant Biology and
75007,
Barley plants defense themselves
with different reactions such as cell-wall appositions, accumulation of
pathogenesis related proteins, production of reactive oxygen species and
hypersensitive cell death. In this study we used a mutant line originally made
from Bowman-Rph 3, mutated with fast neutrons. The phenotype of the mutant is
dark brown spots on the leaves that are visible from the one leaf stage until
the old senescent plant. Microscopic analysis reveled autofluorescence in the
dark spots. To locate the gene responsible to this dramatic phenotype we used
BSA (bulked segregant analysis) in combination with a large number of AFLP
(Amplified Fragment Length Polymorphism) primer combinations. PstI
enzyme was used instead of EcoRI, due to its methylation sensitivity. To perform a
high-resolution genetic mapping a mapping population of 719 mutated F2
plants from the cross Mutant × Proctor were used. As a reference, 12 wildtype F2
plants from the same cross were used. The gene has been mapped to chromosome 7H
by the use of AFLP and Proctor-Nudinka mapping populations. The mutant
supported growth of the biotrophic barley pathogen Puccinia hordei
significantly less than the wildtype. In this study we will investigate
resistance qualities to the necrotrophic barley pathogen Bipolaris sorokiniana.
Small
Mapping Crosses and Their Use to Establish a Broad Based QTL Map for Barley
S.
J. Rae1, R. Keith1,
F. J. Leigh2, A. Mackie3,
D. Matthews2, G. Felix3,
P. C. Morris3,
P. Donini2 and
W. T. B. Thomas1
1Scotish
Crop Research Institute, Invergowrie, Dundee, DD2 5DA, UK, E-mail:
srae@scri.sari.ac.uk;
2Molecular Research Group, NIAB,
Cambridge, CB3 0LE, UK; 3School of Life Sciences, Heriot-Watt University, Riccarton,
Edinburgh, EH14 4AS, UK
Quantitative
traits (QTLs) are identified by selecting and hybridising parental lines that
differ in one or more QTL. Thus to examine numerous agronomic traits many
different crosses are required: This cross specific nature limits the adoption
of marker assisted breeding (MAS) in barley. We propose to maximise the number
of cross specific populations thereby enabling the production of one robust,
broad based QTL map: We have used 23 small mapping populations of a target size
of 20 individuals. These F1 doubled haploid (DH) populations were derived from
pairs of crosses made from
Genetic
Control of Grain Damage in a Spring Barley Mapping Population
R.
Rajasekaran, W. T. B. Thomas, A. Wilson, P. E. Lawrence, G. R. Young and
R. P. Ellis
Scottish Crop Research Institute, Invergowrie, Dundee,
DD2 5DA, UK,
E-mail: wthoma@scri.sari.ac.uk
A
genetic map was constructed, using a wide range of DNA-based markers, in a
barley mapping population developed to explore the genetic control of traits
concerned in grain damage of spring barley cultivars. Quantitative trait loci
(QTL) were located for husk skinning, gape between the lemma and palea and
splitting of the fused pericarp/testa/aleurone tissues. QTLs for the traits
were clustered at loci on chromosomes 1H, 4H, 5H, 6H and 7H along with QTL for
grain shape parameters. Historical changes in grain size and shape therefore
appear to have resulted in a greater propensity for grain splitting in some
modern cultivars. QTL analysis indicates the possibility of transgressive
segregation for grain splitting in random inbred lines from the Tankard × Livet
population as some Tankard alleles decrease splitting. The breeding of more
extreme lines is of potential concern to the malting industry as, without
extensive phenotypic assessment of splitting, such lines could potentially be
commercialised and thus put malting barley supplies at risk. Our genetic
mapping highlights regions of the genome that confer a splitting risk and
identifies potential markers for selection against the potential for grain to
split. Validation of the markers in a wider range of germplasm will establish
their potential value in marker-assisted selection.
Mapping
of Resistance Genes to Powdery Mildew in Barley
J.
Repkova1, Z. Kyjovska1,3,
P. Lizal1, A. Dreiseitl2 and A. Jahoor3
1Department
of Genetics and Molecular Biology, Faculty of Sciences, Masaryk University
Brno,
611 37
2Agricultural Research Institute Kromeriz, Ltd., 767 01
The
introduction of fully effective resistance genes from the wild barley Hordeum vulgare ssp. spontaneum
(PI354949 and PI466495) into H. vulgare
was performed by the cross with the variety Tiffany. Powdery mildew resistance
tests on plants of F2 generation revealed the segregation ratio 15:1 in
Tiffany × PI354949 and 3 : 1 in Tiffany × PI466495, which indicated presence of
2 and 1 dominant resistance genes, respectively. To identify individual R
genes, technology of microsatellite DNA markers and linkage recombination
analysis was applied. One of resistance genes to Blumeria graminis f.sp. hordei
in PI354949 was determined to be
located in Mla locus linked with Bmac0213 and the other on
chromosome 1(7H) linked with Bmag0507 (14.0 cM in proximal position). In
PI466495, R gene was linked with Bmac0213 (8.4 cM in distal
position) and it is also located in Mla
locus. The perspective aim of this work is fine-mapping of the promising gene
located on 7H chromosome and identification of tightly linked DNA markers for
marker assisted selection.
Single
Nucleotide Polymorphism Mapping of the Barley Genes Involved in Abiotic
Stresses
N. Rostoks1,
L. Cardle1, S. Mudie1,
J. T. Svensson2, H. Walia2,
E. M. Rodriguez2,
S. Wanamaker2,
P. E. Hedley1, H. Liu1, L. Ramsay1,
J. Russell1, T. J. Close2,
D. F. Marshall1 and
R. Waugh1
1Scottish Crop Research Institute, Invergowrie, Dundee,
DD2 5DA, UK, E-mail: rwaugh@scri.sari.ac.uk; 2Department
of Botany and Plant Sciences, University of California, Riverside, CA 92521,
USA
Changes in environmental conditions
and need for increased agricultural production require plant varieties tolerant
to various abiotic stresses. Recently, numerous genome-wide transcription
profiling studies have emerged identifying large number of genes involved in
plant response to abiotic stresses. In order to facilitate associations of
these genes with known plant traits, the information about genetic map location
is crucial. The advent of large-scale EST sequencing projects has provided
information about significant proportion of expressed plant genes and, thus,
offers an opportunity to identify mappable polymorphisms at the nucleotide
sequence level. As an additional benefit, Single Nucleotide Polymorphism (SNP)
markers are amenable to automation and high-throughput. Our aim is to identify
and characterize SNPs in 1000 barley genes known to be associated with
responses to abiotic stress in barley or homologous to such genes in other
plant species. The SNPs are discovered by sequencing 3 regions of the genes
from the parent lines of Steptoe × Morex, Owb D × Owb R and Lina × HS92 mapping
populations. The discovered SNPs are assembled into haplotypes which are mapped
in one or several of the mapping populations. The co-localization of the
haplotypes and known barley Quantitative Trait Loci (QTL) is investigated. Here
we report preliminary results on SNP discovery, mapping and SNP database
creation.
Development
of Gene Based Molecular Markers in Barley
J.
R. Russell, L. Ramsay, A. Booth, M. MACaulay,
L. Cardle, D. F.
W. Powell and
R. Waugh
Scottish Crop Research Institute, Invergowrie, Dundee,
DD2 5DA, UK,
E-mail: rwaugh@scri.sari.ac.uk
Novel gene based markers have an
enormous potential for exploitation in barley. Uses include the dense coverage
of the genome in genetic linkage maps through to the development of diagnostic
markers for agronomic and quality traits of interest. To date many of the
markers used successfully to delineate the barley genome have been based
primarily on anonymous DNA sequences, and therefore give little information on
the underlying genes that control traits. With the advent of large-scale EST
sequencing projects in barley there exists a rapidly expanding database of gene
sequences for the cereals, which can be used to develop molecular markers that
are associated with genes of interest. In order to construct a highly defined
gene map in barley, existing sequences can be mined for slight variations such
as single base changes (SNPs), repeat sequence expansion and contraction (SSRs)
and insertion and deletion events (indels) or alternatively variants can be
discovered from additional sequencing guided by existing information. The
resultant sequence-based genetic map will form the basic platform for many
applications including candidate gene isolation, marker-assisted selection,
genetic diversity studies, molecular taxonomy and association genetics studies.
Here we report results of ongoing work on the gene-based marker discovery,
mapping and utilisation with emphasis on a number of genes identified in grain
development and germination.
Identification
of Barley Semi-Dwarf Gene uzu
D.
Saisho1, K. Tanno1,
M. Chono2,
1Research
Institute for Bioresources, Okayama University of Science, Okayama 710-0046,
Japan,
E-mail: saisho@rib.okayama-u.ac.jp;
2Department of Wheat and Barley, National Institute of Crop
Science, National Agricultural Research Organization, Tsukuba 305-8518, Japan; 3Department
of Physiology and Quality Science, National Institute of Vegetables and Tea
Science, National Agricultural Research Organization, Ano, Mie 514-2392, Japan,
4Graduate School of Bioagricultural Science, Nagoya
University, Nagoya, 464-8601, Japan
Semi-dwarf
varieties of cereal crops have contributed to high yield and resistance to
lodging. The adaptation of these varieties is well known as the green
revolution in rice and wheat since 1960s. In barley, there is a single
recessive gene, called uzu, which shows a plant type similar to semi-dwarf
varieties of rice and wheat. The uzu lines are distributed only in
QTL
Analysis in Hordeum
bulbosum L. for Interspecific
Crossability and Hybrid Formation with Barley
H.
Salvo-Garrido, L. Fish, D. A. Laurie and
J. W. Snape
John Innes Centre, Norwich Research Park, Colney lane,
Norwich, NR4 7UH, UK,
E-mail: john.snape@bbsrc.ac.uke
H.
bulbosum is important in barley
breeding for haploid production and for the introgression of useful genes. An
RFLP map has been created which covers about 90 of the H. bulbosum
genome (Salvo-Garrido et
al. 2001). The two parents of this
mapping population also produce very different results when used as pollinators
onto barley. PB1 has high crossability and gives high rates of haploid
production; in contrast, PB11 has a lower crossability and gives a high
frequency of hybrids. Experiments were carried out to map QTL involved in these
different responses. To phenotype the 75 recombinant clones, all were
pollinated onto a barley cultivar, Triumph, under controlled environmental
conditions. For QTL mapping, MapQTL, version-3 was used. QTLs for crossability
were mapped in chromosomes 2Hb, 3Hb, 6Hb and 7H6. A novel QTL promoting hybrid
formation, heterozygous in the PB11 parent, was mapped in chromosome 6HA, and
explained about 40 of the phenotypic variation for this trait. This QTL would
enable more efficient selection of H. bulbosum
clones for use in introgressing traits from H. bulbosum
into barley. The QTLs for crossability identified will enable the
identification of more efficient clones for haploid production by MAS.
Identification of Quantitative Trait Loci Controlling
Morphological and Physiological Traits, which Are Characteristic between
Oriental and Occidental Barley Cultivars (Hordeum vulgare L.)
M.
Sameri and T. Komatsuda
Genetic Diversity Department, National Institute of
Agrobiological Sciences (NIAS), Tsukuba 305-8602,
A
total of 99 recombinant inbred lines (RILs) derived from a cross of Oriental
six-rowed × Occidental two-rowed cultivars were grown during the two seasons
2001 and 2002 to identify quantitative trait loci (QTL) controlling agronomic
traits such as spike characters (length, density, grain number, triplet number,
awn length), plant height, tiller number and days to heading. The RILs showed
wide variations for each trait, showing the wide range of genetic
diversification between Oriental and Occidental varieties and segregation in
the progenies. The composite interval mapping identified three QTLs affecting
plant height, of which one QTL on chromosome 7HL was newly identified in
addition to uzu gene and dsp1 gene. For spike length and density one new QTL was
identified on chromosome 2HL, which is closely linked to the cleistogamy gene
and Fusarium head blight resistance QTL. For heading date one new major QTL was
identified on chromosome 5HL at the same interval covering the Sgh2
locus, however lines with the spring parents allele were later heading. This
study showed the Oriental × Occidental barley RILs are good resources for
discovering of novel genes with agronomic importance.
High-Resolution
Mapping of Non-Brittle rachis1 (btr1) in Barley
D.
V. Saraswathi, P. Azhaguvel and T. Komatsuda
Department of Genetic Diversity, National Institute of
Agrobiological Sciences,Tsukuba, Ibaraka 305-8602,
High-resolution mapping has been
crucial for success of map-based cloning and precise placement of the gene of
interest. Among abundant variations in barley genome,
non-brittle rachis is a valuable trait in study of evolution and domestication
pattern of barley. AFLP markers around the btr1
locus previously mapped on 3HS chromosome were converted into sequence tagged
sites. Converted STS markers were taken for the high-resolution linkage map
study. OUH602 is a wild barley line having Btr1
and Btr2 complementary genes, while Kanto Nakate Gold is
non-brittle having btr1btr1Btr2Btr2. 672 F2 plants of KNG × OUH602 were raised in the year
20022003. Brittleness was evaluated 24 weeks after maturity. Linkage
analysis showed the btr1 locus is flanked between two STS markers at 0.3 cM
proximal and 0.5 cM distal. The order of marker loci was the same in F2
population of Azumamugi × Kanto Nakate Gold with 960 plants. The recombinants
were self-pollinated to produce F3 lines for further study. The F2
population of OUH602 × KNG showed good separation of the AFLP markers that were
previously mapped as clusters in 99 RILs of Azumamugi × Kanto Nakate Gold. This
high-resolution linkage map will serve as basis for map-based cloning of
non-brittle rachis locus, with the identification of BAC clones spanning the
region.
Retrotransposons as Genomic Sculptors and Molecular
Markers in Barley
A.
H. Schulman1,2, R. Kalendar1,
O. Manninen2, C. Stuart-Rogers1,
J. Tanskanen1
and C. Vicient1,3
1Plant
Genomics Laboratory, Institute of Biotechnology, University of Helsinki,
FIN-00014 Helsinki, Finland, E-mail: alan.schulman@helsinki.fi; 2Plant Breeding Biotechnology, MTT Agrifood Research Finland, Plant Production
Research Unit, FIN-31600
Jokioinen, Finland; 3Currently Departamento
de Genetica Molecular CID, CSIC, E-08034 Barcelona,
Spain
Only
a minor fraction, 5 to 10% of the barley genome is genic; most of the rest is
composed of retrotransposons and their derivatives. The retrotransposon life
cycle resembles the intracellular phase of retroviruses, and new copies are
integrated into the genome, without excision of existing copies. We are
studying the life cycle of elements representing the major classes
retrotransposons in the barley genome: BARE-1, Bagy-2, and two major classes of
non-autonomous, parasitic elements, TRIMs and LARDs. Because they represent a
major share of the genome, cause easily detectable genetic changes having known
ancestral and derived states, and contain conserved regions for which PCR
primers may be designed, retrotransposon insertions can be exploited as
powerful molecular marker systems. We develop and apply four key
retrotransposon-based methods: SSAP, IRAP, REMAP, and RBIP. The SSAP, IRAP, and
REMAP methods are multiplex and generate anonymous marker bands; RBIP scores individual loci, much as microsatellite-based marker
systems do. The methods are variously suited to marker detection on agarose and
polyacrylamide slab gels, slab and capillary sequencing devices, and arrays on
solid supports. We apply them to marker-assisted breeding, phylogenetic
analyses, biodiversity determinations, and evolutionary studies.
Physical
Mapping and Identification of Candidate Genes
at the Virus Resistance Gene Locus rym4/5
N.
Stein1, D. Perovic1,
B. Pellio2, J. Kumlehn1,
F. Ordon3 and A. Graner1
1Institute
of Plant Genetics and Crop Plant Research (IPK), D-06466 Gatersleben, Germany,
E-mail: stein@ipk-gatersleben.de;
2Institute of Crop Science and Plant Breeding I, Giessen
University, D-35392 Giessen, Germany;
3Institute of Epidemiology and Resistance, Federal Centre for
Breeding Research on Cultivated Plants (BAZ), D-06449 Aschersleben, Germany
In Europe barley yellow mosaic
disease is caused by a complex of three different viruses: BaMMV, BaYMV and
BaYMV-2. They belong to the bymovirus group- a subclass of the Potiviridae,
and are naturally transmitted via the soilborne fungus Polymyxa graminis. A number of recessive resistance genes has been
localised on at least 5 independent loci of the barley genome. The two genes rym4 and
rym5 reside at the same locus on chromosme 3HL. While rym4 confers
resistance to BaMMV and BaYMV, rym5 confers resistance to all three virus strains. In a
map based cloning effort, a physical BAC contig of over 450 kb has been
established at this locus and sequencing of over 350 kb has been accomplished.
200 kb of this region co-segregating with the gene in over 5,000 meiotic events
harbour two genes. One is a strong candidate for rym4/5
based on the BLASTX annotation. In a set of resistant and susceptible
accessions several rym4/5-diagnostic non-synonymous SNPs were detected in two
exons. Functional complementation of the candidate gene is in progress.
Molecular
Markers and Marker Assisted Selection
in Winter Barley Breeding
V.
Sudyova, M. Hudcovicova, L. Klcova and
J. Kraic
Division of Applied Genetics and Breeding, Research
Institute of Plant Production, 921 68 Piestany, Slovakia, E-mail:
sudyova@vurv.sk
Molecular
markers linked to BaYMV-1, BaYMV-2, and BaMMV resistance genes ym4
and ym11 was introduced into a winter barley varieties.
Acceptors of resistance genes were cultivars Copia and Tiffany. Cultivar
Romanze has been used as a donor of ym4
gene and landrace Russia 57 as a gene ym11 donor.
Total number of analyzed individuals of F2
progenies was 150. The codominant STS marker MWG838 linked to BaYMV/BaMMV
resistance gene ym4 showed a clear differentiation between
individuals possessing resistant- and susceptible-linked markers. 37 plants
with marker-based genotype ym4ym4
from the crosses Copia × Romanze and
Tiffany × Romanze were identified. Calculated values of 2
and probability (P > 0.05) of the observed F2
genotype segregation ratio from the type of crosses indicate that difference
between the expected (1:2:1) and observed segregation ratio are not
statisticaly significant. Microsatellite marker HVM3 linked to resistance gene ym11
distinguished 30 F2 individuals from the cross Copia × Russia 57. Difference
between the expected and observed segregation ratio is not statisticaly
significant (23:1 =
2.000, P = 0.1572). New created winter barley lines homozygous
at ym4- and ym11-linked marker loci possess with high probability
desired resistance genes against BaYMV/BaMMV virus complex. Their effective
resistance against virus complex will be further tested.
Genetic Analysis of Cleistogamy in Barley
Y.
Turuspekov1, Y. Mano2, I. Honda1,
Y. Watanabe1 and
T. Komatsuda2
1National
Institute of Crop Science, Tsukuba 305-8518, Japan,
E-mail: yerlan@affrc.go.jp;
2National Institute of Agrobiological Sciences, Tsukuba
305-8602, Japan
Cleistogamy (CL) in barley could be
potentially helpful in resistance to pathogens arising during the flowering
time and in restriction of gene flow from genetically modified products. Knowledge
of the genetic control of CL can also help better understand important
theoretical aspects of biology. In this work we have attempted to study the
genetic control of CL using different segregating populations. Using 99 RILs
population of Azumamugi × Kanto Nakate Gold, where F1
was chasmogamous (CH), cly1 locus has been mapped on the long arm of chromosome
2H. This locus was co-segregating with AFLP marker e11m19-3 and 3.3 cM proximal
to RFLP marker ABC153. Also, mapping analysis of F2
population between Misato Golden and Satsuki Nijo (F1
was CL and in F2 CL dominated over CH with a ratio 3:1) revealed the
location of Cly2 locus in the same position of chromosome 2H. In this
population, the distances from STS-e11-m19-3 and ABC153 were 1.1 cM and 4.9 cM,
respectively. Considering the opposite gene domination in two different mapping
populations and that both loci were mapped in the same region, it was concluded
that the expression of CL in barley is could be under the control of two
closely located genes.
Genetic
and Physical Mapping of Genic Microsatellites
in Barley (Hordeum vulgare
L.)
R.
K. Varshney1, U. Haehnel1,
T. Thiel1, N. Stein1,
L. Altschmied1, P. Langridge2 and
A. Graner1
1Institute
of Plant Genetics and Crop Plant Research (IPK), D-06466 Gatersleben, Germany,
E-mail: rajeev@ipk-gatersleben.de; 2University
of Adelaide, Waite Campus, Glen Osmond, SA 5064, Australia
A set of
111,090 barley ESTs (corresponding to 55.9 Mb), generated at IPK, was employed
for searching of microsatellites (or simple sequence repeats, SSRs) as a source
for the development of SSR markers. With the help of a PERL5 script (MISA,
http://pgrc.ipk-gatersleben.de/misa/) 9,564 microsatellites were identified in
a total of 8,766 ESTs (SSR-ESTs). Cluster-analysis revealed 2,823 non-redundant
SSR-ESTs. From this, a set of 756 primer pairs was designed and as a result,
190 microsatellite (EST-SSRs) loci were placed onto the barley genetic map.
These markers show a uniform distribution on all the linkage groups ranging
from 22 markers (on 7H) to 36 markers (3H). The polymorphism information
content (PIC) for the markers developed ranges from 0.24 to 0.78 with an
average of 0.48. Developed markers represent functional class of marker system
as they are derived from ESTs (genes). Bioinformatic analyses suggested a
putative function for 55.8% (107) markers of which 53.2% (57) SSR-ESTs were
assigned to functional categories as per FunCat catalogue of proteins. A total
of 74% barley markers showed their transferability in wheat and rye while 41%
did so in rice. Furthermore in the direction of physical mapping of barley
genome, a PCR-based strategy was established to screen the BAC library with
markers. By using this strategy with Morex-BAC library BAC addresses were
obtained for a total of 132 mapped EST-SSRs which may provide anchoring points
to correlate the barley genetic map with a future physical map. Moreover,
eleven groups of markers were recognized on 6 chromosomes of barley which
contain one or more BAC clones on which at least two markers of that group
representing different genes are located. This analysis leads to the
identification of gene-rich regions in barley genome.
Multi-QTL
Mapping of Caryopsis Dormancy and Seedling Desiccation Tolerance of Barley
F.
Zhang1,3, G. Chen2, Q. Huang2,
O. Orion2, T. Krugman2,
T. Fahima2, A. B. Korol2,
E. Nevo2
and Y. Gutterman1
1Wyler
Department of Dryland Agriculture, Jacob Blaustein Institute for Desert
Research, Ben-Gurion University of the Negev, Sede Boker Campus 84990, Israel; 2Institute
of Evolution, University of Haifa,
Haifa 31905, Israel, E-mail: nevo@research.haifa.ac.il;
3Institute of Forestry, Chinese Academy of Forestry, Haidian
District, Beijing 100091, P. R. China
The
genomic regions controlling caryopsis dormancy and seedling desiccation
tolerance were identified with 152 F4
lines derived from a cross between Mona, a Swedish cultivar, and Wadi Qilt, an
Israeli wild barley (Hordeum
spontaneum). Dormancy, the inability of
a viable seed to germinate, and desiccation tolerance, the ability of the
desiccated seedlings to revive after rehydration, were characterized by fitting
the germination and revival data with growth curves, using three parameters
instead of initial scores: minimum, maximum, and speed of germination or
revival rate derived by least square method. The genetic map (2040 cM in
length) was constructed with Eighty-five genetic markers (SSRs, AFLPs, TSs and
Dhn) and the QTL mapping was conducted with MultiQTL package
(http://www.multiqtl.com). Seventeen QTLs were detected, and ten out of these
affected both dormancy and desiccation tolerance traits. No QTL was identified
for germination speed, while six QTLs were for revival speed. More QTLs were
for minimum germination than maximum germination, whereas more QTLs were for
maximum revival rate than minimum revival rate. Most of the QTLs with strong
effects were located on chromosome 4H. Both xeric wild barley and cultivar Mona
contributed favorite alleles for caryopsis dormancy and seedling desiccation
tolerance. The results revealed that the QTL effects on minimum germination
rate underlay the genetic control of caryopsis dormancy, and that the seedling
desiccation tolerance was based on the QTL effects on the maximum revival rate
and revival speed.