Copyright 2004 by the Genetics Society of America
A Chromosome Bin Map of 2148 Expressed Sequence Tag Loci of
Wheat Homoeologous Group 7
K. G. Hossain,* V. Kalavacharla,* G. R. Lazo,†J. Hegstad,* M. J. Wentz,* P. M. A. Kianian,*
K. Simons,* S. Gehlhar,* J. L. Rust,* R. R. Syamala,* K. Obeori,* S. Bhamidimarri,*
P. Karunadharma,* S. Chao,‡,1O. D. Anderson,†L. L. Qi,§B. Echalier,§B. S. Gill,§
A. M. Linkiewicz,¶,2A. Ratnasiri,¶J. Dubcovsky,¶E. D. Akhunov,¶J. Dvor ˇa ´k,¶
Miftahudin,&K. Ross,** J. P. Gustafson,** H. S. Radhawa,††M. Dilbirligi,††
K. S. Gill,††J. H. Peng,‡‡N. L. V. Lapitan,‡‡R. A. Greene,§§
C. E. Bermudez-Kandianis,§§M. E. Sorrells,§§O. Feril,&
M. S. Pathan,&H. T. Nguyen,&J. L. Gonzalez-Hernandez,¶¶,3
E. J. Conley,¶¶J. A. Anderson,¶¶D. W. Choi,&&
D. Fenton,&&T. J. Close,&&P. E. McGuire,‡
C. O. Qualset‡and S. F. Kianian*,4
*Department of Plant Sciences, North Dakota State University, Fargo, North Dakota 58105,†USDA-ARS Western Regional Research Center,
Albany, California 94710-1105,‡Genetic Resources Conservation Program, University of California, Davis, California 95616,
§Department of Plant Pathology, Wheat Genetics Resource Center, Kansas State University, Manhattan, Kansas 66506-5502,
¶Department of Agronomy and Range Science, University of California, Davis, California 95616,&Department of
Agronomy, University of Missouri, Columbia, Missouri 65211, **USDA-ARS Plant Genetics Research Unit,
Department of Agronomy, University of Missouri, Columbia, Missouri 65211,††Department of Crop and
Soil Sciences, Washington State University, Pullman, Washington 99164-6420,‡‡Department of Soil and
Crop Sciences, Colorado State University, Fort Collins, Colorado 80523-1170,§§Department of Plant
Breeding, Cornell University, Ithaca, New York 14853,¶¶Department of Agronomy and
Plant Genetics, University of Minnesota, St. Paul, Minnesota 55108 and&&Department
of Botany and Plant Sciences, University of California, Riverside, California 92521
Manuscript received December 12, 2003
Accepted for publication June 1, 2004
The objectives of this study were to develop a high-density chromosome bin map of homoeologous
group 7 in hexaploid wheat (Triticum aestivum L.), to identify gene distribution in these chromosomes,
and to perform comparative studies of wheat with rice and barley. We mapped 2148 loci from 919 EST
clones onto group 7 chromosomes of wheat. In the majority of cases the numbers of loci were significantly
lower in the centromeric regions and tended to increase in the distal regions. The level of duplicated
loci in this group was 24% with most of these loci being localized toward the distal regions. One hundred
nineteen EST probes that hybridized to three fragments and mapped to the three group 7 chromosomes
were designated landmark probes and were used to construct a consensus homoeologous group 7 map.
An additional 49 probes that mapped to 7AS, 7DS, and the ancestral translocated segment involving 7BS
also were designated landmarks. Landmark probe orders and comparative maps of wheat, rice, and barley
were produced on the basis of corresponding rice BAC/PAC and genetic markers that mapped on
chromosomes 6 and 8 of rice. Identification of landmark ESTs and development of consensus maps may
provide a framework of conserved coding regions predating the evolution of wheat genomes.
bases per haploid cell, which is 35 times larger than
OMMON wheat (Triticum aestivum L., 2n ? 6x ?
42, AABBDD) has a genome of ?16 million kilo-
that of rice (Oryza sativa L.) and ?110 times that of
Arabidopsis (Bennett and Smith 1976). It is composed
of three genomes, contributed by T. uratu Tum. ex
Gand. (A genome), Aegilops speltoides Tausch or an ex-
tinct close relative (B genome), and Ae. tauschii Coss.
(D genome; McFadden and Sears 1946; Kihara 1954;
Nishikawa1983;Dvor ˇa ´kand Zhang1990).Theorder
of loci in these three genomes is thought to be colinear
except for a 4A–5A–7B translocation, a putative 2B–6B
translocation, and two inversions on chromosome 4A
(Devos et al. 1995; Mickelson-Young et al. 1995).
A complete series of aneuploid wheat lines missing
1Present address:USDA-ARS Biosciences ResearchLaboratory, Fargo,
2Present address: Plant Breeding and Acclimatization Institute, Radzi-
kow 05-870 Blonie, Poland.
3Present address: Department of Plant Sciences, North Dakota State
University, Fargo, ND 58105-5051.
4Corresponding author: Department of Plant Sciences, North Dakota
State University, Fargo, ND 58105.E-mail firstname.lastname@example.org
Genetics 168: 687–699 (October 2004)
688K. G. Hossain et al.
an entire chromosome or an arm of a chromosome has
been developed (Sears 1954). More recently, a system
of generating an unlimited number of deletion lines
has become available; a chromosome with gametocidal
properties from Ae. cylindrica host was used to generate
frequent chromosome breaks in the wheat background
(Endo 1988). The deletions were isolated in a wheat
background as the breaks were caused only in the ga-
metes lacking the alien chromosome. Most of the dele-
tions were from a single break followed by the loss of
the chromosome region distal to the breakpoint (Endo
1990). The systematic production of common wheat
stocks containing terminal chromosomal deletions of
involving all 21 wheat chromosomes have been isolated
(Endo and Gill 1996). From this collection, deletion
lines were selected for the present study that provided
extensive coverage of the wheat genome, subdividing it
into 159 chromosome bins (Qi et al. 2003).
Physical maps of RFLPs produced using deletion
stocks have been reported for the chromosomes of all
seven homoeologous groups of hexaploid wheat (Gill
et al. 1993; Kota et al. 1993; Hohmann et al. 1994, 1995;
Delaney et al. 1995a,b; Mickelson-Young et al. 1995;
Gill et al. 1996a,b). Arm-specific physical maps and
identification of gene-rich areas and genes controlling
phenotypic traits have also been reported (Endo and
Mukai 1988; Endo et al. 1991; Endo and Gill 1996;
Faris et al. 2000; Sandhu et al. 2001; Weng and Lazar
2002). The physical maps of homoeologous group 6
and group 7 chromosomes and a comparative map of
chromosomes 7 of wheat and barley (Hordeum vulgare
L.) have been reported (Werner et al. 1992; Hohmann
et al. 1995; Weng et al. 2000). Landmark loci, which
represent cDNA clones and single- or low-copy genomic
DNAs that correspond to highly conserved coding re-
in understanding genome evolution among the species
of Triticeae. Conserved linkages with similar gene con-
tent and gene order have been reported among many
related species (Tanksley et al. 1992; Ahn and Tanks-
ley 1993; Sorrells et al. 2003). The high colinearity
of molecular markers between wheat and barley ge-
which will accelerate integrative mapping among species
(Devos et al. 1995; Van Deynze et al. 1995; Dubcovsky
et al. 1996). Analyzing the degree of linkage conserva-
tion and synteny of chromosome segments between the
homoeologous group 7 chromosomes of wheat and bar-
ley,Hohmann etal.(1995)identified extensivehomolo-
gies between these chromosomes.
of cDNA clones that correspond to mRNA and facilitate
the identification of many genes (Adams et al. 1991).
These sequences have been used to develop new molec-
ular markers to analyze genome structure and to dis-
cover genes in many organisms, such as human, mouse,
rat, Medicago trunculata, maize (Zea mays L.), and rice
(Adams et al. 1991, 1995; Hillier et al. 1996; Covitz et
al. 1998; Ewing et al. 1999; Marra et al. 1999; Scheetz
et al. 2001; Fernandes et al. 2002). Previously, 238 genes
with orthologous locations among the three genomes
of wheat were identified. Thirty-nine (16.5%) of these
genes were localized in the chromosomes of group 7
(McIntosh etal. 2003). Recently,a consortiumof scien-
tists (Lazo et al. 2004) identified ?117,000 ESTs devel-
oped from the sequences of cDNAs of different tissues
and developmental stages primarily of hexaploid wheat
ESTs from this collection representing wheat uni-
genes were physically mapped to individual chromo-
somes/chromosomal intervals using wheat nullisomic
and ditelosomic lines and deletion stocks (Sears 1966;
Endo and Gill 1996). This study summarizes the map-
ping of ?2000 EST loci to the three homoeologous
group 7 chromosomes of wheat, an assessment of con-
served loci, and the distribution of mapped EST loci to
the chromosome bins defined by the deletion stocks.
Patterns of distribution and duplication of loci within
and among the group 7 chromosomes of wheat and
comparisons with rice and barley genomes were investi-
gated. This is the first report of the mapping of such a
large number of ESTs to this homoeologous group, and
location of these genes in the rice genome offers the
possibility of positioning similar genes across grass ge-
MATERIALS AND METHODS
Genetic stocks: In this study various cytogenetic stocks of
the hexaploid wheat cultivar Chinese Spring (T. aestivum)
osomic (DT), and 101 deletion lines (del) lines (Sears 1954,
1966; Sears and Sears 1978; Endo andGill 1996).Adetailed
description of these stocks is provided in Qi et al. (2003).
The fraction length (FL) value of each deletion breakpoint
identifies the position of the breakpoint from the centromere
relative to the length of the complete arm. A bin is defined
by two deletion breakpoints and is given a name followed by
the arm fraction-length endpoints for which the deletion is
diagnostic; e.g., 7AL16-0.86-0.90 designates the region from a
breakpoint at 86% of the 7AL arm to one at 90%. These
aneuploid and deletion stocks provide a complete coverage
of the wheat genome, subdividing it into 159 chromosome
bins. All the genetic stocks selected for EST mapping were
cytologically and/or molecularly verified by C-banding and
Southern hybridization with ?500 EST clones (Qi et al. 2003).
EST singletons: The clones used in this study were devel-
oped from cDNA libraries of different tissues and develop-
mental stages of wheat and other related species in the Triti-
ceae tribe (Lazo et al. 2004; Zhang et al. 2004). The cDNA
clones were sequenced and clones with unique sequences
(unigenes) were used in this study as probes for mapping all
loci along the chromosomes of wheat genomes approximates
mapped-EST distribution in wheat.
Atthe U.S.Department ofAgriculture (USDA)-Agricultural
689Physical Map of EST Clones in Wheat Homoeologous Group 7
Research Service (ARS) Western Regional Research Center
(Albany, CA), ?117,000 ESTs were produced from 43 cDNA
libraries (primarily of wheat) representing a wide range of
tissues, developmental stages, and environmental stresses (Lazo
et al. 2004). Amplified PCR products for unigenes (inserts)
were prepared and sent to 10 mapping laboratories (http:/ /
wheat.pw.usda.gov/NSF) for Southern hybridization.
Southern hybridization: Procedures used for genomic DNA
isolation, restriction endonuclease digestion, gel electropho-
resis, and DNA gel blot hybridization were as described in Qi
et al. (2003) and are available on-line at http:/ /wheat.pw.usda.
genomic DNA was digested with EcoRI. Lambda DNA digested
with HindIII and BstEII was used as a size marker. Images of
all autoradiographs are available on line at http:/ /wheat.pw.
Localization of ESTs: EST loci were assigned to a specific
chromosome, arm, and/or deletion bins on the basis of the
presence or absence of the restriction fragments in a given
set of DNA lanes of a Southern blot (Sears 1954, 1966; Endo
and Gill 1996). Details of the mapping procedure can be
found at http:/ /wheat.pw.usda.gov/NSF/project/mapping.
Map construction: The group at North Dakota State Univer-
sity was responsible for analyzing the mapping data for homo-
eologous group 7 chromosomes (http:/ /wheat.pw.usda.gov/
cgi-bin/westsql/map_locus.cgi). On the basis of the physical
and on the relative length of chromosome intervals (bins),
the expected number of EST loci for each was calculated
(Gill et al. 1991; Endo and Gill 1996). The ?2-test was used
to test for randomness of the distribution patterns of EST loci
among the chromosomes, chromosome arms, and deletion
bins of group 7 of wheat.
The EST probes that hybridized to only three RFLP frag-
ments and mapped across the three genomes were identified.
The homoeologous map positions of the loci produced by
these probes and those associated with the ancestral transloca-
tion involving 7BS were identified on the basis of the overlap-
ping FL values in the bins and were used in the construction
of a consensus physical map of group 7.
Those ESTs that hybridized to more than three fragments,
many of which mapped onto homoeologous group 7 chromo-
somes, identified duplicated loci. On the basis of the pattern
of duplicated loci, different classes of duplicated regions were
evident. Whenever duplicated fragments of a particular EST
mapped in the same deletion bin it was considered as an
intrabin duplication. Interchromosomal duplication was de-
fined as those events where duplicated loci of a particular EST
mapped to chromosome deletion bins other than those in
group 7. The consensus duplication was defined for those
ESTs whose loci mapped to consensus positions across the
chromosomes of homoeologous group 7 as well as to consen-
sus positions across the chromosomes of another homoeolo-
for those ESTs whose loci were duplicated into the same or
another arm of chromosomes of homoeologous group 7.
EST density: The proportion of the chromosome for each
deletion bin was calculated on the basis of Gill et al. (1991).
The physical length, arm ratio data of a chromosome, and
proportion of arm missing in chromosome bins were used
in calculating the megabase values in deletion breakpoint-
defined chromosome bins.
Chi-square statistics to test the homogeneity of EST content
frequencies of ESTs in chromosome bins were tested against
the null hypothesis of uniform EST distribution along the
chromosome arms. Under the null hypothesis, the expected
number of ESTs is proportional to the length of the bin. The
distribution of EST loci along the physical length of each
missing segment was analyzed by estimating the ratio of the
percentage of mapped loci to the relative percentage of miss-
ing arm in the deletion breakpoint-defined regions (Weng
and Lazar 2002).
Ordering ESTs into chromosome bins and comparison of
map positions: Deletion mapping provides a fast and efficient
method of locating many loci within a chromosome bin; how-
ever, the order of loci within a bin cannot be determined. A
putative order of ESTs can be inferred using in silico compari-
son to the rice genome sequence as reported by Sorrells et
al. (2003). Each mapped EST locus is a unique EcoRI restric-
tion digest signature of known molecular weight, marking a
specific expressed segment of an individual chromosome of
wheat. We considered only ordering of the ESTs placed on
tifiedby searchingtheESTs athttp:/ /www.gramene.org/perl/
SeqTable and genetically mapped molecular markers corre-
sponding to BAC/PACs were identified. On the basis of centi-
morgan distances of these markers in different rice chromo-
somes, therelative orderof ESTsin thedifferent chromosome
bins was determined, and the map positions were compared.
somes of group 7 of wheat and barley, map positions of barley
chromosome 7 markers were compared to those of wheat
Corresponding rice BAC/PACs with markers were identified
by searching the sequences of RFLP markers of chromosome
7 of barley at http:/ /www.gramene.org/perl/SeqTable (Ku ¨n-
zel et al. 2000). Identified BAC/PACs were compared with
the BAC/PACs and ESTs previously identified in the wheat
Chromosome bin maps of 7A, 7B, and 7D: Nine hun-
dred nineteen EST probes were mapped in homoeolo-
gous group 7 chromosomes of wheat, identifying 2148
661 loci, 549 mapped on 7B and identified 719 loci,
and 613 mapped on 7D and identified 768 loci. The
distributions of ESTs without duplication and showing
no ambiguous loci in 7A, 7B, and 7D are presented
in Figure 1, A, B, and C, respectively. The ?2analysis
indicated a significantly higher number of EST loci
A total of 267 probes mapped to all three chromo-
somes of homoeologous group 7. Of these, 119 probes
were unique, identifying only three loci. These probes
were highly conserved among the genomes and could
map of group 7 was developed with 117 (consensus
position could not be resolved for two ESTs) of these
probes, providing a framework map for chromosome 7
(Figure 2, A and B). An additional 49 probes could be
added to this group if the ancestral translocation event
involving 7BS was considered. These markers mapped
to 7AS, 7DS, and the 5AL-4AL segment derived from
7B (Figure 2B).
Distribution of the loci and gene density: All of the
23 deletion breakpoints defined bins among the homo-
690K. G. Hossain et al.
Figure 1.—Physical EST maps of chromosomes of group 7 showing distribution of ESTs mapped in different deletion bins.
The deletion bins for each chromosome are marked on the left. The intrabin duplicated loci are represented only once per
bin. The number of loci per bin is that presented in Table 1 minus the duplications. Information on the exact locus designation
and restriction fragment mapped to each bin can be found at http:/ /wheat.pw.usda.gov/cgi-bin/westsql/map_locus.cgi. The
short arm of each chromosome is oriented toward the top while the long arm is toward the bottom. (A) physical map of
chromosome 7A; (B) physical map of chromosome 7B; (C) physical map of chromosome 7D.
eologous group 7 chromosomes contained different
numbers of EST loci (Table 1). A trend for increasing
numbers of EST loci mapped from proximal to distal
regions of all chromosome arms was observed. All of
the centromeric bins except C-7BS1-0.27 contained a
significantly lower number of EST loci than expected
on the basis of the size of the bins (Table 1). Except
for the distal bin of chromosome arm 7BS (7BS1-0.27-
numbers of EST loci. The relative density of ESTs was
expressed as the percentage of mapped loci per unit of
physical length for each deletion bin and chromosome
is physically 32% of the arm length and 39% of the loci
to this bin (Table 1). Assuming the physical length of
7AL is 100 units then the ratio of mapped loci per unit
arm length is ?1.22. From Table 1, it is clear that in
except for the short arm of chromosome 7B. The trend
in 7BS can be explained by a double translocation event
involving this arm (details presented later). The per-
chromosome deletion bins of all chromosome arms var-
ied from 0.71 to 3.75.
The megabase content of DNA and gene densities
for all deletion breakpoint-defined regions are summa-
691Physical Map of EST Clones in Wheat Homoeologous Group 7
rized in Table 1. For example, in the short arm of chro-
mosome 7A, four deletion breakpoint-defined regions
four loci mapped to bin 7AS1-0.89-1.00, while 108 loci
mapped to bin 7AS5-0.59-0.89, and 46 loci mapped to
bin 7AS8-0.45-0.59. On average, 72% of all mapped loci
in homoeologous group 7 chromosomes were located
in the distal regions, and the number of loci mapped
in these regions was about seven times higher than
that in the centromeric chromosome bins. The highest
density of EST loci was observed in bin 7AL16-0.86-
0.90 followed by bin 7BL10-0.78-1.00. The megabase
contents of these chromosome bins were 16.30 and
119.02, and 45 and 184 loci were mapped in these bins,
genomes of homoeologous group 7 chromosomes, 63
(24%) identified duplicated loci either on the chromo-
somes of other homoeologous groups or on the same
of these ESTs identified duplicated loci placed on the
consensus regions across the three genomes of groups
3,5,and 6.Eighty-five(31%)ESTs identifiedduplicated
loci in the same deletion bin where they were placed.
The distribution of ESTs producing duplicated loci
along the length of the consensus chromosome 7 is
presented in Figure 3 and the number of ESTs with
each type of locus duplication pattern is presented in
7BS ? 4AL and 5AL ? 7BS translocations: Of the
919 ESTs mapped to homoeologous group 7, 44 were
mapped on the short arms of chromosomes 7A and 7D,
7B they mapped to the long arm of chromosome 4A
(Figure 2B). Out of these 44 probes 31 mapped to bin
7AS1-0.89-1.00 and 13 mapped to 7AS5-0.59-0.89. All of
these 44 probes mapped to the distal bin of 7DS (7DS4-
0.61-1.00). These probes also mapped to the distal 41%
of the long arm of chromosome 4A. Twenty-nine
mapped into bin 4AL4-0.80-1.00, 13 into bin 4AL5-0.66-
0.80, and 2 mapped into bin 4AL13-0.59-0.66.
Five of the probes mapped to the long arms of chro-
692K. G. Hossain et al.
region on chromosome 5A, they mapped to the short
arm of chromosome 7B (Figure 2B).
Ordering the EST loci among chromosome bins and
comparison of map positions with rice and barley: Of
the 117 probes (excluding those involved in the 7BS
logous group 7, the possible order for 38 probes was
determined on the basis of the molecular markers
mapped to chromosomes 6 and 8 of rice (Figure 4), on
the assumption of retained colinearity of loci among
these species (Sorrells et al. 2003). In general, the
while all those mapping to rice chromosome 8 mapped
near the wheat centromere (Table 3, Figure 4). All the
ESTs in interval 0.45–0.59 of the short arm corre-
mapped to rice chromosome 8 (Table 3, Figure 4).
Eleven molecular markers mapped on chromosome 7
of barley identified corresponding EST probes mapped
to consensuschromosome 7ofwheat,eight ofwhichmain-
tained the same relative order in both genomes. Three
EST probes (BE500615, BE446380, and BE424174)
mapped in reverse order into the 0.45–0.59 region on
the short arm of wheat chromosome 7 (Figure 4).
Chromosome bin maps of 7A, 7B, and 7D: A total of
661, 719, and 768 loci were mapped to wheat chromo-
somes 7A, 7B, and 7D, respectively. On the basis of
the relative sizes of these homoeologous chromosomes
(7B ? 7A ? 7D), significantly higher numbers of EST
loci were mapped to 7D followed by 7B and 7A (Figure
1, A, B, and C, respectively), which was in agreement
with the findings of Qi et al. (2003). The reduced num-
ber of loci on 7B might be explained by the reciprocal
translocation, where unequal size fragments were ex-
changed between 4AL, 7BS, and 5AL. There are 44 7BS-
specific ESTs on 4AL and 5 5AL-specific ESTs translo-
cated to 7BS. Due to this uneven exchange, 7BS had
693Physical Map of EST Clones in Wheat Homoeologous Group 7
Figure 2.—Consensus map of homoeologous group 7 of wheat including the ancient translocation involving 7BS, 4AL, and
5AL. (A) consensus map of the long arm of group 7; (B) consensus map of 7AS and 7DS (on the left) with landmark probes
shared with 7BS (0.00–0.59 interval). Forty-four EST detected loci mapped to 7AS and 7DS (0.59–1.00 interval) and translocated
from 7BS to 4AL. A smaller segment of 5 EST detected loci representing an unequal translocation from 5AL to 7BS. Probes in
boldface type were ordered on the basis of the corresponding markers mapped on the linkage maps of chromosomes 6 and 8
the lowest number of EST loci as compared to other
Distribution of loci and gene density: Physical RFLP
maps produced using deletion stocks have been re-
ported for each of the seven homoeologous chromo-
some groups and for some chromosome arms (Werner
et al. 1992; Kota et al. 1993; Hohmann et al. 1994, 1995;
Delaney et al. 1995b; Mickelson-Young et al. 1995;
Endo and Gill 1996; Gill et al. 1996a,b; Faris et al.
2000; Sandhu et al. 2001; Weng and Lazar 2002). In
most cases the numbers of markers analyzed were rela-
tively low, and a majority of the markers were of un-
known function or were genomic probes. The highest
gous chromosome 7’s was 111, and only 21 of these
were cDNA probes (Hohmann et al. 1995).
A higher marker density was generally observed in
the distal regions as compared to the proximal regions
of chromosome arms. Akhunov et al. (2003a) analyzed
694K. G. Hossain et al.
Chromosome bins with relative physical length, number of loci per bin, and ratio of
mapped loci per unit arm length of group 7 of wheat
Chromosome arm and
DNA content (Mb)a
arm lengthChromosome bin % armMb DNA
7AS ? 407.53, ?2? 0.226C-7AS8-0.45
7AL ? 407.53, ?2? 0.226
7BS ? 360.65, ?2? 0.57
7BL ? 540.98, ?2? 0.38
7DS ? 346.90, ?2? 0.620
7DL ? 381.59, ?2? 0.51
aRelative distributions of loci mapped per chromosome were 7A ? 815.06, ?2? 5.49, *P ? 0.05; 7B ? 901.63, ?2? 6.41,
**P ? 0.01; and 7D ? 728.49, ?2? 22.33, ***P ? 0.005.
bThe loci presented here are those that were unambiguously assigned to each bin and not to the entire region, chromosome
arm, or chromosome.
cSignificance levels were *P ? 0.05, **P ? 0.01, and ***P ? 0.005.
the distribution of EST loci in all chromosomes of the
wheat genome using a subset of this project’s mapped
EST database and determined that in each arm the
density of mapped loci increases from the centromeric
in our study along the arms of group 7 chromosomes
supports that pattern.
The highest density of EST loci, as revealed by ?2and
ratio of percentage of mapped loci per unit arm length,
was observed in bin 7AL16-0.86-0.90, which agrees with
Hohmann et al. (1995). The second-highest density of
megabase content of bin 7AL16-0.86-0.90 is 16.30 and
45 loci were mapped in this bin; therefore, on average,
1 EST locus was mapped for every 362 kb in this region.
The difference in distribution of recombination along
the chromosome lengthmeans that the amountof DNA
per centimorgan varies depending on the location of
the gene on a chromosome. A higher density of EST
Number of ESTs in different duplication patterns
Duplication patternNo. of ESTs
Unique (consensus) duplication
Figure 3.—Pattern of distribution of EST loci duplicated
along the length of the arms of consensus chromosome 7.
The vertical axis is the number of loci. The horizontal axis
dividesthe shortandlong armsintoapproximately aproximal
one-third and distal two-thirds portions.
695Physical Map of EST Clones in Wheat Homoeologous Group 7
Figure 4.—Comparison of map positions of
molecular markers of rice and barley with ten-
of homoeologous group 7.The linkage map of
barleychromosome7,adaptedfrom Ku ¨nzel et
al. (2000), is depicted on the right. The step-
onal lines were used in wheat group 7 to differ-
entiate among the regions.
loci in distal regions was correlated with a higher rate
of recombination. Conversely, a lower density of EST
loci in proximal regions was correlated with a lower rate
of recombination (Akhunov et al. 2003a). Thus, the
majority of genes in wheat appear to be located in the
high-recombination areas, allowing for effective devel-
opment and use of map-based cloning strategies to
clone genes of interest.
Consensus map: Because many ESTs detected three
orthologousloci amongthethree homoeologousgroup
7 chromosomes, and the loci appear to be colinear,
it was possible to construct a consensus chromosome
deletion bin map of homoeologous group 7. The con-
sensus map provides a detailed resolution of the relative
positions of mapped orthologous loci. Because of the
colinearity of these loci across the chromosomes of
group 7 and a lack of duplication anywhere else in the
wheat genome, we identified these 166 loci as landmark
markers for this homoeologous group.
Landmark loci, presented here as EST clones, could
correspond to highly conserved regions and could be
of orthologous genes. These regions may be of signifi-
cance in understanding genome evolution among Triti-
ceae species by analyzing chromosome structural re-
arrangements, recombination hot spots, suppression of
recombination, and gene distribution, duplication, and
elimination events in the genome. Hohmann et al.
(1995) designated 10 landmark RFLP loci, 5 each for
the short and long arms of consensus chromosome 7.
They suggested that these loci could be useful in tar-
geting specific genes to specific regions of consensus
chromosome 7. In our study 117 loci were colinear
across the homoeologous group, and 68% (81/117)
were mapped into the region close to the centromere.
These loci mapped to the proximal region are possibly
conserved over large evolutionary distances and could
be linked to, or possibly represent, critical genes that
necessitated their presence in the genomes during the
establishment of polyploid species (Akhunov et al.
2003b). Because of their evolutionary importance we
believe that these 117 loci should be present in closely
696K. G. Hossain et al.
Putative order of wheat ESTs based on corresponding rice BAC/PAC and genetic markers
(cM) in rice
Short arm of wheat consensus chromosome 7
Long arm of wheat consensus chromosome 7
aThe intervals of each consensus chromosome arm are presented from proximal to distal.
Duplication: There are duplicated loci (paralogous)
on almost all of the RFLP linkage and physical maps of
Hohmann et al. 1994; Nelson et al. 1995; Marino et al.
1996; Weng et al. 2000; Weng and Lazar 2002). In a
mapping study of the T. monococcum L. genome, Dub-
mosomal duplications. In our study, 24% of the ESTs
mapping to group 7 identified duplicated loci either
on the chromosomes of other homoeologous groups
or on the same arm or different arms of chromosomes
loci in the same deletion bin (intrabin) and these could
have resulted from the internal cut site of EcoRI within
a locus. The observed rate of duplication does not re-
flect the total duplicated loci of the wheat genome since
we mapped only the chromosomes of homoeologous
group 7. In an effort to map physically 6421 ESTs in
the rice genome, Wu et al. (1998) reported only 2.4%
duplicated loci. Hence, there appears to be an order
of magnitude more of duplicated loci per gene motif
within the homoeologous group 7 chromosomes of
wheat than in the small genome of rice. The growth or
shrinkage of the plant genomes has been attributed
697Physical Map of EST Clones in Wheat Homoeologous Group 7
to the growth and shrinkage of repeated nucleotide
sequences (Bennetzen 2002; Sanmiguel et al. 2002);
however, the growth of the wheat genomes also appears
to have been accompanied by the concomitant accumu-
lation of dispersed gene duplications. The similar pro-
portions of duplicated loci to overall genome size of
wheat and rice suggest that the accumulation or dele-
tion of repeated sequences and genes could have been
coupled and controlled by a common mechanism.
The duplicated loci tend to be located in the distal
regions of chromosome arms (Figure 3), whereas the
landmark loci were mostly proximal (Figure 2). The
was highly correlated with the recombination rates
along hexaploid wheat chromosome arms and along
chromosome arms in diploid species of the Triticum-
Aegilops alliance (Dvor ˇa ´k et al. 1998; Akhunov et al.
2003a). This relationship has been attributed to either
selection or linkage of neutral loci to mildly deleterious
genes not favored by natural selection (Charlesworth
1994). In both scenarios, there is a greater chance for
a neutral locus and, by extension, for polymorphism for
a duplicated locus, to be eliminated if it is in a low-
recombination region than if it is in a high-recombina-
tion region. Therefore, it could be suggested that poly-
morphisms for neutral locus duplications are expected
to survive and become fixed preferentially in high-
recombination regions. Identification of duplicated re-
gions between homoeologous group 7 and homoeolo-
gous groups 3, 5, and 6 of wheat implied that these
duplications existed prior to polyploidization (Qi et al.
7BS ? 4AL and 5AL ? 7BS translocation: On the
basis of the location of structural genes on chromo-
somes 4BL, 4DL, and 5AL (Ainsworth et al. 1983),
and of endosperm peroxidase on 4AL, 7AS, and 7DS
(Kobrechel and Fillet 1975), Naranjo et al. (1987)
proposed a double translocation, 4AL to 5AL, 5AL to
7BS, and 7BS to 4AL, in the genome of Chinese Spring
wheat. Anderson et al. (1992) analyzed these transloca-
tions by RFLP analysis using genomic probes and sup-
ported the translocations proposed by Naranjo et al.
(1987). Werner et al. (1992) reported a segment of the
short arm of chromosome 7B had been translocated to
the long arm of chromosome 4A and suggested that
?20% of the distal region of the 4AL chromosome was
derived from a translocation of 7BS. In the present
study, we identified loci corresponding to 44 probes
mapped to bins 7AS1-0.89-1.00, 7AS5-0.59-0.89, 7DS4-
0.61-1.00, 4AL4-0.80-1.00, 4AL5-0.66-0.80, and 4AL13-
0.59-0.66 (Figure 2B). Even ifwe consider that a portion
4AL13-0.59-0.66wasmapped bythelociof theseprobes,
at least 34% of the 4AL chromosome arm at the distal
region was derived from a distal translocation event
location between 5AL and 7BS by assigning a 5AL-spe-
cific fragment ofthe probe BCD87 tochromosome 7BS.
We examined the probes mapped to 7BS and identified
loci corresponding to five probes (Figure 2B) that
mapped distal on 7BS, 5BL, and 5DL. We could not
Our analysis supported the proposed translocation be-
involved in this translocation appears much smaller
than the translocation between 7BS and 4AL, which is
in agreement with Jiang and Gill (1994).
Ordering EST loci and comparison of map position
with rice and barley: Homoeology between wheat and
rice genomes was first studied by Ahn et al. (1993) fol-
lowed by Kurata et al. (1994) and Van Deynze et al.
(1995) at the macro level. Sorrells et al. (2003) com-
pared rice and wheat genomes at the micro/DNA se-
quence level. All those studies indicated that rice chro-
mosomes 6 and 8 are homoeologous with Triticeae
group 7 chromosomes. Of the 117 ESTs located on the
group 7 consensus map, 38 were located to rice BAC/
PACs with corresponding genetic markers, and 11 of
the BAC/PACs correspond with the sequence of RFLP
markers mapped to chromosome 7 of barley (Table 3
and Figure 4). The terminal regions (100.4–118.9 cM)
of the long arm of rice chromosome 8 corresponded
with the centromeric region (0.0–0.59) of the long arm
of consensus chromosome 7 of wheat (Figure 4). The
short arm region (13.7–39.7 cM) of rice chromosome
8 corresponded with the centromeric region of the long
arm (0.0–0.39) of consensus chromosome 7 of wheat.
About 39% of the distal region of the long arm of con-
sensus chromosome 7 corresponded with a 26-cM re-
a putative homoeologous relationship of genes involved
in these regions of wheat and rice. The present study
between the wheat group 7 consensus chromosome and
rice chromosomes 6 and 8. Orthology of these loci with
rice suggests a possible ancestral origin of these loci and
and rice lineages. Hence the proximal low-recombina-
tion region of wheat chromosomes could be a region
the findings of Akhunov et al. (2003a) and Sorrells
et al. (2003). Using the rice genome as a template one
can predict colinearity with the wheat genomes; how-
ever, microsynteny studies have suggested that, in most
cases, colinearity will need to be verified at the DNA
sequence level (Han et al. 1999; Bennetzen and Rama-
krishna 2002). The ordering of mapped ESTs within
chromosome bins would be an important enhancement
for the wheat/rice comparative analysis.
Although the wheat homoeologous group 7 map is
based on consensus physical maps that combine dele-
698 K. G. Hossain et al.
Bennett, M. D., and J. B. Smith, 1976
angiosperms. Trans. R. Soc. Lond. Ser. B Biol. Sci. 274: 227–274.
Science 296: 60–63.
Bennetzen, J. L., and W. Ramakrishna, 2002
arrangements of gene content, order and orientation differenti-
ate grass genomes. Plant Mol. Biol. 48: 821–827.
ificity. Curr. Biol. 4: 545–546.
Covitz, P. A., L. S. Smith and S. R. Long, 1998
tags from a root-hair-enriched Medicago truncatula cDNA library.
Plant Physiol. 117: 1325–1332.
Delaney, D., S. Nasuda, T. R. Endo, B. S. Gill and S. H. Hulbert,
somes of wheat. Theor. Appl. Genet. 91: 568–573.
Delaney, D., S. Nasuda, T. R. Endo, B. S. Gill and S. H. Hulbert,
somes of wheat. Theor. Appl. Genet. 91: 780–782.
Devos, K. M., J. Dubcovsky, J. Dvor ˇa ´k, C. N. Chinoy and M. D.
Gale, 1995Structural evolution of wheat chromosomes 4A, 5A,
and 7B and its impact on recombination. Theor. Appl. Genet.
Dubcovsky, J., M.-C. Luo, G.-Y. Zhong, R. Bransteiter, A. Desai
et al., 1996Genetic map of diploid wheat, Triticum monococcum
L., and its comparison with maps of Hordeum vulgare L. Genetics
Dvor ˇa ´k,J., andH.B. Zhang,1990
sequence sheds light on the phylogeny of the wheat B and G
genomes. Proc. Natl. Acad. Sci. USA 87: 9640–9644.
Dvor ˇa ´k, J., M. C. Luo and Z. L. Yang, 1998
length polymorphism and divergence in the genomic regions of
Aegilops species. Genetics 148: 423–434.
Endo, T. R., 1988Induction of chromosome structural changes by
a chromosome of Aegilops cylindrical L. in common wheat. J.
Hered. 79: 366–370.
Endo, T. R., 1990 Gametocidal chromosomes and their induction
of chromosome mutations in wheat. Jpn. J. Genet. 65: 135–152.
Endo, T. R., and B. S. Gill, 1996
wheat. J. Hered. 87: 295–307.
suppression gene of Triticum aestivum L. based on partial deletion
in the long arm of chromosome 5A. Jpn. J. Genet. 69: 13–19.
Endo, T. R., Y. Mukai, M. Yamamoto and B. S. Gill, 1991
mapping of male-fertility gene of common wheat. Jpn. J. Genet.
Ewing, R. M., A. B. Kahla, O. Poirot, F. Lopez, S. Audic et al.,
1999 Large-scale statistical analyses of rice ESTs reveal corre-
lated patterns of gene expression. Genome Res. 9: 950–959.
Faris, J. D., K. M. Haen and B. S. Gill, 2000
of a gene-rich recombination hot spot region in wheat. Genetics
Fernandes, J., V. Brendel, X. Gai, S. Lal and V. L. Chandler, 2002
ComparisonofRNA expressionprofilebasedon maizeexpressed
sequence tag frequency analysis and micro-array hybridization.
Plant Physiol. 128: 896–910.
Gill, B. S., B. Friebe and T. R. Endo, 1991
nomenclature system for description of chromosome bands and
structural aberrations in wheat (Triticum aestivum). Genome 34:
Gill, K. S., B. S. Gill and T. R. Endo, 1993
specific mapping strategy reveals gene-rich telomeric ends in
wheat. Chromosoma 102: 374–381.
Gill, K. S., B.S. Gill, T. R. Endo andE. V. Boyko, 1996a
tion and high-density mapping of gene-rich regions in chromo-
some group 5 of wheat. Genetics 143: 1001–1012.
Gill, K. S., B. S. Gill, T. R. Endo and T. Taylor, 1996b
tion and high-density mapping of gene-rich regions in chromo-
some group 1 of wheat. Genetics 144: 1883–1891.
Han, F., A. Kilian, J. P. Chao, D. Kudrna, B. Steffenson et al., 1999
Sequence analysis of a rice BAC covering the syntenous barley
Rpg1 region. Genome 42: 1071–1076.
Hillier, L., G. Lennon, M. Becker, M. F. Bonaldo, B. Chiapelli
sequence tags. Genome Res. 6: 807–828.
Hohmann, U., T. R. Endo, K. S. Gill and B. S. Gill, 1994
Nuclear DNA amounts in
tions from 7A, 7B, and 7D chromosomes that differ
in size and in amount of heterochromatin, the results
obtained by comparing themto the barley chromosome
7 linkage map are in good agreement with those re-
ported by Werner et al. (1992), Hohmann et al. (1995),
and Ku ¨nzel et al. (2000). Except for the reverse order
of four ESTs mapped into the 0.45–0.59 regions on the
group 7 consensus map of wheat, the relative positions
chromosome 7 and barley chromosome 7 (Figure 4).
The reverse order of these ESTs corresponding to the
tified a minor inversion of the similar region of group
7 chromosomes that has been reported by Hohmann
et al. (1995). Thus, these results suggest the ordering
chromosomes of homoeologous group 7 with chromo-
somes of rice and barley.
Thelocalization anddistribution ofEST lociinto bins
reflects the distribution of genes and gene-rich regions
of this group in wheat. Identification of landmark
probes and putative map positions in rice and barley
genomes suggested a detailed analysis of ESTs mapped
in the wheat genome could provide valuable informa-
tion in mapping and identification of genes across grass
Numerous small re-
We thank S. S. Maan for his valuable guidance throughout this
project. This material is based upon work supported by the National
Science Foundation under cooperative agreement no. DBI-9975989.
The deletion stocks of common
Adams, M. D., J. M. Kelley, J. D. Gocayne, M. Dubnick, M. H.
Polymeropoulos et al., 1991
Adams, M. D., A. R. Kerlavage, R. D. Fleischmann, R. A. Fuldner,
C. J. Bult et al., 1995 Initial assessment of human gene diversity
and expression patterns based upon 83 million nucleotides of
cDNA sequence. Nature 377: 3–17.
Ahn, S. N., and S. D. Tanksley, 1993
of the rice and maize genomes. Proc. Natl. Acad. Sci. USA 90:
Ahn, S. N., J. A. Anderson, M. E. Sorrells and S. D. Tanksley,
1993Homoeologous relationships of rice, wheat and maize
chromosomes. Mol. Gen. Genet. 241: 483–490.
Ainsworth, C. C., M. D. Gale and S. Bird, 1983
?-amylase isozymes in wheat. I. Allelic variation among hexaploid
varieties and intrachromosomal gene locations. Theor. Appl.
Genet. 66: 39–49.
Akhunov, E. D., A. W. Goodyear, S. Geng, L. L. Qi, B. Echalier
et al., 2003aThe organization and rate of evolution of wheat
genomes are correlated with recombination rates along chromo-
some arms. Genome Res. 13: 753–763.
Akhunov, E. D., A. R. Akhunova, A. M. Linkiewicz, J. Dubcovsky,
D. Hummel et al., 2003b Synteny perturbations between wheat
homoeologous chromosomes caused by locus duplications and
deletions correlate with recombination rates. Proc. Natl. Acad.
Sci. USA 100: 10836–10841.
Anderson, J. A., Y. Ogihara, M. E. Sorrells and S. D. Tanksley,
1992Development of a chromosomal arm map for wheat based
on RFLP markers. Theor. Appl. Genet. 83: 1035–1043.
Complementary DNA sequenc-
Comparative linkage maps
Standard karyotype and
The genetics of
A chromosome region-
699 Physical Map of EST Clones in Wheat Homoeologous Group 7
son of genetic and physical maps of group 7 chromosomes from
Triticum aestivum L. Mol. Gen. Genet. 245: 644–653.
Hohmann, U., A. Graner, T. R. Endo, B. S. Gill and R. G. Herr-
mann, 1995Comparison of wheat physical maps with barley
linkage maps for group 7 chromosomes. Theor. Appl. Genet. 91:
Jiang, I. P., and B. S. Gill, 1994
and plant genome mapping: the first 10 years. Genome 37: 717–
Kihara, H., 1954Consideration on the evolution and distribution
of Aegilops species based on the analyzer-method. Cytologica 19:
Kleinhofs, A., A. Kilian, M. A. Saghai Maroof, R. M. Biyashev,
P. Hayes et al., 1993 A molecular, isozyme and morphological
map of barley (Hordeum vulgare) genome. Theor. Appl. Genet.
Kobrechel, K., and P. Fillet, 1975
chromosomes involved in peroxidase synthesis of wheat seeds.
Can. J. Bot. 53: 2326–2344.
Kota, R.S., K. S.Gill, B.S. Gill andT. R. Endo,1993
cally based physical map of chromosome 1B in common wheat.
Genome 36: 548–554.
Ku ¨nzel, G., L. Korzun and A. Meister, 2000
grated physical restriction fragment length polymorphism maps
of barley genome based on translocation breakpoints. Genetics
Kurata, N., G. Moore, Y. Nagamura, T. Foote, M. Yano et al.,
1994 Conservation of genomic structure between rice and wheat.
Bio/Technology 12: 276–278.
Lazo, G. R., S. Chao, D. D. Hummel, H. Edwards, C. C. Crossman
et al., 2004 Development of an expressed sequence tag (EST)
resource for wheat (Triticum aestivum L.): unigene analysis, probe
selection and bioinformatics for a16,000-locus bin map. Genetics
Marino, C. L., J. C. Nelson, Y. H. Lu, M. E. Sorrells, P. Leroy et
al., 1996 Molecular genetic maps of the group-6 chromosomes
of hexaploid wheat (Triticum aestivum L). Genome 39: 359–366.
Marra, M., L. Hillier, T. Kucaba, M. M. Allen, R. Barstead et al.,
McFadden, E. S., and E. R. Sears, 1946
spelta and its free threshing hexaploid relatives. J. Hered. 37:
McIntosh, R. A., Y. Yamazaki, K. M. Devos, J. Dubcovsky, W. J.
Rogers et al., 2003 Catalogue of gene symbols for wheat, Vol.
4, pp. 1–34 in Proceedings of the 10th International Wheat Genetics
Symposium, edited by N. E. Pogna, M. Romano, E. Pogna and
G. Galterio. Instituto Sperimentale per la Cerealicotura, Rome.
ladder map of the wheat homoeologous group-4 chromosomes.
Theor. Appl. Genet. 90: 1007–1011.
Naranjo, T., P. Roca, P. G. Goicoechea and R. Giraldez, 1987
Arm homoeology of wheat and rye chromosomes. Genome 29:
Nelson, J. C., M. E. Sorrells, A. E. Van Deynze, Y. H. Lu, M.
and rearrangements in homoeologous groups 4, 5, and 7. Genet-
ics 141: 721–731.
Nishikawa, K., 1983
ancestors as viewed from isozyme variation, pp. 59–63 in Proceed-
ings of the 6th International Wheat Genetics Symposium, edited by S.
Sakamoto. Plant Germplasm Institute, Faculty of Agriculture,
Kyoto University, Kyoto, Japan.
Qi, L. L., B. Echalier, B. Friebe and B. S. Gill, 2003
characterization of a set of wheat deletion stocks for using in
chromosome bin mapping of ESTs. Funct. Integr. Genomics 3:
Sandhu, D., J. A. Champoux, S. N. Bondareva and K. S. Gill,
markers to a major gene-rich region on wheat 1S chromosomes.
Genetics 157: 1735–1747.
Sanmiguel, P., W. Ramakrishna, J. L. Bennetzen, C. S. Busso and
J. Dubcovsky, 2002Transposable elements, genes and recom-
bination in a 215-kb contig from wheat chromosome 5A. Funct.
Integr. Genomics 2: 70–80.
Scheetz, T. E., M. R. Raymon, D. Y. Nishimura, A. McClain, C. W.
Roberts et al., 2001 Generation of a high-density rat EST map.
Genome Res. 12: 648–655.
Sears, E. R., 1954The Aneuploids of Common Wheat. Bull. 572,Univer-
sity of Missouri Agricultural Experiment Station, Columbia, MO.
Sears, E. R., 1966Nullisomic-tetrasomic combinations in hexaploid
wheat, pp. 29–45 in Chromosome Manipulation and Plant Genetics,
edited by R. Riley and K. R. Lewis. Oliver & Boyd, Edinburgh.
Sears, E. R., and L. M. S. Sears, 1978
Wheat GeneticsSymposium, editedby S.Ramanujam. IndianSociety
for Genetics and Plant Breeding, New Delhi, India.
Sorrells, M. E., M. La Rota, C. E. Bermudez-Kandianis, R. A.
Greene, R. Kantety et al., 2003
analysis of wheat and rice genomes. Genome Res. 13: 1818–1827.
Tanksley, S. D., M. W. Ganal, J. P. Price, M. C. DeVicente, M. W.
Bonierbale et al., 1992High density molecular maps of tomato
and potato genomes; biological inferences and practical ap-
proaches. Genetics 132: 1141–1160.
Van Deynze, A. E., J. C. Nelson, E. S. Yglesias, S. E. Harrington,
D. P.Braga et al.,1995 Comparative mapping ingrasses. Wheat
relationships. Mol. Gen. Genet. 248: 744–754.
Werner,J.E., T.R.Endoand B.S.Gill,1992
cally based physical map of the wheat genome. Proc. Natl. Acad.
Sci. USA 89: 11307–11311.
Weng, Y., and M. D. Lazar, 2002
group-6 short arm physical maps of wheat and barley reveals a
similar distribution of recombinogenic and gene-rich regions.
Theor. Appl. Genet. 104: 1078–1085.
Weng,Y., N.A. Tuleenand G.E.Hart,2000
and a consensus physical map of the homoeologous group-6
chromosomes of wheat (Triticum aestivum L.). Theor. Appl. Genet.
Wu, J. Z., N. Kurata, H. Tanoue, T. Shimokawa, Y. Umehara et al.,
1998Physical mapping of duplicated genomic regions of two
chromosome ends in rice. Genetics 150: 1595–1603.
Zhang, D., D. W. Choi, S. Wanamaker, R. D. Fenton, A. Chin et
al., 2004Construction and evaluation of cDNA libraries for
large-scale expressed sequence tag sequencing in wheat (Triticum
aestivum L.). Genetics 168: 595–608.
Species relationship of wheat and its putative
Nonisotopic in situ hybridization
Identification of genomes and
The telocentric chromosomes
Comparative DNA sequence
The origin and of Triticum
Comparison of homoeologous
Communicating editor: J. P. Gustafson