BMC Genomics BioMed Central

Research article Open Access

DArT markers: diversity analyses and mapping in Sorghum bicolor
Emma S Mace†1, Ling Xia†2, David R Jordan1, Kirsten Halloran1,
Dipal K Parh3,4, Eric Huttner2, Peter Wenzl2 and Andrzej Kilian*2

Address: 1The Department of Primary Industries & Fisheries, Queensland (DPI&F), Hermitage Research Station, Warwick, QLD 4370, Australia,
2Diversity Arrays Technology P/L, PO Box 7141, Yarralumla ACT 2600, Australia, 3School of Land and Food Sciences, University of Queensland,

Brisbane, QLD 4072, Australia and 4Bureau of Sugar Experiment Station, DNRP, 50 Meiers Road, Indooroopilly, QLD 4068, Australia
Email: Emma S Mace - emma.mace@dpi.qld.gov.au; Ling Xia - ling@diversityarrays.com; David R Jordan - david.r.jordan@dpi.qld.gov.au;
Kirsten Halloran - kirsten.halloran@dpi.qld.gov.au; Dipal K Parh - d.parh@bses.org.au; Eric Huttner - e.huttner@diversityarrays.com;
Peter Wenzl - p.wenzl@diversityarrays.com; Andrzej Kilian* - a.kilian@diversityarrays.com
* Corresponding author †Equal contributors

Published: 22 January 2008 Received: 28 June 2007

Accepted: 22 January 2008
BMC Genomics 2008, 9:26 doi:10.1186/1471-2164-9-26
This article is available from: http://www.biomedcentral.com/1471-2164/9/26
© 2008 Mace et al; licensee BioMed Central Ltd.
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0),
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract
Background: The sequential nature of gel-based marker systems entails low throughput and high
costs per assay. Commonly used marker systems such as SSR and SNP are also dependent on
sequence information. These limitations result in high cost per data point and significantly limit the
capacity of breeding programs to obtain sufficient return on investment to justify the routine use
of marker-assisted breeding for many traits and particularly quantitative traits. Diversity Arrays
Technology (DArT™) is a cost effective hybridisation-based marker technology that offers a high
multiplexing level while being independent of sequence information. This technology offers
sorghum breeding programs an alternative approach to whole-genome profiling. We report on the
development, application, mapping and utility of DArT™ markers for sorghum germplasm.
Results: A genotyping array was developed representing approximately 12,000 genomic clones
using PstI+BanII complexity with a subset of clones obtained through the suppression subtractive
hybridisation (SSH) method. The genotyping array was used to analyse a diverse set of sorghum
genotypes and screening a Recombinant Inbred Lines (RIL) mapping population. Over 500 markers
detected variation among 90 accessions used in a diversity analysis. Cluster analysis discriminated
well between all 90 genotypes. To confirm that the sorghum DArT markers behave in a Mendelian
manner, we constructed a genetic linkage map for a cross between R931945-2-2 and IS 8525
integrating DArT and other marker types. In total, 596 markers could be placed on the integrated
linkage map, which spanned 1431.6 cM. The genetic linkage map had an average marker density of
1/2.39 cM, with an average DArT marker density of 1/3.9 cM.
Conclusion: We have successfully developed DArT markers for Sorghum bicolor and have
demonstrated that DArT provides high quality markers that can be used for diversity analyses and
to construct medium-density genetic linkage maps. The high number of DArT markers generated
in a single assay not only provides a precise estimate of genetic relationships among genotypes, but
also their even distribution over the genome offers real advantages for a range of molecular
breeding and genomics applications.

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Background
Sorghum is a major staple food and fodder crop grown
worldwide, with an annual average production of 61 mil-
lion tonnes over the past decade [1]. The crop is tolerant
of many biotic and abiotic stresses and is often grown in
more marginal cropping areas. In developing countries it
tends to be a staple food and forage of the poor. In devel-
oped countries it is used primarily as an animal feed. Sor-
ghum is often preferentially grown in both situations as it
is better adapted to water limited environments than
other cereal crops.
Investment in sorghum breeding and genomic resources
has been less than for the other major cereals rice, wheat,
maize and barley. Interest has focused on the crop due to
its drought resistance and small genome size (~760 Mb)
compared to close relatives maize (~2500 Mb) and sugar-
cane (2550 to 4200 Mb) [2-4]. In recent years the poten-
tial of sorghum as a biofuel crop has led to additional
investment culminating in the sequencing of the sorghum
genome [5].
Numerous studies have demonstrated that sorghum is
very diverse crop, with cultivated sorghums exhibiting
great phenotypic variability. The cultivated germplasm
has been classified into five major races (bicolor, cauda-
tum, durra, guinea and kafir) and 10 intermediate races
based on panicle and spikelet morphology [6]. In order to
exploit this diversity at the genotypic level, an efficient
marker system is required. Many molecular marker tech-
nologies have been developed and applied to studying
patterns of genetic diversity in sorghum germplasm col-
lections and in breeding programs, including RFLPs [7-9],
RAPDs [10,11], ISSRs [12], SSRs [13-19] and AFLPs
[17,20]. However, the major limitation to the widespread
use of current marker technologies in applied sorghum
breeding programs and germplasm collections is the high
cost per data point. Applications that require whole
genome scans such as pedigree analysis [21], association
mapping [22] and mapping as you go (MAYG) [23], or
large scale genotyping of germplasm collections [24] are
not cost effective using current technologies.
The current molecular marker technologies have charac-
teristics which additionally affect the level of genome cov-
erage, their discrimination ability, reproducibility and
technical and time demand. A number of the limitations
associated with the different marker technologies can be
overcome by utilising specialised hardware such as high
throughput capillary electrophoresis machines, which can
impact on discrimination ability, reproducibility and
speed. However, the majority of the limitations are related
to the sequential nature and high assay costs of the marker
technologies, in addition to reliance on DNA sequence
information. Diversity arrays technology (DArT) can over
http://www.biomedcentral.com/1471-2164/9/26
come these limitations and has been developed as a
hybridisation-based alternative to the majority of gel-
based marker technologies currently in use. The DArT gen-
otyping method was developed originally for rice [25] and
has subsequently been applied to many other plant spe-
cies, including barley [26], cassava [27], Arabidopsis [28],
pigeonpea [29] and wheat [30]. DArT has been also
applied to a number of animal species and microorgan-
isms [31]. The DArT methodology offers a high multiplex-
ing level, being able to simultaneously type several
thousand loci per assay, while being independent of
sequence information. DArT assays generate whole-
genome fingerprints by scoring the presence versus
absence of DNA fragments in genomic representations
generated from genomic DNA samples through the proc-
ess of complexity reduction.
This paper reports the results of a study designed to (1):
develop a sorghum diversity array for DArT genotyping,
(2): determine linkage map positions of polymorphic
DArTs and (3): assess utility of DArT technology in diver-
sity analyses on a set of diverse sorghum lines, including
selected lines from the Department of Primary Industries
and Fisheries (DPI&F) sorghum breeding program. In
order to evaluate the discriminatory ability of DArTs,
efforts have been made to include the same genotypes
used in genetic diversity studies based on other molecular
marker technologies.
Results
Evaluation of complexity reduction methods and array
development
The initial tests of DArT performance in sorghum were
done on eight sorghum genotypes (Additional File 1).
Based on our positive experience with PstI-based genomic
representations [25] we initially evaluated several combi-
nations of PstI with different frequently cutting restriction
enzymes (RE) as a complexity reduction approach for sor-
ghum. DNA samples from the eight sorghum genotypes
were digested with PstI and several frequently cutting RE
(PstI+TaqI, PstI+MseI, PstI+ApoI, PstI+AluI, PstI+BanII,
PstI+BstNI and PstI+AflIII), ligated to a PstI adapter and
then amplified with the PstI-0 primer. Gel analysis of the
PCR products showed that a uniform smear (without
major bands) only appeared in the PstI+BanII combina-
tion. Other RE combinations gave a smear with one or
more dominant bands. Such strong bands represent
highly amplified restriction fragments and correspond to
abundant repetitive sequences in the representation; a fea-
ture which is highly detrimental to DArT performance
[32].
AFLP-like analysis was performed to estimate the frag-
ment number in the PstI+BanII representation following
the approach utilized by Xia et al. [27]. Four primers con-
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BMC Genomics 2008, 9:26
taining 3 selective bases at the 3'end were used for the
amplification. For all four primers, a large number of frag-
ments were visible on the gel, making a precise estimate of
the total number of fragments impossible (data not pre-
sented). Interestingly, a similar approach resulted in an
easily quantifiable number of bands in cassava which has
a similar genome size to sorghum and even in barley,
which has genome size almost seven times larger than sor-
ghum. The results suggest that the PstI+BanII representa-
tion in sorghum has a larger number of unique fragments
than the PstI+BstNI representation in barley which was
reported to contain 1,546 markers placed on the inte-
grated map of barley genome [26].
Based on the extended PstI+BanII sub-libraries and the
additional libraries generated based on the genomic sub-
traction (SSH) method [33], the best DArT markers from
the initial experiments were "cherry picked" and a "re-
array library" with 768 polymorphic clones was created.
In addition to these 768 polymorphism-enriched clones,
a further 5367 clones from PstI+BanII Library C were used
for all genotyping work reported here.
Genetic relationships between sorghum lines revealed by
DArT
The selected DArT clones were tested for their ability to
resolve genetic relationships among a set of 90 lines. The
reproducibility of the DArT genotyping array was success-
fully validated by independent assays from the same
DNA. The genotypes selected represent a significant pro-
portion of the genetic variation in sorghum with all 5
races represented and 5 intermediate races also repre-
sented. In addition, the germplasm set included elite lines
from breeding programs some of which had high levels of
co-ancestry.
DArTsoft analysis (see Materials and Methods) identified
508 markers polymorphic among 90 genotypes typed on
the array. The PIC values of these 508 markers were very
high with over 69% of the markers having a PIC value
between 0.4 and 0.5 (Table 1). The average PIC was 0.41,
higher than the previous DArT studies in barley (0.38
[26]) and comparable with cassava (0.42 [27]). The rela-
tionship between the quality of the DArT markers (meas-
ured as the % of total variance which existed between the
Table 1: Polymorphism Information content (PIC) values for 508
DArT markers
PIC value # DArTs % DArTs
0.5-0.4 353 69.5
0.4-0.3 82 16.1
0.3-0.2 46 9.1
0.2-0.1 21 4.1
0.1-0 6 1.2
http://www.biomedcentral.com/1471-2164/9/26
two clusters: present and absent) and the performance of
the DArT markers as determined through call rate and PIC
was analysed (Table 2). The PIC values were largely inde-
pendent of marker quality, with only a small reduction in
PIC value observed in the lowest quality class (0.40 in the
lowest quality class vs. 0.44 and 0.42 for the two higher
quality classes). As expected, the average call rate
decreased with average Q value. The markers with the
highest Q values (above 90% of total variance between
the clusters) had very high average call rates (98%), while
the markers in the lower quality marker classes had lower
average call rates and higher standard deviations.
Cluster analysis based on the DICE dissimilarity index
and the unweighted neighbour-joining method was per-
formed on the 508 DArT markers for 90 genotypes (Figure
1). This cluster analysis discriminated well between all 90
genotypes and has a cophenetic correlation value of
0.9308, indicating an excellent fit of the similarity matrix
data to the tree topology. Thirteen main clusters were
identified which correspond well with race and origin of
the genotyped lines. In particular a single predominant
race or origin could be identified in 9 out of the 13 clus-
ters. Cluster 1 contained 13 genotypes in total, of which
11 were kafir or kafir-caudatum. Cluster 4 contained 9
genotypes, of which 5 were of race durra. Additionally,
two Ethiopian durra types (IS 12555C and B35) grouped
together within this cluster with a boot strap value of
100%. Cluster 5 consisted of 6 Chinese genotypes of com-
plex racial background. Cluster 6 contained the wild spe-
cies, S. propinquum and the weedy subspecies, S. bicolor
subsp. verticilliflorum (formerly S. arundinaceum). Cluster
9 consisted of 2 caudatum genotypes (IS 12656C and IS
10302). Cluster 10 consisted of 11 genotypes in total, of
which 8 were caudatum or caudatum-derived. Cluster 11
consisted of a tight cluster of restorer (R) lines predomi-
nately from Australian breeding programs; R9990066,
R999017 and R999003 are all progeny of the line
R31945-2-2. Cluster 12 is a looser cluster consisting of 15
lines of which 7 were caudatum or caudatum-derived.
Finally cluster 13 consisted of 4 genotypes of which 2 are
bicolor or bicolor intermediate and S. bicolor subsp. drum-
mondii which is very similar morphologically to the
bicolor race.
Table 2: The relationship between the quality and the
performance of the DArT markers
100 > Q > 90 90 > Q > 80 80 > Q > 70
Number of markers 37 236 235
Call Rate 98.03 ± 1.69 92.75 ± 3.51 88.01 ± 4.83
PIC 0.44 ± 0.08 0.42 ± 0.09 0.40 ± 0.11
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B Tx 3197
70
68 B OK 11
41 D warf R edlan
SD S 1948-3
48
27 IS 8525
B Tx 398
64 67
B Tx 399
98
RTx 7000
B Tx 3042
C1
21 B Tx 623
IS 3151
62
67 R io
IS 3511
R9403463-2-1
54
44 QL41
22 QL12
R 9188
4

23
IS 12661
Wray
C2
1 IS 4546C
K aura (blac k glumes )
99
67 IS 1596

69
IS 22525

IS 1237
IS 2263 C3
90
98 M35-1
92 IS 8336
1
34
91
IS 12572C
K arper 669
IS 2403C
C4
96
IS 12555C
100
B 35
17 LR2659
94
75 A i4
54 LR 9198
100 LR 2528
0 10
100
LR2931-2
LR 2844 C5
S. propinquum
94
34 S. bic olor s ubs p. v ert ic illif lorum

2 51
R Tx 2536
296B
C6
84 R Tx 2895
48 R Tx 2737
IS 12543C
2
45
87
IS 3614-2
IS 17214
C7
R Tx 2903
2 IS 24756
97

7
IS 12656C
IS 10302
MLT 135
C8
79
46
K uy uma
MP531
C9
27 ICSV 400
93 IS 12568C
D orado
49 42
IS 12611C

69
48
81
IS 12663C
R S29
C10
IS 22457C
51 IS 11435
Mac ia
R Tx 430
21 100
28 R 931945-2-2
100 R 999066
R999017
23 99
100
44
R999003
QL36
C11
R 890562
TA M428
9 B 923171
100
94 B 923296
56 QL39
8 Green Leaf
32
IS 8337C
100
20 TA M422

9
LR2490-3
IS 12608C
K S 115 C12
46
2 SC 170-6-8
IS 2816C
29
2 18 Sureño
ICSV 745
H egari
64
IS 8163C
IS 12179C
79
S. bic olor s pp. drummondii
91
50
IS 11080C
IS 12648C
C13
0 0.1

Neighbor-joining
Figure 1 anlaysis of diverse sorghum genotypes based on 508 DArT markers using the DICE similarity coefficient
Neighbor-joining anlaysis of diverse sorghum genotypes based on 508 DArT markers using the DICE similarity
coefficient. 13 clusters have been defined. The numbers on the branches indicate bootstrap values (expressed in percentages;
based on 100 replications).

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Mapping of DArT loci markers were excluded. Interestingly, the difference in

To confirm that the sorghum DArT markers behave in a
Mendelian manner, we constructed a genetic linkage map
for a cross between R931945-2-2 and IS 8525. We selected
370 DArT markers with Q-values greater than 80% and
merged with the segregation data for 286 markers, consist-
ing of 55 SSRs, 229 AFLPs and 2 morphological markers
derived from the original map [34].
In total, 596 markers could be placed on the integrated
linkage map, which spanned 1431.6 cM (Table 3). The
genetic linkage map had an average marker density of 1/
2.39 cM, with an average DArT marker density of 1/3.9
cM. The 358 DArT markers included on the map mapped
to all 10 chromosomes and are distributed across the
genome in a similar way as the 47 SSR and 188 AFLP
markers (Figure 2), suggesting that the density of both
groups of markers roughly followed the distribution of
DNA polymorphism across the genome. The DArT mark-
ers accounted for approximately 50% of the framework
markers used in the initial construction of each linkage
group, with a high proportion of the DArT markers acting
as delegates (Table 3; see M&M for explanations of frame-
work, delegate and attached markers) indicating a higher
level of redundancy compared to the other marker types.
We therefore compared the level of redundancy, as deter-
mined through co-location, between the DArT markers
generated from the 6 subtraction libraries (SSH) versus
those developed from the extended PstI+BanII sub-librar-
ies. Of the 358 DArTs included on the map, 172 were SSH-
dervied and of these 50 (29%) were redundant, whereas
the 186 non-SSH derived DArTs exhibited a reduced level
of redundancy (26%). Overall redundancy in the map
dropped from 33.7% to 24% when SSH-derived DArT
Table 3: Summary of the genetic linkage map based on a cross between R31945-2-2 and IS 8525. The genetic linkage map was
constructed using DArTs, AFLPs and SSRs. The total length of each chromosome, the total number of markers, the total number of
DArTs and the number of framework, delegate and attached markers per chromosome are detailed. For the last three columns, the
number of DArTs in each class is given in parentheses
apparent redundancy levels between SSH-derived and
"normal" DArT markers was smaller compared to what we
observed for tomato and sugarcane (DArT P/L, unpub-
lished) and in fern species Asplenium and moss species
Garovaglia (DArT P/L and collaborators, unpublished
data).
There was no statistically significant difference between
DArT and non-DArT markers in the distribution of paren-
tal alleles across the genome. Twelve DArTs were removed
during the course of map construction either because of
lack of linkage to other markers, or as they formed small
linkage groups containing only DArTs which did not link
to the known chromosomes.
Discussion
Sorghum DArT markers
This is the first report of the use of DArT technology in sor-
ghum and our results demonstrate that the sorghum DArT
markers are high quality, as assessed by their call rate,
scoring reproducibility and PIC values. The DArT marker
quality parameters measured for the sorghum array are
comparable to those obtained for pigeonpea [29], barley
[26] and cassava [27] and wheat [30].
Utility of DArTs for diversity analysis
Among the criteria for genetic markers that are to be used
for fingerprinting and marker-assisted selection is a high
level of polymorphism [15]. Clearly, sorghum DArTs
meet this criterion, with over 69% of the 508 DArTs
revealing polymorphism between the set of 90 sorghum
lines having a PIC value between 0.5-0.4; 0.5 being the
highest PIC value expected for a bi-allelic marker system.

SBI Length (cM) Total # markers # DArTs # framework* # delegate* # attached*

1 188.1 94 65 31 (16) 25 (24) 38 (25)

2 135.6 48 31 31 (19) 9 (9) 9 (3)
3 83 31 19 21 (11) 4 (4) 6 (4)
4 133.9 70 49 28 (16) 18 (18) 24 (15)
5 130.4 81 43 38 (17) 18 (13) 25 (13)
6 157.1 61 23 36 (13) 9 (5) 16 (5)
7 120.5 40 24 26 (15) 5 (5) 9 (4)
8 184.5 75 51 41 (26) 14 (11) 20 (14)
9 149.3 40 19 24 (7) 9 (7) 7 (5)
10 149.2 56 34 26 (9) 6 (5) 25 (20)

Totals 1431.6 596 358 302 (149) 117 (101) 179 (108)

* 'Framework' markers are defined as those that could be ordered with a jack-knife value of 90% or greater using the MultiPoint software; Delegate
markers are defined as those that map to the same location as the representative framework marker for a specific locus; Attached markers are
those that initially were excluded from the framework map as they caused unstable neighborhoods but were included in the final, complete map by
assigning them to the best intervals on the framework map.

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SBI-01 SBI-02 SBI-03 SBI-04 SBI-05

0.0 sPb-0906
5.2 AAG+CAA1
11.7 Xtxp65
12.5 sPb-7047
0.0 AAG+CTA1 13.3 AG+CTG8
sPb-5611 sPb-0939 16.6 sPb-5256
9.1 sPb-4492 sPb-6160 0.0 sPb-6640 sPb-7308
19.9
sPb-5668 sPb-8237 18.5 AGG+CAG2 sPb-2941
31.9
12.0 sPb-0602 20.9 AGG+CAA5 AAG+CAT11 AGG+CAT3
13.0 AAG+CAG4 22.2 sPb-1932 sPb-6627 sPb-9815
13.9 Xtxp350 Xtxp325 23.5 sPb-6958 sPb-9580 sPb-2539 sPb-4131
39.7
15.3 sPb-5015 sPb-8773 0.0 sPb-2187 32.4 AG+CTG7 sPb-7638 sPb-3290
16.6 sPb-7096 7.6 AAG+CAT2 34.0 AGG+CAT4 sPb-1227 sPb-3977
17.3 sPb-7481 22.9 sPb-4195 42.2 Xtxp22 sPb-8881
45.3
64.0 AAG+CAC7 32.3 ACC+CAA7 0.0 sPb-6690 sPb-5618 sPb-2138
47.2 sPb-9544
sPb-2704 sPb-1694 35.9 Xtxp211 2.0 sPb-6982 sPb-2461 sPb-9635 sPb-4928 AG+CAT13
48.6
64.8 AG+CAT15 AGC+CTG4 38.8 sPb-4864 2.6 sPb-0892 46.3 sPb-2009 sPb-4039 sPb-5805
50.0
AAG+CAA2 41.7 sPb-9687 sPb-1127 5.7 sPb-5491 sPb-6559 sPb-8296 sPb-8316
50.4
65.5 sPb-1119 45.6 sPb-8647 6.4 AG+CTA6 sPb-2699 50.7 sPb-9009
71.8 sPb-8261 50.9 sPb-4453 8.5 sPb-4226 sPb-3343 sPb-7210 51.1 ACT+CAA4
83.3 sPb-6780 52.7 ACG+CAA1 9.2 AAC+CAG4 ACC+CAA3 sPb-6231 ACT+CAA5
51.5
87.7 sPb-0422 54.4 sPb-6693 22.5 sPb-6122 sPb-8521 sPb-9393 sPb-0932
57.1
88.5 sPb-0333 Xtxp88 56.8 ACT+CAT2 24.2 sPb-5108 46.6 sPb-1811 sPb-0258
62.0 AAG+CAA4
89.3 AAG+CTG1 58.2 AAG+CAC4 25.8 sPb-1294 sPb-1901 sPb-0543 sPb-8138
69.4 AGC+CAG1
sPb-5808 sPb-2070 59.5 Xtxp13 26.2 sPb-1137 Sb5-236 sPb-8378 sPb-1841
69.8 Xtxp15
sPb-9514 sPb-8794 67.6 sPb-1784 26.5 AAG+CTC6 sPb-5276 sPb-8972
89.8 70.2 AG+CTG6
sPb-2823 sPb-6061 sPb-4311 sPb-2639 28.4 sPb-0357 sPb-0703 sPb-1574
70.9 sPb-0738
AAC+CAC2 AG+CAT10 68.9 sPb-6700 sPb-1017 30.3 ACA+CAC1 46.8 sPb-0279 sPb-5530
sPb-3031 sPb-6508
sPb-0090 sPb-3386 sPb-2075 47.0 sPb-2246 sPb-5742 71.5
sPb-5194
sPb-6883 sPb-9050 74.3 sPb-5087 54.0 ACT+CAG1 55.2 AG+CTA4
90.2 72.2 AG+CAC2
sPb-0813 sPb-2058 75.1 sPb-1684 67.0 sPb-6882 sPb-4921 63.5 AGG+CAC2
72.4 sPb-3766
sPb-8947 AG+CAC4 75.9 Xtxp56 67.7 sPb-5267 68.1 AG+CTA8
72.5 AG+CTA2
92.4 sPb-0232 sPb-2544 76.7 ACC+CTC2 68.5 Xtxp285 72.7 XXtxp343 sPb-7534
73.3 AG+CAT6 sPb-0368
94.5 sPb-7229 sPb-9930 78.5 AG+CAC6 69.5 ACG+CAA4 73.1 AAG+CTA2
74.3 AAG+CTG4
96.8 sPb-0274 83.4 sPb-0073 70.7 sPb-7186 73.4 sPb-1147
75.2 AG+CTC1
99.1 AAC+CAC1 85.6 sPb-7702 71.8 sPb-9977 sPb-4629 77.7 sPb-4825
77.0 AG+CTA10
Xtxp37 sPb-3930 89.2 Xgap84 74.1 ACA+CAC2 79.0 sPb-9303
112.2 77.6 sPb-1501
sPb-0509 91.5 sPb-4444 74.3 AGC+CTG3 80.2 sPb-8013 sPb-6115
78.7 sPb-9347
124.5 ACC+CTC1 93.7 sPb-1925 74.5 AG+CTG4 81.8 Xgap10
79.7 sPb-8677
130.7 sPb-3801 sPb-9203 99.0 sPb-0549 83.0 AGG+CAA3 82.6 sPb-7324
80.0 sPb-6636
sPb-3103 sPb-4638 103.6 sPb-0971 sPb-9511 83.9 sPb-9995
133.9 sPb-6347 sPb-0205
sPb-7833 104.3 AAG+CAC6 87.7 sPb-9713 80.3
sPb-1104
137.1 sPb-7958 104.8 sPb-6663 Xtxp315 90.4 sPb-3838 sPb-4734
81.0 ACC+CTA5
150.1 sPb-4029 sPb-8132 111.5 sPb-4054 sPb-4473 sPb-4198 sPb-4806 sPb-5209
93.6
155.2 AGG+CAG1 113.2 sPb-7153 sPb-9304 AAG+CAG1 AAG+CTG8
160.2 Xtxp61 sPb-0277 114.8 sPb-1413 Xtxp207 94.0 sPb-8337 81.2 AG+CTT1 AGC+CTA3
sPb-7154 sPb-0383 124.1 AAG+CTG9 94.3 AGC+CAA2 AAG+CTG2
162.7 sPb-4593 AAC+CAT2 99.5 sPb-4070
127.1 sPb-5077 AGC+CAA3 AG+CAA3
165.2 AAC+CAA5 127.8 AGC+CTA4 AG+CTA9 105.2 Xtxp177 81.4 AAC+CAA3 AG+CAT3
168.4 sPb-0233 128.5 sPb-7647 sPb-8896 Xtxp41 ACA+CAA3 AAG+CTA3
106.6 Xtxp51
171.6 Xtxp319 135.6 sPb-7015 82.9 sPb-6604
174.1 sPb-0725 108.0 Xtxp60 sPb-8973
84.8
176.5 sPb-1984 126.0 Xtxp265 sPb-7203
86.6
sPb-8066 sPb-8275 126.7 ACA+CAG1 ACA+CAT2 sPb-6323 AAG+CAT10
177.8 127.3 sPb-7434 87.5
sPb-5527 sPb-5832 sPb-6855
178.7 sPb-5975 133.9 sPb-5550 88.1 AAG+CAT8
179.5 sPb-0494 93.1 sPb-3817
181.0 sPb-7331 sPb-6066 98.5 AG+CAA8
182.4 sPb-3861 AAG+CAA7 AGG+CAA7 AG+CTT4
183.5 sPb-4420 sPb-7850 98.9 sPb-1152
184.6 Xtxp248 99.2 sPb-1454
AGG+CAG5 AG+CAG3 105.4 AG+CTT5
185.1 AGG+CAC6 ACG+CAA5 108.8 SbKAFGK1_I
sPb-2729 113.2 SBKAFGK1
185.5 Xtxp316 119.4 sPb-2874
188.1 ACA+CAA2 130.4 AAC+CAT1

SBI-06 SBI-07 SBI-08 SBI-09 SBI-10

0.0 Xtxp273
1.1 sPb-6589
0.0 sPb-4183 2.2 AGC+CAA4 AG+CAA7
2.9 sPb-4937 6.3 AGG+CAG6
5.3 sPb-5182 15.1 sPb-2736 sPb-3464
7.6 AAG+CAT3 sPb-5390 sPb-7126
9.0 sPb-2551 23.9 0.0 sPb-3912
sPb-9818
11.8 sPb-3539 0.0 AG+CAA4 sPb-5825 6.8 sPb-8306 sPb-2041
26.2
14.8 AG+CTG9 sPb-9595 sPb-7064 sPb-3396 sPb-5079 RL
2.7 28.4 7.8
16.9 Xtxp6 sPb-4740 sPb-3995 AGG+CAT1 AAG+CAT6 AG+CTC8
52.2
20.6 AG+CAT2 3.1 sPb-2034 ACC+CAA4 0.0 sPb-0005 8.7 sPb-4129
55.9
23.5 sPb-8422 3.4 ACC+CTA4 sPb-9372 sPb-5005 4.9 Xtxp258 58.1 AG+CTA1
59.6
24.9 AG+CTC3 4.7 sPb-9223 sPb-2140 10.2 AGC+CAG2 60.8 sPb-8858
63.0
26.5 AAG+CAC5 AAG+CTC4 6.0 sPb-9931 sPb-4934 10.5 AG+CAG2 63.4 sPb-3432 sPb-0248
64.0
26.7 ACC+CAA6 11.3 AGG+CAA1 sPb-2641 15.1 ACC+CTA2 sPb-3287 sPb-8150
65.0 69.9
26.9 AAG+CTT2 AAG+CAA6 12.9 AG+CAA1 68.0 ACA+CAT6 20.6 ACA+CAT1 sPb-0817
27.2 AG+CAT1 ACA+CAT7 18.6 sPb-9513 71.7 ACC+CAA1 36.5 AAG+CAA5 sPb-2149 sPb-5391
19.6 sPb-3361 70.5 sPb-8497 sPb-0562
sPb-5564 sPb-1395 72.5 AAG+CAG6 50.8 ACC+CTA3
27.5 20.6 sPb-6518 53.7 sPb-9989 71.1 sPb-2836
sPb-7169 AGG+CAA6 sPb-1058
sPb-8198 24.7 AGC+CTG1 73.2 56.6 sPb-9688 73.7 sPb-3549 sPb-9999
27.8 sPb-2474 sPb-3195
28.1 sPb-8362 28.0 Xtxp159 73.6 AG+CAA6 59.3 sPb-8873 sPb-4786 76.2 AAG+CTG5
29.5 AGC+CAA1 31.3 sPb-8216 74.0 AG+CAT14 59.6 AG+CTA11 79.9 AAC+CAG3
ACC+CAA5 AAG+CTC3 42.8 AG+CTC5 74.4 sPb-2568 Xtxp210 59.9 sPb-1324 sPb-6852 80.7 AAG+CAA3
30.3 43.2 ACC+CTC3 AG+CTC2 sPb-5312 sPb-7643 sPb-3958
AAG+CAT7 76.1 ACA+CAG5
43.5 sPb-2566 61.3 sPb-9215 sPb-5512
30.5 ACC+CTA1 77.8 AAG+CTT3 sPb-9272 sPb-3158 81.1
30.6 AG+CTC7 47.0 sPb-6942 79.1 AAG+CAT1 AAG+CTC5 ACT+CAA1 sPb-6875
48.3 sPb-2774 62.4 81.4 ACT+CAA3
38.5 sPb-9146 79.8 sPb-0599 sPb-9841 AG+CTG3
57.7 sPb-1543 49.5 Xtxp227 sPb-9762 64.8 AG+CAC3 82.0 AAG+CTT4 sPb-4944
82.5
60.2 Xtxp145 ACA+CAG3 ACA+CAT4 sPb-9700 65.2 ACT+CAA2 82.6 AAG+CTG7
51.5 86.4
AG+CAT12 sPb-2757 65.9 AG+CAA2 83.4 sPb-1244
64.4 87.0 sPb-9468
Xtxp265-1 ACC+CAG1 sPb-1414 sPb-8019 66.7 AGG+CAA2 84.2 ACC+CAG2
65.9 53.4 88.9 sPb-7889
67.4 Xtxp274 sPb-8258 67.1 ACC+CAA2 85.8 ACA+CAA1
99.2 sPb-1881
70.2 AG+CTG11 AG+CTG5 68.1 AG+CTC6 67.5 AGG+CAC5 87.0 SvPEPCAA
sPb-1595 Xtxp18
73.0 AGC+CTG5 73.9 sPb-5594 100.3 AAG+CAC3 sPb-1850 75.6 AGG+CAA8 88.7 sPb-6889
78.0 AAG+CTT5 91.9 sPb-7086 sPb-7375 76.3 sPb-5142 90.3 sPb-5841
101.3
86.4 AAG+CTT6 95.3 sPb-3691 sPb-0031 sPb-2228 77.0 AAG+CTT1 93.7 AAG+CTC2
91.5 CC 97.5 Xtxp295 AGC+CTA2 111.3 93.5 sPb-3971 sPb-6288 sPb-4480
sPb-9743 sPb-3014 94.2
99.4 AAG+CAT5 99.7 AG+CTA5 sPb-3954 sPb-0833 sPb-9107 sPb-4853 sPb-2318 AG+CTA3
113.2 94.5
115.3 sPb-5374 107.8 sPb-4306 Xtxp105 sPb-9227 94.6 AGC+CTA1
115.1
125.6 sPb-3962 118.4 sPb-1014 AAC+CAA2 95.4 Xgap32 97.5 AG+CAT11
122.5
sPb-0017 sPb-5603 119.1 Xtxp168 AAG+CAG3 sPb-7367 sPb-8368 100.3 AAG+CAG2
126.2 148.7
131.9 120.5 sPb-7549 sPb-9091 102.3 ACA+CAG2
sPb-1486 sPb-4732 126.4 sPb-6918
134.2 Xtxp17 Xtxp95 126.6 AGG+CAA4 149.3 sPb-7955 108.8 ACA+CAT3
136.4 Xtxp57 129.0 sPb-5401 126.5 sPb-3003 sPb-1701
138.9 sPb-5212 sPb-6076 133.1 sPb-1272 133.6 AGG+CAC1 sPb-1660
141.4 AG+CTT2 133.5 sPb-8993 sPb-9584 140.7 Xtxp141 Xgap325
142.9 sPb-9667 133.9 sPb-6960 149.2 sPb-5678
144.3 sPb-7428 134.2 sPb-5250 sPb-2846
146.6 ACA+CAG4 134.5 sPb-7648 sPb-4385
148.8 ACA+CAT5 135.2 AG+CTG10
150.7 AG+CTG1 135.9 sPb-3434
157.1 ACG+CAA3 140.2 sPb-0087
178.2 sPb-0144
180.3 sPb-5473
181.7 sPb-3274
182.4 sPb-1997
183.8 sPb-9449
184.5 sPb-2770

Figure 2linkage map for a cross between R31945-2-2 and IS 8525

Genetic
Genetic linkage map for a cross between R31945-2-2 and IS 8525. Genetic distances are expressed as cumulative map
distances from position 0.0 (first locus of LG) in cM (Kosambi estimates). Locus names in bold indicate framework markers;
locus names in italics indicate attached and delegate markers.

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BMC Genomics 2008, 9:26
Additionally, this study meets all the criteria proposed for
evaluating the use of molecular markers for large scale
germplasm diversity analyses [35]; a large number of
markers, thorough representation of these markers on a
genetic linkage map of the species and the selection of
monolocus probes. The 508 DArT markers permitted the
unique identification of all 90 genotypes including closely
related backcross derived lines. The cluster analysis
revealed that the 90 genotypes examined showed a clear
demarcation of the germplasm according to their racial
classification, consistent with previous studies using
RFLPs [9] and SSRs and AFLPs [17]. The bicolor race was
found to be highly variable in this study, shown by its
presence in multiple clusters, as also noted by previous
studies [8,9,14]. In fact, it has been noted previously that
race bicolor resembles spontaneous weedy sorghums and
is thought to be the race most closely related to wild sor-
ghums [18] and also the most primitive grain sorghum
[10]. Indeed, the bicolor race accession IS 12179C
included in this study, groups together with the weedy
sorghum, S. bicolor subsp. drummondii, in cluster 13 (Fig-
ure 1).
The race caudatum is one of the most important agro-
nomically, providing genes for high yield and excellent
seed quality. It has become one of the most important
sources of germplasm in modern breeding programs
throughout the world and in this study, the caudatum race
genotypes were clearly demarcated in clusters 9, 10 and
12. In contrast, a previous study [18] found no significant
differences in diversity between the races, except the kafir
race which was less diverse. Low genetic diversity was also
observed in the kafir race in this study, with all the kafir
race accessions constituting a specific cluster (cluster 1).
These results are in agreement with the recent origin and
restricted geographic distribution of this race, together
with recent studies using SSRs [14] and RFLPs [9].
In addition to the observed clustering on racial groups,
differences between the genotypes based on their status as
B (maintainer female) and R (male parental restorer) lines
was also noted. In this study, there was less variation
among the B-lines than the R-lines, with the B-lines clus-
tering tightly together in only 2 of the 13 clusters, whereas
the R-lines grouped more loosely in 5 clusters. It has also
been noted [17] that the lower levels of diversity among
elite B-lines vs. R-lines is expected since B-lines are
required to produce high quality male-sterile A-lines. The
development of new A/B-lines is more difficult compared
to R-line development and hence B-line development is
more restrictive and slower to incorporate new germ-
plasm. Where pedigree data was available, groupings asso-
ciated with these pedigree relationships were observed.
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Comparisons of the discrimination ability for DArT mark-
ers and their ability to reflect pedigree backgrounds in
contrast to other marker types is made much simpler
when over-lapping sets of germplasm are used in a
number of studies. Seventeen genotypes included in this
study were examined by Menz et al. [17], 54 genotypes
were in common with Ritter et al. [20] and 11 with Tao et
al. [7]. Very similar groupings of genotypes were observed,
e.g. the over-lapping genotypes in cluster 1 also group
together in previous studies [17,20], e.g. BTx3197 and
BOK11 grouping tightly together in Menz et al. [17], in
addition to RTx7000 and Btx3042. Similarly, ICSV400,
MP531 and Macia group together based on AFLPs [17,20]
and DArTs (cluster 10), and QL41 and R9188 group
together based on RFLPs [7] and DArTs (cluster 2).
The separation of "wild" sorghums from cultivated germ-
plasm seems to be less pronounced compared to the
results based on SSR data [18]. This is not surprising, as
the array used for genotyping all materials in this study
was developed using mostly DNA from cultivated materi-
als. Even though three accessions from wild relatives were
included in library construction, their "private" alleles
were highly "diluted" by common alleles and those which
were "private" to the cultivated material. The design of the
array did not substantially change the topology of trees
generated from DArT data, but would reduce the distance
between the wild and cultivated samples. We expect that
the distance reduction would be up to two-fold, as we
were effectively finding unique "0" scores for the wild
materials, but less effectively unique "1" scores. Such
ascertainment bias is not limited to DArT, but is a feature
of many marker systems. In fact the ascertainment bias
will be significantly larger for SNP technologies, as most
SNP markers are developed from a small number of sam-
ples/chromosomes in a limited number of populations,
even in well resourced human studies [36,37]. The effect
of such "marker source sampling bias" was recognised in
many human studies, e.g. on population migration rate
[38], population mutation and recombination rate esti-
mates [39] and on Linkage Disequilibrium (LD) estimates
[40]. Marker panels in DArT are developed using large and
representative sampling from target populations [32] and
can be easily expanded by cloning libraries from the germ-
plasm pools for which precise relatedness estimates
would be required.
Distribution of DArT loci in the sorghum genome
The integrated genetic linkage map comprising DArTs,
AFLPs and SSRs clearly demonstrates that the new sor-
ghum DArT markers behave in a Mendelian manner. In
total, 358 DArTs were mapped to 257 unique loci. The
higher level of redundancy of the DArT markers is
reflected by the higher number of AFLP and SSR markers
having unique segregation patterns. However, the markers
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BMC Genomics 2008, 9:26
on the DArT array used in this study were not filtered for
redundancy, whereas the SSR/AFLP data set had previ-
ously undergone curation [34], to remove markers with
redundant segregation patterns. Also, the total number of
DArT markers was higher than in other marker classes,
therefore the apparent redundancy would need to be also
corrected for sample size. After applying such sample-size
correction to STMP markers in the linkage map of a cross
between two wheat cultivars Cranbrook and Halberd, a
lower level of redundancy was found for the DArT mark-
ers [30].
The total length of the integrated genetic linkage map was
1431.6 cM, with an average DArT marker density of 1 per
every 3.9 cM. The total map length is comparable to other
recently reported sorghum genetic maps, being slightly
shorter than the 1713 cM high-density genetic linkage
map based on 2926 AFLP, RFLP and SSR markers [41],
and slightly longer than the 1059.2 cM genetic linkage
map based on 2050 RFLP probes [42]. Although the DArT
markers are distributed across the genome in a similar way
to the non-DArT markers, there are genomic regions con-
taining significant excesses or paucity of markers, e.g. the
centromeric region of SBI-05 has a higher than average
marker density of 1/0.64 cM and the distal ends of SBI-01
and SBI-10 have marker-poor regions with gaps spanning
over 30 cM. These areas of low marker density may corre-
spond to regions of similar ancestry or identity by descent
(IBD) in the germplasm included in the initial diversity
representation. In addition, lines with photoperiod sensi-
tivity and tall stature were under represented in the diver-
sity set used to develop the DArT markers. These regions
of low marker density may be therefore associated with
genomic regions that were identical by descent or that had
very limited genetic variability in the initial diversity rep-
resentation. The marker-dense regions appear to corre-
spond to the centromeric regions, a feature that has been
observed previously [42]. This is also supported by the
recent observation that the pericentromeric heterochro-
matic regions of sorghum chromosomes show much
lower rates of recombination (~8.7 Mbp/cM) compared
to euchromatic regions (~0.25 Mbp/cM), with the average
rate of recombination across the heterochromatic portion
of the sorghum genome being ~34-fold lower than recom-
bination in the euchromatic region [43]. It should how-
ever be noted that clustering around the centromeres is
observed for both DArT and non-DArT markers, due to
the centromeric suppression of recombination. Interest-
ingly, DArT markers were significantly less clustered at
most centromeric regions of barley chromosomes com-
pared to non-DArT markers on the integrated map con-
taining approximately 3,000 markers [26]. We will be in
position to rigorously test if this difference in marker posi-
tion holds true in sorghum only after completion of
building the consensus map integrating approximately
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1,000 DArT markers with similar number of other types of
markers [Mace et al in preparation].
Conclusion
We have successfully developed DArT markers for Sor-
ghum bicolor and have demonstrated that DArT provides
high quality markers that can be used for diversity analy-
ses and to construct medium-density genetic linkage
maps. The high number of DArT markers generated in a
single assay not only provides a precise estimate of genetic
relationships among genotypes, but also their even distri-
bution over the genome offers real advantages for a range
of molecular breeding and genomics applications. Addi-
tionally, the availability of the sorghum whole genome
sequence by the end of 2007 offers very exciting opportu-
nities for assessing the colinearity of the DArT markers on
the genetic linkage maps with the markers on the
sequence map. As DArT assays are performed on highly
parallel and automated platforms the cost of datapoint (a
few cents per marker assay) is reduced by at least an order
of magnitude compared to current, gel-based technolo-
gies.
Methods
Source of DNA
The sorghum accessions used to prepare DArT libraries
represent the genetic diversity present in the cultivated
species (S. bicolor subsp. bicolor) with all 5 races and 5
intermediate races represented, two weedy subspecies (S.
bicolor subsp. drummondii and subsp. verticilliflorum) and a
wild species, S. propinqum (Additional File 1). In addition,
the germplasm set included elite lines from breeding pro-
grams some of which had high levels of co-ancestry. DNA
was extracted using a modified CTAB-based extraction
protocol [44,45].
Development of DArT for sorghum
Several DArT arrays were built in the course of this study.
For each of these arrays, a genomic representation was
generated from a mixture of sorghum lines using the PstI-
based complexity reduction method previously described
[26]. Libraries were prepared as described previously [25].
Two extended PstI+BanII sub-libraries were subsequently
generated using DNA from 31 and 94 genotypes, respec-
tively (Additional File 2). In addition, six libraries were
generated by applying the SSH method to genomic repre-
sentations [33]. Drivers and testers used in the subtraction
libraries construction were as shown in Table 4. DNA of
drivers and testers was digested by PstI/BstNI and ligated
to the PstI adaptor. The digestion/ligation products were
amplified using the PstI-0 primer. The resulting PCR prod-
ucts were digested with a RE mixture containing DpnII,
HpyCH4IV, MseI and NlaIII. Subtraction was done in a
30:1 ratio of driver to tester and carried out in one and two
rounds of subtractive hybridization. After final amplifica-
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BMC Genomics 2008, 9:26
tion, the subtraction products were cloned as detailed pre-
viously [27]. The sorghum libraries utilized for the initial
marker discovery are summarized in the supplementary
material (Additional File 3). Clones from all libraries with
the exception of PstI+BanII Library C were used to create
arrays in order to genotype several hundred sorghum
accessions (data not presented). The best markers from
the initial experiments were "cherry picked" to assemble
the "re-array library" with 768 polymorphic clones.
Clones from this library together with 5367 clones from
PstI+BanII Library C were used for all genotyping work
reported here. The re-array library was created using the
PstI+BanII and the subtraction (SSH) libraries. Details of
the re-array library and other libraries used for sorghum
genotyping are included in the supplementary material
(Additional Files 2 &3).
DArT genotyping
Genotyping was performed essentially as described in ref-
erences 26 and 30. Briefly, each genomic DNA sample is
subjected to the PstI+BanII complexity reduction method.
The resulting genomic representation is labelled with flu-
orescent nucleotides and hybridised on a microarray
printed with the DArT clones. A typical experiment is per-
formed on about 94 samples of genomic DNA. Following
hybridisation and washing, the microarrays for an experi-
ment are scanned, the images are analysed and the score
of each marker is calculated for each sample by dedicated
software DArTsoft: markers are scored 1 for presence, 0 for
absence and X for inability to score. The quality parameter
Q for each marker is calculated by dividing the variance of
the hybridisation level for the marker between the 2 clus-
ters (present and absent) by the total variance of hybridi-
sation level of the marker, in the experiment.
Diversity analysis
A group of 90 sorghum lines (subset detailed in Addi-
tional File 1) were genotyped on the re-array library as
described previously [30]. The sorghum lines were chosen
to provide a reasonable representation of sorghum genetic
diversity as well as including elite inbred lines from the
DPI&F and other sorghum breeding programs. Some of
the elite lines were quite closely related and were included
to demonstrate the discrimination possible with DArT.
Table 4: Drivers and testers used in the subtraction libraries
Library 1 Library 2
Subtraction method One round One round
Library Subtraction-1 Subtraction-2
Driver ISCV745 ISCV745
tester Tester mixture* 90562
* The tester mixture was comprised of 90 sorghum genotypes without the four parental genotypes (ICSV745, B90562, R31945-2-2 and IS 8525).
http://www.biomedcentral.com/1471-2164/9/26
The marker scores were subjected to cluster and principal
coordinate analysis using the DARwin [46] to visualize
the genetic relationships among the lines. Additionally,
the polymorphism information content (PIC) of each
DArT marker was determined as follows; PIC = 1-ΣPi2,
where Pi is the frequency of the ith allele in the examined
genotypes [47].
Genetic mapping
The lines R931945-2-2, a commercially accepted restorer
line in Australia and IS 8525, an Ethiopian line (kafir
race), in addition to 92 lines of a F5 recombinant inbred
line (RIL) population derived from a cross between the
two lines were typed using the genotyping array. Clones
with Q > 80% and a call rate of at least 80% were selected
for mapping. DArTs markers were merged with an existing
mapping data set consisting of 286 markers including 55
SSRs and 229 AFLPs [34]. A genetic linkage map was con-
structed using MultiPoint software [48]. The RIL_Selfing
population setting was selected and a maximum thresh-
old rfs value of 0.40 was used to initially group the mark-
ers into ten linkage groups. Multipoint linkage analysis of
loci within each LG was then performed and marker order
was further verified through re-sampling for quality con-
trol via jack-knifing [49]. Markers that could be ordered
with a jack-knife value of 90% or greater were included as
'framework' markers, with any remaining markers causing
unstable neighborhoods being initially excluded from the
map, including redundant markers mapping to the same
location. Following a repeated multipoint linkage analy-
sis with the reduced set of markers for each LG to achieve
a stabilised neighbourhood, the previously excluded
markers were attached by assigning them to the best inter-
vals on the framework map and labelled as attached mark-
ers. The redundant markers were also included on the
final, complete map but labelled as delegates. Finally, the
linkage groups were assigned to sorghum chromosomes,
SBI-01 to SBI-10 according to recent nomenclature [50].
The Kosambi [51] mapping function was used to calculate
the centimorgan (cM) values. The graphical representa-
tion of the map was drawn using MapChart software [52].
Authors' contributions
ESM carried out the diversity and mapping analyses and
drafted the manuscript. LX co-developed the DArT tech-
Library 3 Library 4 Library 5 Library 6
One round One round One round Two rounds
Subtraction-3 Subtraction-4 Subtraction-5 Subtraction-6
90562 IS8525 31945-2-2 Mixture of drivers
ISCV745 31945-2-2 IS8525 Tester mixture*
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BMC Genomics 2008, 9:26
nology for sorghum. DRJ conceived of the study, and par-
ticipated in its design and coordination, germplasm
development and selection and helped to draft the manu-
script. KH was involved in the optimisation of the meth-
odology and participated in the mapping analysis. DKP
participated in the mapping analysis. EH participated in
the development of the DArT technology and editing of
the manuscript. PW contributed to ongoing improve-
ments of the DArT technology and the quality assessment
of sorghum clones. AK supervised development of the
DArT technology, participated in the study's design and
coordination and helped to draft the manuscript.
Additional material
Additional File 1
List of sorghum genotypes used for the development of the sorghum DArT
array and the diversity analyses. The table includes details of genotype IDs
and aliases, race and origin and inclusion status in both the methodology
developmental stages and diversity analyses.
Click here for file
[http://www.biomedcentral.com/content/supplementary/1471-
2164-9-26-S1.doc]
Additional File 2
Summary of sorghum libraries. The table includes details of the number
of genotypes used in the development of each sorghum library and the
number of clones identified.
Click here for file
[http://www.biomedcentral.com/content/supplementary/1471-
2164-9-26-S2.doc]
Additional File 3
Libraries used for generation of the genotyping array. The table details the
barcode for each sorghum library.
Click here for file
[http://www.biomedcentral.com/content/supplementary/1471-
2164-9-26-S3.doc]
Acknowledgements
We thank the Australian Grains Research and Development Cooperation
[53] for financial support. We thank Robert Henzell, Bert Collard, Mandy
Christopher, Sally Dillon and colleagues at Diversity Arrays Technology P/
L for helpful discussions and comments on the manuscript.
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Wang Y, Kresovich S, Schertz KF, Paterson AH: A high-density scientist can read your work free of charge
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