Nagy et al. BMC Genomics 2012, 13:469
http://www.biomedcentral.com/1471-2164/13/469
RESEARCH ARTICLE
Open Access
A high-density genetic map of Arachis duranensis,
a diploid ancestor of cultivated peanut
Ervin D Nagy1, Yufang Guo1, Shunxue Tang1, John E Bowers1, Rebecca A Okashah1, Christopher A Taylor1,
Dong Zhang1, Sameer Khanal1, Adam F Heesacker1, Nelly Khalilian1, Andrew D Farmer2, Noelia Carrasquilla-Garcia3,
R Varma Penmetsa3, Douglas Cook3, H Thomas Stalker4, Niels Nielsen4, Peggy Ozias-Akins5* and Steven J Knapp1
Abstract
Background: Cultivated peanut (Arachis hypogaea) is an allotetraploid species whose ancestral genomes are most
likely derived from the A-genome species, A. duranensis, and the B-genome species, A. ipaensis. The very recent
(several millennia) evolutionary origin of A. hypogaea has imposed a bottleneck for allelic and phenotypic diversity
within the cultigen. However, wild diploid relatives are a rich source of alleles that could be used for crop
improvement and their simpler genomes can be more easily analyzed while providing insight into the structure of
the allotetraploid peanut genome. The objective of this research was to establish a high-density genetic map of
the diploid species A. duranensis based on de novo generated EST databases. Arachis duranensis was chosen for
mapping because it is the A-genome progenitor of cultivated peanut and also in order to circumvent the
confounding effects of gene duplication associated with allopolyploidy in A. hypogaea.
Results: More than one million expressed sequence tag (EST) sequences generated from normalized cDNA libraries
of A. duranensis were assembled into 81,116 unique transcripts. Mining this dataset, 1236 EST-SNP markers were
developed between two A. duranensis accessions, PI 475887 and Grif 15036. An additional 300 SNP markers also
were developed from genomic sequences representing conserved legume orthologs. Of the 1536 SNP markers,
1054 were placed on a genetic map. In addition, 598 EST-SSR markers identified in A. hypogaea assemblies were
included in the map along with 37 disease resistance gene candidate (RGC) and 35 other previously published
markers. In total, 1724 markers spanning 1081.3 cM over 10 linkage groups were mapped. Gene sequences that
provided mapped markers were annotated using similarity searches in three different databases, and gene ontology
descriptions were determined using the Medicago Gene Atlas and TAIR databases. Synteny analysis between
A. duranensis, Medicago and Glycine revealed significant stretches of conserved gene clusters spread across the
peanut genome. A higher level of colinearity was detected between A. duranensis and Glycine than with Medicago.
Conclusions: The first high-density, gene-based linkage map for A. duranensis was generated that can serve as a
reference map for both wild and cultivated Arachis species. The markers developed here are valuable resources for
the peanut, and more broadly, to the legume research community. The A-genome map will have utility for fine
mapping in other peanut species and has already had application for mapping a nematode resistance gene that
was introgressed into A. hypogaea from A. cardenasii.
* Correspondence: pozias@uga.edu
5
Department of Horticulture, University of Georgia, Tifton, GA 31793, USA
Full list of author information is available at the end of the article
© 2012 Nagy 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.
Nagy et al. BMC Genomics 2012, 13:469
http://www.biomedcentral.com/1471-2164/13/469
Background
Cultivated peanut (Arachis hypogaea L.) is a major crop
in most tropical and subtropical areas of the world and
provides a significant source of oil and protein to large
segments of the population in Asia, Africa and the
Americas. In the U. S., peanut is a high-value cash crop
of regional importance, with major production areas
concentrated in the Southeast. Plant breeding efforts to
pyramid genes for disease and insect resistances, quality,
and yield is hampered by the polyploid genetics of the
crop species, the multigenic nature of many traits (e.g.,
yield), and the difficulty of selecting for many traits
in the field (e.g., soil borne diseases). Thus, secondary
selection methods that are environmentally neutral
would greatly facilitate crop improvement efforts. Molecular markers fit this criterion, but only recently have
markers been developed that reveal sufficient polymorphisms in A. hypogaea and related species to have widespread application in peanut breeding. Preliminary steps
for utilizing molecular markers for crop improvement
are developing collections of polymorphic markers and
utilizing them to construct dense and high-resolution
genetic maps.
Constructing a high-quality genetic map depends
largely upon finding one or more marker systems that
can detect high levels of polymorphism between two
individual parents. Unfortunately, low levels of molecular polymorphism were observed within tetraploid
(2n = 4x = 40) A. hypogaea throughout the 1990s and
early 2000s with the marker systems available at that
time [1,2]. However, compared with the limited numbers
of polymorphic markers detected for the tetraploid, the
same marker systems can uncover high levels of molecular polymorphism within and between the diploid
(2n = 2x = 20) peanut species. This polymorphism led
researchers to create molecular maps for Arachis. The
first molecular map in peanut was constructed between
the diploids A. stenosperma Krapov. and W.C. Gregoryx
and A. cardenasii Krapov. and W.C. Gregory by Halward
et al. [3] who used Restriction Fragment Length Polymorphisms (RFLPs) to associate 117 markers into 11
linkage groups. Additional maps were subsequently published using Randomly Amplified Polymorphic DNA
(RAPD) [4] and Simple Sequence Repeats (SSRs) [5,6].
Burow et al. [7] published the first tetraploid map
in peanut based on 370 RFLP loci across 23 linkage groups by utilizing the complex interspecific cross,
Florunner × 4x [A. batizocoi Krapov. and W.C. Gregory
(A. cardenasii × A. diogoi Hoehne)]. Another interspecific tetraploid linkage map of 298 loci and 21 linkage
groups was derived from a backcross population between A. hypogaea and a synthetic amphidiploid [8].
Only recently have linkage maps been developed from
crosses between A. hypogaea genotypes, most with less
Page 2 of 11
than 200 loci and with more than the expected 20 linkage groups [9-13]. An exception is the recently published
map containing 1114 loci across 21 linkage groups
that was constructed in part with highly polymorphic
markers derived from sequences harboring miniature
inverted repeat transposable elements [14]. Therefore,
there is a continuing need to generate dense linkage
maps for the cultivated tetraploid peanut that will not
only cluster the markers into the expected 20 linkage
groups to cover the haplotype chromosomes, but also
to facilitate marker-trait association and eventually assist
in its genetic improvement.
The domesticated peanut is thought to have arisen
from a single hybridization event between two diploid
wild species followed by whole genome duplication
approximately 3,500 years ago [15]. This short evolutionary history, along with hybridization barriers between diploids and the tetraploid have resulted in a
narrow genetic base for the cultivated tetraploid peanut.
On the contrary, diploid Arachis species are genetically
diverse, have simpler inheritance patterns, and most importantly, contain a rich source of agronomically important traits for peanut improvement. Due to these
attributes, diploid Arachis species have been proposed as
model systems to map the peanut genome. Because the
genomes of progenitor diploid species [i.e., A. duranensis
(A-genome donor) and A. ipaensis (B-genome donor)]
are closely allied to the cultivated peanut [16], mapping
the genome of one or both of these species should be
useful for predicting the positions of loci in the cultivated
peanut. This approach has been employed in wheat
[17,18], alfalfa [19,20], oat [21], and other crop species.
One accession of A. ipaensis and 67 accessions of
A. duranensis have been collected in South America.
The largest concentration of A. duranensis is in southern
Bolivia and northern Argentina, with a few populations
being reported in Paraguay and one in central Brazil
[22,23]. The species is morphologically diverse and the
Bolivia and Argentina types can be separated cytogenetically and morphologically [24]. Due to the availability of
diverse accessions to produce intraspecific crosses in the
greenhouse, a dense linkage map in the diploid species
A. duranensis was produced using large numbers of
molecular markers derived from transcribed sequences.
Results and discussion
Species relationships
A preliminary study of SSR marker variation among 37
A. duranensis accessions using 556 markers indicated
that the species is highly polymorphic at the molecular
level and individual accessions could be separated based
on a cluster analysis (Figure 1). Interestingly, we found
that A. ipaensis, the proposed B-genome (BB) progenitor
species, clustered with the A-genome (AA) species
Nagy et al. BMC Genomics 2012, 13:469
http://www.biomedcentral.com/1471-2164/13/469
Page 3 of 11
A. duranensis
(AA)
PI468201
PI468202
PI468203
PI468319
Grif 14248
PI468325
PI468328
PI468322
Grif 7731
A. stenosperma
PI591351
PI468329
PI468326
PI468327
A. batizocoi
(BB)
Grif15033
Grif15030
A. ipaensis
Figure 1 Genetic relationships among A- and B-genome Arachis species. Clustering of A- (A. duranensis and A. stenosperma) and
B- (A. ipaensis and A. batizocoi) genome species according to analysis of data from SSR markers. The two parents used for mapping are
indicated by arrows.
A. stenosperma and not with the B-genome species A.
batizocoi. Recent molecular cytogenetic analysis of Aand non-A- (i.e., B-) genome species suggests that karyotype diversity among non-A-genome species is extensive
enough to support separation into additional genome
classes where A. ipaensis remains in B sensu stricto while
A. batizocoi is placed into a separate group [25]. Therefore, A. batizocoi is less typical of B-genome species.
The number of polymorphic SSR markers between
paired A. duranensis accessions ranged from 160 to 375
out of 556, which is 29 to 67% of the total number of
SSR markers screened. This is a significant amount of
variation, which indicates the high genetic diversity
within the species. Based on cluster analysis, success of
crosses, and fertility of F1s, accessions PI 475887 and
Grif 15036 were selected for subsequent mapping studies using 94 F2 progenies. Screening of the parental
accessions with 2,138 SSR markers derived from A.
hypogaea EST sequences resulted in 1,768 (82.7%) that
were scorable (detected by ABI3730XL genotyping systems) and 896 (41.9%) that were polymorphic (Guo Y et al:
Comparative mapping in intraspecific populations
uncovers a high degree of macrosynteny between A- and
B-genome diploid species of peanut, Submitted). The
same markers were used to create a map between
two A. batizocoi accessions and to determine syntenic
relationships between the A and B genome species
(Guo Y et al: Comparative mapping in intraspecific populations uncovers a high degree of macrosynteny between
A-and B-genome diploid species of peanut, submitted).
Arachis duranensis genetic map
The total number of published SSR markers has now
risen beyond the 2,847 cataloged in a related paper by
Guo et al. (Guo Y et al: Comparative mapping in intraspecific populations uncovers a high degree of macrosynteny between A-and B-genome diploid species of
peanut, submitted) to around 6,000 [26]. Those most
recently reported include: 14 by Gimenes et al. [27]; 51
by Mace et al. [28]; 188 by Proite et al. [29]; 104 by Cuc
et al. [30]; 138 by Yuan et al. [31]; 33 by Song et al. [32];
123 by Wang et al. [33]; 290 by Liang et al. [34]; and
1,571 by Koilkonda et al. [35]. Five hundred and ninetyeight of these markers are included in the A. duranensis
map (Figure 2). Of the 34 genomic SSR markers mapped
in the current study (Table 1), 24 were mapped previously in an interspecific population between A. duranensis and A. stenosperma [6,36]. These markers served to
anchor and align the current and previously published
peanut maps (Figure 2). Linkage group assignments of
all markers were consistent between the current map
and that of Bertioli et al. [36] except for the marker
GM117 (AC3C02 on map in reference 36 derived from
GenBank accession DQ099133) that was localized on
Nagy et al. BMC Genomics 2012, 13:469
http://www.biomedcentral.com/1471-2164/13/469
1A
2A
3A
Page 4 of 11
4A
5A
6A
7A
8A
0.0
0.6
2.3
0.0
0.3
95.9
97.0
98.7
99.1
101.1
101.9
102.4
102.7
103.0
103.3
103.8
104.0
104.2
105.1
108.4
109.3
110.0
110.4
111.4
112.0
114.5
116.3
116.9
118.7
119.3
119.8
120.5
121.7
122.3
122.9
124.6
124.7
124.8
125.2
126.5
127.1
127.6
127.7
128.8
129.4
129.8
130.5
131.2
132.1
132.6
133.2
136.0
137.4
138.4
138.6
138.9
139.9
140.8
141.1
141.8
143.5
145.6
69.4
70.2
70.8
71.2
71.7
72.0
72.4
73.8
74.9
75.9
77.0
79.0
80.2
81.0
81.5
82.6
83.2
85.8
87.5
90.2
91.1
94.5
95.3
96.8
97.8
98.9
99.9
100.4
101.0
101.3
102.0
102.9
106.3
109.1
112.0
112.9
115.8
59.4
59.9
60.1
60.8
61.4
61.9
62.3
62.5
63.2
63.6
64.3
65.4
66.4
66.5
67.0
67.8
68.4
68.9
69.6
69.7
70.0
70.1
70.3
71.1
71.7
72.3
72.8
73.5
74.3
75.2
75.8
77.0
77.2
77.7
80.1
80.7
83.2
83.5
85.0
86.8
89.6
92.6
93.1
93.6
94.2
95.9
98.0
107.8
108.3
SNP410
SNP294
111.7
112.0
SNP1066*
GM1702*
GM1162 SNP62*
GM1944*
112.1
116.4
GM1352*
131.7
SNP42*
99.1
99.3
100.4
100.9
101.5
104.8
105.3
109.8
32.1
32.2
32.7
33.0
34.6
36.9
37.3
38.4
39.6
40.7
41.0
43.2
44.3
46.5
48.9
51.2
52.5
55.2
55.6
58.6
59.2
62.3
64.3
67.3
70.2
71.3
74.9
75.5
77.2
78.0
78.4
78.5
79.0
79.6
80.6
82.3
57.3
57.6
57.7
57.8
57.9
58.1
58.4
59.6
60.0
60.4
61.3
61.6
62.1
62.5
62.6
62.9
63.3
63.7
63.9
64.4
64.6
64.9
65.1
65.5
66.0
66.3
66.9
67.3
67.4
67.6
67.9
68.2
68.7
68.9
69.0
69.2
69.6
69.9
70.2
70.7
71.5
71.7
71.8
72.6
73.8
74.4
74.8
75.2
75.5
75.7
77.3
GM950
GM1373
12.9
GM2266
SNP107 GM2082
GM951
GM1562
GM2577
SNP372*
SNP304* SNP530*
SNP614* SNP805*
GM810*
16.3
SNP321
18.0
SNP157 GM1107
SNP527 SNP828
SNP838
SNP994
RGC33
GM2316
SNP1004
SNP147
RGC1
GM1561
SNP254 SNP404
SNP510
11.3
SNP350
12.6
SNP640
12.7
14.6
SNP516*
21.7
GM2438
GM1600* GM324#9*
SNP441*
SNP599* SNP1017*
SNP887* GM2120*
GM1338*
GM1237*
SNP149* SNP774*
SNP948*
SNP577* GM1952*
GM1876*
SNP364* SNP437*
SNP521*
SNP385* SNP439*
GM1559* SNP65*
GM1650*
SNP975* GM1732*
SNP378*
GM1893*
SNP218* SNP226*
SNP919* SNP1009*
SNP346*
SNP679*
SNP356*
SNP525*
SNP630*
SNP47*
GM1796*
GM66*
SNP860*
SNP296* SNP413*
SNP482* SNP711*
RGC144b*
SNP84* SNP111*
SNP402* SNP465*
SNP1057*
SNP952*
SNP672* SNP762*
SNP1043*
SNP446*
SNP871*
GM2164*
GM1069*
SNP28*
SNP1052*
SNP38* SNP616*
SNP708* SNP787*
GM2172*
GM2839* SNP96*
SNP733* SNP827*
GM2359*
GM1911*
SNP338* SNP701*
SNP851* SNP934*
SNP1044* SNP1050*
SNP972* GM1979*
GM2792b* GM1834*
GM2778*
GM565*
SNP429* GM328#9*
GM2847*
SNP68* SNP85*
SNP92* GM2553*
GM2292*
SNP735*
GM1851*
GM1411* SNP5*
SNP567* SNP978*
GM2003* GM2389*
GM2843*
GM1291*
SNP36* SNP685*
SNP422* SNP739*
SNP154* SNP874*
SNP951*
SNP953*
GM1841*
SNP790* GM7#9*
GM1998* GM1999*
GM1047*
SNP100* SNP150*
SNP609*
GM2033*
S197*
SNP6*
SNP141*
SNP13* SNP550*
SNP875*
SNP124*
SNP977*
SNP49* SNP292*
SNP584* SNP674*
SNP621*
GS20a*
SNP794*
SNP21*
GM1803*
RGC63*
SNP722*
GM2529*
SNP533* GM1459
SNP884* SNP1055*
GM1117*
GM1329*
GM1312*
GM1607*
SNP1109*
SNP276* SNP748*
SNP457*
SNP738 GM1199*
GM2081 GM1389
SNP305*
SNP1110*
SNP537*
23.7
24.3
24.7
27.0
28.2
28.7
28.9
30.0
30.5
33.3
34.4
36.2
36.6
36.7
37.1
37.5
38.0
39.0
39.6
41.5
42.0
42.3
43.2
43.7
44.2
44.8
45.4
46.5
47.1
48.3
48.9
49.9
51.2
51.5
51.9
52.3
52.6
53.2
53.7
54.5
55.1
55.5
55.6
55.7
55.9
56.2
56.6
56.7
57.3
57.9
58.4
59.5
59.7
59.9
60.2
60.4
61.0
62.4
63.4
65.8
66.2
67.3
70.0
70.3
70.6
73.0
73.3
73.8
78.2
79.8
83.0
86.0
87.7
89.2
89.3
91.1
93.0
94.1
94.2
95.0
97.4
97.8
99.4
100.5
101.0
104.6
106.5
18.7
19.5
20.7
21.2
21.9
23.0
24.1
25.5
25.9
30.8
31.2
31.7
SNP746 GM1163
GM1624
GM2041
34.1
SNP158
37.0
37.2
39.6
40.7
41.3
43.2
44.2
45.3
47.2
47.3
48.1
49.0
50.3
50.4
51.3
51.8
52.6
83.8
91.6
96.7
97.3
99.0
100.2
103.8
SNP879
GM1764
GM2140
GM2029
SNP541
GM2058
GM926
GM927
SNP603
GM1195
GM2303
SNP230
GM2722
SNP121
GM1868
GM1481
SNP724
SNP57 SNP376
SNP715 SNP1006
SNP303
SNP781
SNP99 SNP426
SNP506 SNP731
SNP996
SNP788 GM1243
GM2320
SNP709
SNP590
GM2383
SNP829
SNP250
GM1341
GM2040
GM2084
GM2207
GM1563
GM784
GM2582 GM1322
SNP109 SNP119
SNP202 SNP380
SNP486 SNP651
SNP683 SNP695
SNP865 SNP922
SNP993 GM1303
GM117#2
GM1526 GM1933
SNP532
GM1875
SNP234 SNP257
SNP627 SNP749
GM2104 GM1460
GM1172
SNP409
GM1581
SNP243
SNP133
GM2348 GM1408
SNP677
SNP240
GM1453
GM2531
SNP606 SNP658
SNP802
SNP649
GM1527
GM2283
SNP618 SNP923
SNP488
GM2148
SNP105 SNP1029
SNP244 SNP933
SNP444
SNP528
GM2218
SNP703
SNP193
SNP669
GM1520c
SNP581 SNP878
GM1886 GM1520a
GM1520b
SNP297
SNP548 GM857
SNP671
SNP547
SNP173 SNP1021
SNP406
SNP98
108.4
GM2166
53.1
53.4
53.8
54.4
55.1
55.6
55.9
56.3
56.7
56.8
57.3
57.6
57.9
58.2
58.4
58.5
58.6
58.7
58.8
59.1
59.6
59.8
60.1
60.3
60.7
61.2
62.3
63.3
63.6
64.2
64.7
65.4
65.7
67.0
69.1
70.8
71.8
72.0
76.0
78.6
79.7
80.6
80.7
81.5
82.4
83.3
83.4
1.5
GM1865
SNP88
67.7
68.0
68.7
59.3
31.1
32.0
GM2001
5.5
6.3
8.4
8.9
9.0
9.2
10.1
12.2
1.0
SNP687
67.4
67.5
59.1
30.9
56.6
0.0
0.5
SNP897
103.7
103.9
66.0
66.3
66.7
67.0
64.1
64.7
64.8
65.9
66.2
69.1
69.3
69.4
69.5
69.7
70.5
70.6
70.8
71.1
71.2
71.3
71.5
73.2
74.5
76.7
76.8
77.4
77.9
79.8
80.5
81.1
82.7
83.3
83.9
84.4
84.8
88.9
91.0
91.1
91.7
94.2
95.9
96.5
97.1
58.4
53.0
53.1
54.1
54.2
55.0
55.6
56.1
56.3
56.4
0.0
93.8
96.3
100.7
65.6
57.1
58.1
58.2
58.3
52.8
SNP467
GM1174
SNP673
SNP477
GM1856 GM1835
SNP615
SNP256
1.5
85.7
GM1930
64.0
64.4
64.7
65.2
56.1
51.5
51.6
51.8
52.7
SNP358*
4.7
5.3
6.6
7.0
7.4
7.9
8.5
1.0
84.6
63.3
54.9
55.0
56.1
57.1
57.6
58.2
58.7
58.8
59.0
60.0
60.4
61.3
63.0
63.5
63.8
50.7
51.3
51.7
52.6
52.8
53.3
54.5
54.9
55.2
55.9
50.1
50.6
50.8
50.9
51.2
0.0
0.5
78.6
78.8
79.1
79.4
79.6
80.1
80.6
81.6
83.1
83.9
58.6
58.9
59.3
59.8
60.8
62.1
62.9
53.8
54.4
54.8
49.1
50.3
48.7
49.0
49.3
49.4
49.6
49.7
0.0
78.5
58.4
53.1
53.2
53.4
53.5
53.6
53.7
37.1
38.9
41.1
42.4
44.8
45.7
46.2
47.3
48.4
10A
2.0
78.3
57.2
57.5
57.8
57.9
58.0
58.2
51.5
52.9
36.1
12.3
12.6
13.6
14.4
15.1
15.4
15.5
15.6
16.5
16.7
16.8
16.9
17.3
17.8
17.9
18.6
19.2
20.7
20.9
21.2
21.7
21.9
22.0
22.6
22.9
23.1
23.5
23.9
24.4
24.7
25.4
25.8
26.7
27.1
27.3
28.1
28.7
29.0
29.3
29.4
30.4
1.5
64.1
64.4
65.0
65.6
66.6
67.6
69.4
71.7
71.8
75.5
76.4
77.4
77.9
SNP438
SNP277
SNP986
SNP270
SNP269 SNP831
GM1565 SNP814*
SNP1015* GM2246*
GM1021*
GM1720*
GM1755
GM1296*
GM2589*
SNP268*
SNP329* SNP468*
SNP909* GM218*
SNP1040* SNP1062*
SNP1073*
SNP470* GM2813*
GM1887*
GM1959*
GM2311*
SNP983*
GM2572*
GM2006*
SNP689* SNP693*
SNP889*
SNP1002*
GM1465*
GM1995* GM1070*
SNP921*
SNP387* SNP582*
GM1805*
SNP430*
SNP453* SNP836*
GM916*
SNP172*
SNP638* SNP1060*
GM1186* GM1643*
RGC89
SNP593* GM1167*
GM96*
GM2142*
GM1013*
SNP935* SNP960*
SNP1024* GM2482*
GM2480*
SNP336*
RGC205* GM1554*
GM162* SNP45*
SNP93* SNP183*
SNP495* SNP982*
GM1190*
GM1143
GM1907*
SNP451* SNP938*
GM1919*
GM1679*
GM1594*
SNP279*
SNP323*
GM1298*
GM1729*
GM1223*
GM2236*
SNP704*
SNP284
SNP714* GM13#4*
GM2583*
GM942*
GM955
GM1062
SNP944*
SNP824*
SNP4
SNP442*
SNP954*
SNP825
GM1817
GM735*
GM1963*
SNP1005
GM1045
GM2755*
SNP290 GM2021
GM2020
55.4
47.6
48.7
49.0
49.8
50.2
50.7
51.0
GM226
GM1567
SNP39 SNP286
SNP920
GM1548
SNP76
GM1547
SNP898 SNP450
SNP950
GM2170 SNP497
SNP517 SNP929
GM1286
SNP726
SNP1033
GM1385
GM1968 SNP394
SNP1007
GM2844
SNP128
SNP408* SNP123*
SNP331*
SNP911 SNP9
SNP718
SNP752 SNP311
SNP864
GM2063
SNP956 SNP267
SNP607
SNP417
GM664 GM959
GM1505 SNP694
SNP971
GM2701
GM1185 SNP418
GM92#6
GM2702
SNP449
GM988
SNP1023
GM1461 SNP155
SNP352 SNP459
SNP480 SNP539
SNP575 SNP1032
GM1035 SNP1
SNP44 SNP136
SNP179 SNP224
SNP310 SNP557
SNP578 SNP642
SNP705 SNP793
SNP821 SNP876
SNP1049 GM1096
GM1962 GM1896
GM83
SNP625 SNP238
SNP624
SNP1027
GM2366* SNP41*
SNP67
SNP958 GM2306
SNP511
GM1909
SNP464 RGC53a
RGC53b
SNP728
GM1617
SNP681
RGC93b
GM1748
SNP789
GM900
SNP400
SNP969 GM2425
SNP322
SNP569 SNP461
SNP573
SNP187
SNP215
GM1572
SNP195 SNP151
SNP730 SNP51
GM2817
GM255#6
SNP1058
SNP239
GM847
GM1498
SNP312
SNP509
SNP721
GM2446
SNP991 SNP273
SNP299 SNP490
SNP553 SNP834
SNP858 SNP995
SNP1041 SNP203*
SNP907 GM2109*
GM2110
SNP75
SNP456
SNP281 SNP635
GM2414*
SNP246* SNP796*
SNP1053 SNP727*
GM2337
GM2767
SNP143 SNP1047
SNP152
SNP676 SNP900
SNP942
GM2066
GM1490
GM1497
SNP700 SNP916
SNP1039
SNP118
GM1506
SNP915*
42.7
43.1
43.2
43.6
44.7
45.3
45.8
46.1
46.6
47.2
48.1
1.0
86.2
88.4
89.3
91.7
93.4
95.0
83.1
63.4
GM1867*
50.8
51.7
51.9
52.8
53.1
47.0
SNP131
24.5
25.4
26.9
31.2
32.8
33.8
34.1
34.5
35.1
41.3
0.5
85.0
66.5
67.7
72.8
72.9
75.3
77.9
78.6
79.6
79.7
81.3
81.8
63.0
48.3
42.6
44.2
45.0
46.6
SNP518
SNP544
GM1940
19.4
11.5
8.2
8.7
9.8
9.9
16.8
17.1
17.8
24.1
25.1
25.6
26.6
27.6
28.4
29.3
30.0
30.4
32.1
35.0
35.8
36.0
36.2
36.9
38.5
39.4
39.8
0.0
SNP806
53.9
54.2
54.5
54.9
55.3
55.5
56.2
56.7
57.2
59.6
62.6
63.7
64.2
64.7
66.1
59.1
59.4
59.7
SNP91 SNP620
SNP778 GM1445
SNP337
SNP503
GM2448
GM1738
SNP1014
SNP472
GM839
SNP302
GM1173
40.8
16.7
17.3
18.0
13.8
2.0
90.9
53.7
54.9
56.5
57.1
57.2
57.7
58.6
60.5
60.7
61.0
61.2
61.8
62.1
62.6
37.9
39.7
40.3
41.7
42.9
43.3
43.4
43.8
44.3
45.1
45.9
40.1
14.2
SNP817
SNP1037
SNP596
GM1521 SNP112
GM1495 SNP27
SNP102 SNP344
SNP888
GM1682*
1.5
84.5
86.4
86.9
88.1
88.9
SNP320*
SNP962*
RGC236
SNP228* SNP968*
SNP872
GM126#2
SNP407 SNP670
SNP729
SNP12 SNP499
GM2474
SNP420
SNP180 SNP883
SNP643
RGC141
SNP90
SNP10 SNP168
SNP185 SNP272
SNP494 SNP505
SNP570 SNP734
SNP756 SNP1003
GM2455 GM1516
GM1503
GM1075
SNP433 SNP880
SNP587
SNP190
SNP863
GM2491
GM1598
GM1386
SNP348
SNP602*
SNP1010
SNP177
SNP101
SNP293
GM2496
SNP867 GM1714
GM2797
SNP391
GM1946
SNP424
GM2047
GM1852
SNP574
GM1724
GM1160
RGC5b
GM2500
GM733 SNP799
GM2016
SNP807
SNP594
SNP646
SNP288
GM1819
51.4
51.9
52.5
53.0
53.2
53.5
GM1647
GM1466
GM2108
10.1
10.6
11.2
13.3
1.0
46.8
48.1
49.0
49.1
49.6
49.8
50.7
51.3
54.6
39.5
SNP313 SNP563
SNP976
GM76#4
SNP571
GM2057 SNP452
SNP210
SNP23 SNP930
GM2473
SNP261
SNP508
SNP713
GM852
GM1633
GM2073
GM963
GM2146
GM2147
SNP434
SNP295 SNP556
SNP198 SNP403
SNP77
GM1621
SNP585
SNP955
SNP1059
SNP2
GM2177
SNP20 GM1929
SNP24 SNP1020
GM1250
GM1175
SNP357 SNP588
SNP782 GM1140
GM2454
GM1785
SNP589
SNP519 GM2050
GM2156
GM2199
SNP564
GM1942
GM1494
SNP381
SNP634
GM1632
SNP469
GM1111
SNP33
GM1114
GM846
GM1688
SNP481
GM2588
GM1023
GM1623
SNP608
SNP595 SNP837
SNP397
SNP217 SNP455
SNP882
GM2222
SNP339 SNP1026
SNP707
GM1858
SNP485
SNP946 SNP959
GM1076
GM1451
SNP592
SNP479
GM2034
SNP611
GM2423 SNP114
SNP138 SNP196
SNP301 GM1651
GM2326
GM1922
GM1937 SNP200
SNP688 SNP985
GM1315
GM1829 SNP318
SNP520 SNP791
SNP260 SNP655
SNP710
SNP375 SNP568
SNP870
GM1477
GM59#7*
SNP766*
GM1219* GM2747*
SNP192*
SNP369*
SNP586* SNP1036*
SNP366* SNP771*
SNP932*
GM2494
GM1701
SNP841
SNP549
SNP795
GM802
SNP940*
SNP343*
SNP784* GM1853*
GM69#7*
SNP275 SNP650
GS3b
GM1214
SNP641
SNP389
GM980
SNP835
GM1347
SNP803 GM1348
GM2526
SNP1016
GM840
GM953
SNP274
0.5
SNP178
SNP106 SNP918
SNP662*
GM2111
GM1668
SNP421*
GS110* SNP384*
RGC233*
SNP97 SNP235
GM1528
SNP225
34.7
35.5
35.6
31.5
7.2
1.1
1.4
2.2
3.1
3.4
4.0
4.6
4.9
5.0
5.3
6.5
6.8
7.0
7.3
7.4
7.8
8.1
8.6
8.8
9.5
0.0
38.0
39.2
40.8
41.0
41.1
42.9
43.5
44.6
28.0
29.1
29.7
32.6
33.1
SNP999
1.5
42.0
42.3
43.5
44.9
45.8
46.3
46.7
47.0
47.7
49.5
49.6
50.0
51.2
51.7
52.1
52.6
53.0
54.0
GM2010
SNP58
2.8
1.0
SNP745*
SNP656*
26.0
0.0
0.5
GM2257*
35.5
36.1
GM1120
SNP822
SNP458
SNP881
0.0
41.4
GM2814*
GM2121*
33.6
14.8
15.9
16.5
16.7
3
SNP427* GM1910*
29.4
29.6
40.2
SNP227
0.9
2
27.2
38.3
39.1
8.1
9.2
9.5
12.4
SNP626
SNP26 SNP431
SNP732
SNP60
SNP768
GM2770
SNP333
GM2703
SNP493 SNP753
RGC44
SNP974
SNP11 GM1898
SNP989
SNP265 SNP936
GM1728
GM1253
GM1519
SNP842
GM948
SNP291
GM1261
SNP869
SNP664
GM2793*
GM1119
SNP252 GM1118a
SNP622 SNP759
SNP804
SNP699
SNP816
SNP325
SNP379
SNP617
GM1317*
SNP559 SNP767
SNP750
GM1154
SNP191 SNP691
GM1313
SNP54
SNP161 SNP162
SNP182 SNP504
GM2771
SNP170 SNP702
SNP964
GM2663
SNP757 SNP1019
GM2662
SNP737 SNP844
GM1987
GM1332
GM2092
SNP134
GM2031
SNP201
GM2043
GM874
GM996 SNP1025
SNP1030
GM1049
SNP144 SNP654
SNP742 GM2820
GM1892
GM2078
GM1257
SNP135
GM68#5 SNP72
SNP86 SNP207
SNP554 SNP810
SNP818 SNP859
SNP862 SNP1000
GM1577
GM2160
GM1482
GM1955 SNP562
SNP847
GM1967
SNP108 SNP538
GM16#5
SNP761
SNP211
GM28#5
SNP139 SNP212
SNP80
SNP411
GM1018
GM2347
SNP156 SNP680
SNP812
GM1890 GM2429
SNP66 SNP205
SNP720 SNP941
GM1218 GM2487
SNP82
SNP237 GM2672
GM2673 GM1782
GM2137
SNP392
SNP129 GM2231
GM2769
GM1788
GM1227
SNP89
SNP171
GM1228
SNP579
GM1878
GM2017
GM1843
SNP580
GM1778
SNP253* SNP902*
SNP282* GM1293*
SNP220*
SNP130*
SNP723*
SNP957*
SNP63* SNP903*
SNP29* SNP40*
SNP199* SNP289*
GM944*
GM1902*
SNP117
SNP71 GM1912
GM1816
GM1904
SNP83 SNP652
SNP64
SNP35
RGC279
1
40.8
38.1
GM2014
SNP306
SNP59
GM2091
0.6
1.6
2.9
4.0
5.6
6.0
6.2
6.7
7.3
9.5
10.1
11.8
12.4
12.7
13.7
15.6
16.8
17.8
18.0
18.5
19.6
20.6
23.8
24.2
25.4
26.0
26.4
27.7
28.3
31.7
32.4
36.3
37.0
38.2
38.3
0
39.9
GM2808*
GM1836*
GM2098*
6.6
2.0
SNP405*
24.4
25.5
25.9
SNP857
GM1840
SNP597
1.5
SNP53*
22.9
SNP142 SNP845
3.5
3.9
4.6
1.0
SNP852*
RGC193*
20.7
0.0
0.5
17.6
18.1
0.0
GM817*
GM2838*
2.0
81.8
83.8
87.1
88.5
89.5
91.4
96.8
15.4
15.8
1.5
61.1
61.4
61.5
61.9
62.2
62.3
62.7
63.1
63.7
64.5
67.3
71.0
73.2
73.8
74.0
74.4
75.9
76.3
76.7
77.1
79.7
80.3
SNP551* SNP772*
GM1655*
SNP3*
11.4
0.0
0.0
1.0
60.7
60.8
60.9
61.0
31.4
31.5
32.7
33.9
35.4
36.6
37.5
37.9
38.0
12.6
GM32#3
SNP712
GM2599
SNP927 GM2598
SNP125
GM2818
SNP931
GM2261
GM2372
GM1760
GM1149
SNP682* GM1609*
GM1191
GM1425*
GM2764* SNP478*
SNP164*
SNP832
GM2087
SNP113* SNP335*
SNP342*
SNP780*
SNP1045*
SNP890*
SNP231*
SNP491*
SNP632*
GM1266*
GM2407*
SNP34* SNP813*
SNP1008*
GM1360* RGC146b
SNP186* SNP314*
SNP904* RGC72a*
GM2693* GM2694*
SNP31* SNP137*
SNP534* SNP552*
GM1733* RGC17b*
SNP194*
SNP566* GM2230*
GM2524*
SNP222* SNP792*
RGC177* RGC170*
RGC70b* RGC182*
RGC249 RGC220
GM2467*
SNP328*
SNP601*
RGC140b
GM1656*
GM1487* SNP165*
SNP447* SNP856*
GM828
GM2206*
GM2364*
SNP660*
GM2352*
GM2184
GM2274
SNP917
SNP653
GM774
SNP330 SNP894
GM1566
GM1034
SNP377*
SNP939
GM898
SNP540
GM1618*
SNP167 SNP823
GM1473
SNP69
SNP30* SNP285*
SNP809* GM1763*
GM19#3*
SNP241* SNP500*
GM2008
SNP926
GM1583
RGC18b*
GM1500
GM1053* SNP572*
GM2449
SNP583 SNP605
SNP777 SNP783
SNP943
SNP1018
GM783
GM2586 SNP811
SNP661
SNP896*
SNP283 GM2402
GM1645
SNP686*
SNP905
GM1659a
SNP524*
GM1681*
GM1443*
GM1369* SNP263*
SNP435*
SNP46* SNP223*
SNP317* SNP354*
SNP383* SNP425*
SNP436* SNP445*
SNP513* SNP561*
SNP666* SNP885*
SNP949* SNP980*
GM1502* GM1717*
GM2215*
SNP56*
GM2556* SNP698*
SNP327* SNP349*
SNP690
SNP262* SNP895*
SNP631
GM1249*
SNP990
SNP233
SNP300 SNP423
SNP542
GM2772 GM1726
SNP163 SNP440
SNP833 GM2051
SNP1111
GM1883
SNP531
SNP1001*
SNP32* SNP623*
SNP645
GM2416
SNP906 SNP908
GM2027
GM743
SNP801
SNP17 SNP600
GM2419
SNP204 SNP543
GM2528
SNP390 SNP463
GM1108 GM1885
GM2069*
SNP371 SNP576
GM866
SNP103 SNP644
SNP1031*
SNP945*
GM2765
GM1084*
SNP176
SNP7* SNP886*
GM2792a*
GM714
SNP264
SNP496
GM1326*
SNP855* SNP970*
SNP776*
SNP819*
SNP153*
GM1954*
SNP219* SNP353*
SNP675* SNP773*
GM1321*
GM1652*
GM2228* SNP255*
SNP326* SNP483*
SNP839*
GM2103*
GM2805*
SNP507* SNP529*
SNP639* SNP966*
GM1213*
GM1591*
SNP912* SNP961*
GM1536*
SNP800*
GM74#3*
SNP741*
SNP725*
SNP668* SNP743*
GM2302*
GM2301*
SNP87*
SNP120* SNP1051*
SNP763*
SNP591*
SNP341* SNP347*
SNP15* SNP612*
SNP914* GM1263*
SNP1038*
SNP8* SNP719*
GM1854
SNP492*
SNP786
SNP175
SNP526*
GM2757
0.5
60.6
0.0
8.1
8.9
9.1
9.3
9.4
9.7
12.1
13.4
14.0
15.7
17.3
20.2
20.4
21.2
25.3
28.3
30.1
0.0
59.1
59.5
59.7
60.2
60.4
2.0
56.9
57.3
57.8
58.0
58.8
GM2845
SNP628
SNP398*
SNP395*
GM2186*
GM2232
SNP853*
SNP522*
GM922*
GM1355* SNP416*
SNP820*
SNP1068
SNP249*
1.5
53.6
53.9
54.3
54.5
55.1
56.0
2.1
3.2
4.0
4.7
6.1
6.2
6.4
6.8
7.0
7.6
8.4
8.7
1.0
53.2
0.0
0.5
50.2
50.3
50.5
50.8
51.0
51.7
52.8
0.0
50.1
1.5
46.7
47.1
48.1
48.7
49.0
49.3
49.5
49.6
49.7
49.9
50.0
1.0
25.4
26.5
28.3
30.9
34.1
34.6
36.3
37.1
37.3
38.7
39.7
40.8
41.0
41.4
42.2
44.0
44.5
45.0
45.6
46.0
46.4
46.6
0.5
24.1
0.0
5.2
5.6
8.2
9.3
12.8
15.1
15.5
16.0
18.4
19.1
20.8
22.9
23.5
SNP770
RGC108b SNP1012
SNP1063
SNP716
SNP370 SNP984
SNP1035
GM24#1
GM1030 GM825
SNP25
SNP850
SNP308
SNP415
SNP717*
SNP498 SNP979
GM1331
SNP454
GM1118b GM1761
GM2022
SNP14 SNP146
SNP169
SNP754
SNP78 SNP363
SNP873
GM1504
SNP797
GM2568*
SNP514 GM1132
SNP1067
SNP73*
GM1208*
SNP555
SNP565
GM2603
SNP546
SNP629 SNP785
SNP636
GM2024
SNP367
GM1423
SNP484
GM1483
GM2173 SNP278
SNP678 SNP988
SNP271 SNP471
GM2153
GM2233
GM2607
GM170#1
SNP1013
GM917
GM2015
GM1171
GM890 GM2597
SNP19 SNP70
SNP79 SNP104
SNP110 SNP122
SNP127 SNP242
SNP258 SNP351
SNP388 SNP460
SNP466 SNP523
SNP536 SNP665
SNP846 SNP848
SNP861 SNP893
SNP901 SNP947
GM22#1 GM1649
GM1992 GM2527
GM1233 GM1972
GM2287
GM1048
GM672
GM824
GM286#1
GM1777
SNP396 SNP899
GM2840
SNP16 SNP216
SNP316 SNP332
SNP647 GM1065
SNP1046 GM1353
SNP604
SNP55
SNP232
SNP779 GM1330
GS22b
SNP747 GM1339
GM933
SNP95
SNP412 SNP692
GM1444
GM2724
SNP808 SNP981
GM1950
SNP515
GM2390
GM1879
SNP213 SNP280
SNP148 GM2783
GM2349 GM1913
SNP866 SNP487
SNP399 SNP373
SNP355 SNP236
SNP50
GM1771
GM2350
SNP558
GM1501
GM1542 SNP462
GM1599
GM2418
GM2132
GM1871
GM2004
SNP189
GM1684
GM2061
SNP1056
SNP307
SNP736
SNP386 GM2653
SNP229
SNP744
GM2053
GM1573
SNP52
GM38#1
SNP706
SNP345
GM1275 GM1620
SNP610 GM1277
SNP160 SNP868
SNP1042 GM1085
GM2344*
GM1750
SNP188 SNP489
SNP1011
GM1612
SNP928*
9A
SNP545
SNP74 SNP197
GM2421
SNP37 SNP126
SNP362
SNP145
SNP997
GM1239 GM1073
GM2269
SNP967
SNP61
SNP184
GM1134
SNP998
RGC206
RGC268
GM1628
GM1320
GM1575
SNP43 SNP340
GM1872
SNP840
GM2055*
GM1932
GM10#8
SNP740
GM2758
SNP48 SNP287
SNP298
GM1961
SNP775 SNP1034
SNP1048
SNP251 SNP324
SNP94 GM883
GM1844
SNP159
SNP1065
GM2400
SNP22
GM1744
SNP393
SNP209 GM2101
SNP475
SNP684 SNP925
SNP1028
GM1136
SNP360 GM58
GM1965
GM1165
SNP208
SNP443
SNP248 SNP501
SNP830
SNP368
GM741
SNP659
SNP598 SNP755
SNP361 SNP613
SNP1085
GM2182
SNP937
SNP758
SNP1022 SNP1061
GM1092
GM2125
SNP319 SNP637
SNP166 SNP259
GM2787
GM2089
SNP132*
GM1281
GM2144
GM2551
SNP18 SNP359
GM2842 GM1613
SNP535 SNP910
GM2788
GM1709
SNP401
GM1201
SNP315 SNP657
SNP798
GM2832
GM2643 GM1001
SNP206
GM1798
SNP115 SNP245
SNP826
GM2644
GM2640
SNP965
GM1903a
SNP619
SNP633
SNP1100 SNP214
GM1240
RGC244 RGC219
GM1629
GM1901
SNP663 SNP1064
GM1905 GM1934
GM2339
GM2312
SNP892
SNP116 SNP266
SNP334 SNP365
SNP877 GM2600
SNP473 SNP849
GM1839
GM2601
GM1936
GM1831
SNP81 SNP221
SNP476
GM2176
SNP924 GM2665
GM2664
GM1917
GM1189
GM1509
GM2827
GM2564 GM1797
SNP419 SNP769
GM2712 GM2713
SNP913
GM71#8
GM1659b
GM1713
GM1238
SNP502 SNP560
SNP140
SNP374
GM2417
SNP432 SNP474
SNP891 GS43b
SNP174 SNP1054
SNP963
GM1957
GM1587
GM2099
110.2
SNP815 GM2032
111.4
SNP696
Figure 2 High-density linkage map of Arachis duranensis including 1,724 markers. SNP and SSR markers are prefixed by ‘SNP’ and ‘GM’,
respectively, resistance gene candidate markers are prefixed by ‘RGC’ and ‘GS’. Twenty-four previously published markers (underlined) were
selected from an interspecific map between A. duranensis and A. stenosperma [36] to establish synteny between the current and former linkage
groups. The original linkage group assignments are given in the marker names separated by the pound (#) sign. Loci with significant segregation
distortion (p = 0.05) are labeled with an asterisk. Graphs to the right of the linkage groups represent recombination frequencies. Each data point
represents genetic distances between adjacent markers averaged for a window of 20 markers.
chromosome 2A (the ‘A’ following a chromosome number is presented in this study to represent chromosomes
in the A genome of peanut) in their interspecific map,
while mapping to chromosome 10A in the A. duranensis
intraspecific map. Although detailed information for parental alleles in the study by Bertioli et al. [36] was not
presented, GM117 amplified only one locus from each
parent in both their population and ours. It is, therefore,
unlikely that the marker location discrepancy was due to
mapping of multiple loci and perhaps could reflect a
small chromosomal rearrangement. Chromosomal rearrangements are not unexpected based on previous cytological observations in the genus [24,37].
EST libraries of A. duranensis were developed to produce Single Nucleotide Polymorphism (SNP) markers
for mapping (Table 2). Of the 1,536 SNP markers developed (Additional file 1), 1,054 were included in the
A. duranensis map (Figure 2). The remaining 482 SNP
markers were either of low quality (GC quality score
<0.25) or they showed segregation patterns (extremely
distorted) that could not be mapped. Of the 1,054
mapped SNP markers, 815 were derived from the cDNA
sequencing project while the other 239 were genomic
legume orthologs.
The A. duranensis map produced in this study contained 1,724 markers combined into 10 linkage groups
with a total genetic distance of 1081.3 cM. MSTMap, a
software program that accommodates large numbers of
markers and utilizes a “minimum spanning tree” algorithm, was used to construct an initial genetic map using
only the codominant markers. The 1,673 codominant
markers were distributed into 810 co-segregating groups
(bins). Although this program has been reported to
be accurate for large-scale mapping projects [38], few
independent studies are available establishing consistency
between MSTMap and other commonly used mapping
software [39]. To confirm the linkage group assignments,
marker orders, and genetic distances determined by alternative software, both codominant and dominant markers
were mapped with Joinmap 3.0. Marker orders and genetic distances were highly consistent between MSTMap
and Joinmap 3.0 (Additional file 2).
Significant segregation distortion (p = 0.05) was
observed for 513 (29.8%) markers (Figure 2, Additional
file 3). Chromosomes 4A and 9A carried particularly
long segments of distorted segregation suggesting largescale chromosomal selection in these regions. Guo et al.
(Guo Y et al: Comparative mapping in intraspecific
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Table 1 Previously published genomic SSR markers mapped in Arachis duranensis
Universal Name
Original Name
Forward (50-30)
Reverse (50-30)
Reference
GM7
Ah1TC1D02
GATCCAAAATCTCGCCTTGA
GCTGCTCTGCACAACAAGAA
Moretzsohn et al. 2005
GM10
Ah1TC1E05
GAAGGATAAGCAATCGTCCA
GGATGGGATTGAACATTTGG
Moretzsohn et al. 2005
GM13
Ah1TC1H04
CATTACTTCCTAGGTTTGTTTTCCA
ATGGCGTGACAACGGAAC
Moretzsohn et al. 2005
GM16
Ah1TC2B01
TTGCAGAAAAGGCAGAGACA
GAAAGAAGCTAAGAAGGACCCATA
Moretzsohn et al. 2005
GM19
Ah1TC2C07
CACCACACTCCCAAGGTTTT
TCAAGAACGGCTCCAGAGTT
Moretzsohn et al. 2005
GM22
Ah1TC2D06
AGGGGGAGTCAAAGGAAAGA
TCACGATCCCTTCTCCTTCA
Moretzsohn et al. 2005
GM24
Ah1TC2E05
GAATTTATAAGGCGTGGCGA
CCATCCCTTCTTCCTTCACA
Moretzsohn et al. 2005
GM28
Ah1TC3A12
GCCCATATCAAGCTCCAAAA
TAGCCAGCGAAGGACTCAAT
Moretzsohn et al. 2005
GM32
Ah1TC3E02
TGAAAGATAGGTTTCGGTGGA
CAAACCGAAGGAGGAACTTG
Moretzsohn et al. 2005
GM38
Ah1TC3H02
CTCTCCGCCATCCATGTAAT
ATGGTGAGCTCGACGCTAGT
Moretzsohn et al. 2005
GM58
Ah1TC4G06
ATTTCACATTCCCTAGCCCC
CATCGACTGACTTGAAAAATGG
Moretzsohn et al. 2005
GM59
Ah1TC4G10
TTCGGTCATGTTTGTCCAGA
CTCGAGTGCTCACCCTTCAT
Moretzsohn et al. 2005
GM66
Ah1TC5D06
GAAATTTTAGTTTTCAGCACAGCA
TTTTCCCCTCTTAAATTTTCTCG
Moretzsohn et al. 2005
GM68
Ah1TC6E01
CTCCCTCGCTTCCTCTTTCT
ACGCATTAACCACACACCAA
Moretzsohn et al. 2005
GM69
Ah1TC6G09
GGAGGTTGCATGCATCATAGT
TCATTGAACGTATTTGAAAGCTC
Moretzsohn et al. 2005
GM71
Ah2TC7A02
CGAAAACGACACTATGAAACTGC
CCTTGGCTTACACGACTTCCT
Moretzsohn et al. 2005
GM74
Ah2TC7E04
GAAGGACCCCATCTATTCAAA
TCCGATTTCTCTCTCTCTCTCTC
Moretzsohn et al. 2005
GM76
Ah2TC7G10
AATGGGGTTCACAAGAGAGAGA
CCAGCCATGCACTCATAGAATA
Moretzsohn et al. 2005
GM83
Ah2TC9C06
CAAATGGCAGAGTGCGTCTA
CCCTCCTGACTGGGTCCT
Moretzsohn et al. 2005
GM92
Ah2TC11A04
ACTCTGCATGGATGGCTACAG
CATGTTCGGTTTCAAGTCTCAA
Moretzsohn et al. 2005
GM96
Ah2TC11C06
TCCAACAAACCCTCTCTCTCT
GAACAAGGAAGCGAAAAGAA
Moretzsohn et al. 2005
GM117
Ah2AC3C02
TCTAACGCACACAAATCGAA
CTTGTACCTGCGCCATTCT
Moretzsohn et al. 2005
GM126
AS1RI1F06
TGTCTCTCTTCCTTTCCTTGCT
CCTTTTGCTTCTTTGCTTCC
Moretzsohn et al. 2005
GM162
AS1RN9C02
CGTTACACTGAGCCAGCAACTC
ACGGCGGCGATAGTTTCA
Moretzsohn et al. 2005
GM170
AS1RN11E05
CTCGGTCCAGAAAACACAGG
GTAGAGGCGAAGAAGGCAGAG
Moretzsohn et al. 2005
GM218
gi-30419832
GCCACTTTATTCTAAGCACTCC
AAGAGACCACACGCTCACA
Moretzsohn et al. 2005
GM226
gi-30419936
TCACAGATCCATAGACTTTAACATAGC
CCGGTGTGGATTCATAGTAGAG
Moretzsohn et al. 2005
GM255
pPGPSeq4H6
CCAACATTGCAGAAGCAAGA
CAAAGAGAGGCACACCACAA
Moretzsohn et al. 2005
GM286
Ah-193
CTTGCTGAAGGCAACTCCTACG
TCGGTTTGTCTCTTTGGTCACTC
Moretzsohn et al. 2004
GM324
Ah-649
GGAAATGCCAAATCCATCCTTC
GTTGTTCGGTGTGAAAACGGTC
Moretzsohn et al. 2004
GM328
Ah-671
AGAAAGAGCACGGGACATTACC
ATGAATGAGTGGTCATACGCGA
Moretzsohn et al. 2004
GM565
pPGSseq17E3
TTTCCTTTCAACCCTTCGTG
AATGAGACCAGCCAAAATGC
Ferguson et al. 2004
GM664
GM664
CTTCACCTCCAAAATCAACCA
ACCGCTGACATTTGATTGTTC
Guo et al. 2011
GM671
GM671
TGGATGCTGTAAGGAATGGAC
TTATCGAGCTTGCCTCAGAAA
Guo et al. 2011
Markers were renamed in order to follow a unified marker nomenclature. The complete list of renamed markers can be found in Guo et al. (Guo Y et al:
Comparative mapping in intraspecific populations uncovers a high degree of macrosynteny between A-and B-genome diploid species of peanut, Submitted).
populations uncovers a high degree of macrosynteny between A-and B-genome diploid species of peanut, Submitted) found that a single linkage group (4/9B) in A.
batizocoi was syntenic with chromosomes 4A and 9A of
A. duranensis implicating inversion and reciprocal
translocation events as the underlying chromosomal
rearrangements in this B-genome species. Recombination
frequencies were generally low in the central, presumably
centromeric chromosomal regions of A. duranensis and
increased toward the telomeres, a pattern typical of many
plant species [40,41]. More even distribution was
observed along chromosome 3A and only slightly suppressed recombination was observed around the presumable location of the centromere (Figure 2).
Across the A. duranensis linkage map, each linkage
group spanned on average 108.1 cM (77.3-145.6 cM)
and included 172.4 markers (119–266) (Table 3). This is
considerably denser than the previously published AA,
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derived from a cross between cultivated peanut and a
synthetic A. ipaensis x A. duranensis tetraploid (OziasAkins, unpublished).
Table 2 cDNA sequence reads generated for single
nucleotide polymorphism (SNP) discovery in Arachis
duranensis*
Accession
Sequencing
Platform
PI 475887
Sanger
22,356
21,487
43,843
PI 475887
454
212,938
266,575
479,513
Grif 15036
454
Total
Tissue type
Total
Developing seed
Gene annotation and comparative mapping
Root
296,242
235,245
531,487
531,536
523,307
1,054,843
* Assembly is deposited at NCBI as Accession: PRJNA50587.
BB, and AABB maps consisting of only a few hundred
markers. For example, the A. ipaensis × A. magna Bgenome map published by Moretzsohn et al. [5] had 149
SSR markers grouped into 10 linkages, whereas the
B-genome SSR-based map in our related paper consists
of 449 loci in 16 linkage groups (Guo Y et al: Comparative mapping in intraspecific populations uncovers a
high degree of macrosynteny between A-and B-genome
diploid species of peanut, Submitted). The A-genome
map produced using the interspecific hybrid A. duranensis × A. stenosperma had 339 SSRs that were mapped
into 11 linkage groups [6,42]. For A. hypogaea, there are
now several maps with the most dense consensus map
containing 324 loci on 21 linkage groups [11].
The map produced in the current study is the first
high-density map available in peanut, and because it was
generated from a progenitor species of A. hypogaea, we
anticipate that it will have significant applications for
analyzing the cultivated genome. For example, the data
generated in this map was used by Nagy et al. [43]
to more precisely map the Rma gene for nematode
resistance that originated from an introgression line
between A. hypogaea and A. cardenasii. The A-genome
SNP array also has been useful at the tetraploid level
for genotyping a recombinant inbred line population
Table 3 Genetic distances and distribution of markers on
the ten linkage groups of A. duranensis
Linkage group
Genetic distance (cM)
Markers
1A
96.8
186
2A
103.9
119
3A
145.6
266
4A
115.8
149
5A
131.7
178
6A
109.8
181
7A
82.3
141
8A
77.3
180
9A
106.5
171
10A
111.4
153
Total
1081.3
1724
Homology search of the 1,724 mapped loci resulted in
significant hits for 1,463 loci in at least one of the three
databases: Medicago, Uniprot and GenBank NR database, and 580 of the mapped loci gave significant similarities in either of the two gene ontology databases:
Medicago Gene Atlas and TAIR (Additional file 4).
Altogether 1,366 gene ontology terms were assigned to
the 580 genes. These were distributed among the three
major gene ontology categories as follows: 521 molecular
functions, 534 biological processes, and 311 cellular
components (Additional file 4).
The sequences used to create the A. duranensis map
also were compared to the genomes of two legumes
where 995 loci on the A. duranensis map could be
mapped to M. truncatula, and 2,711 matches could be
found in G. max (with potentially two hits per mapped
locus). While a majority of the dots in the synteny plots
appear to be random (Figure 3), there are definite clusters of markers, and striking examples of colinearity (red
arrows), especially for the comparisons to Glycine. Presumably there has been extensive single gene movement
since the last common ancestors in one or both species,
but many genes remain in the ancestral locations and
can be detected. Overall, the synteny patterns for G.
max showed the recent whole genome duplication
within Glycine, with each location in peanut showing
corresponding synteny at two locations in Glycine. Colinearity between Medicago and Arachis is much less
conserved than between Arachis and Glycine. This could
be due to extensive inversions in either genome, or more
likely, due to preliminary ordering of sequences within
the Medicago unfinished genome assembly. In general,
the patterns showed strong synteny on the chromosomal
ends in both genetic and physical distance, while the
central regions of chromosomes tended to show less
synteny. Presumably this could be attributed to pericentromeric heterochromatin which is known to define less
recombinogenic regions where genomic rearrangements
are more likely to persist [44]. Chromosome arms tend
to be maintained as syntenic between Glycine and Arachis, but there is evidence that chromosome arms have
been translocated in some cases so that synteny exists at
the chromosome arm level, but less so at the whole
chromosome level.
Conclusions
This investigation provided a large number of de novo
EST sequences that were deposited into GenBank. The
markers developed here are valuable resources for peanut
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Figure 3 Synteny between diploid A-genome peanut (A. duranensis, 2n = 20) and Glycine max (2n = 40). Arrows indicate clusters of genes
in common between the two genomes. For plotting the data on the Y axis, the peanut genome for each chromosome is proportional in size to
the total map size in centimorgans. For the X axis, the unit of measure is scaled to bp within the chromosomal assemblies of the respective
genomes. The plot was obtained with a visual basic program that plotted the x-y coordinates of each hit. The total number of matches for each
pair wise comparison is listed at the upper left corner.
and, more broadly to the legume research community.
This research presents the first high-density molecular
map in peanut with 1,724 markers grouped into the 10
expected linkage groups for an A-genome species.
Because the map was produced with the progenitor
species A. duranensis which contributed the A genome
of A. hypogaea, it will serve as the reference map for
both wild and cultivated species. Lastly, synteny was
found between Arachis and the Glycine and Medicago
genomes, which indicates that markers developed for
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other legume species may be of value for crop improvement in peanut. The A-genome map will have utility for
fine mapping in other peanut species and has already
had application to mapping a nematode resistance gene
that was introgressed to A. hypogaea from A. cardenasii.
polymorphic fragments. The matrix was imported into
Phylip v3.67 [55] for the construction of the neighborjoining tree.
Methods
A total of 101,132 unigenes (37,916 contigs (GenBank Acc.
No. EZ720985-EZ758900) and 63,216 singletons) from
tetraploid peanut ESTs (GenBank Acc. No. CD037499CD038843, ES702769-ES768453, GO256999-GO269325,
GO322902- GO343529 and short-read Sequence Read
Archive accessions SRX020012, SRX019979, SRX019972,
SRX019971) representing ca. 37 Mb of the A. hypogaea
genome were mined for 2,138 EST-SSR markers (GM710GM2847) (Guo Y et al: Comparative mapping in intraspecific populations uncovers a high degree of macrosynteny
between A-and B-genome diploid species of peanut,
Submitted). Unigenes in the transcript assembly were
screened for perfect repeat motifs using SSR-IT http://
www.gramene.org/db/markers/ssrtool) and for imperfect
motifs using FastPCR (http://primerdigital.com/fastpcr.
html). The repeat count (n) threshold for each motif type
was set for n ≥ 5. SSR markers were genotyped on an
ABI3730XL Capillary DNA Sequencer (Applied Biosystems, Foster City, CA) using forward primers labelled
with FAM, HEX, or TAMRA fluorophores. PCR was performed in a 12 μL reaction mixture containing 1.0 × PCR
buffer, 2.5 mM Mg++, 0.2 mM each of dNTPs, 5.0 pmol
of each primer, 0.5 unit of Taq polymerase, and 10 ng of
genomic DNA. Touchdown PCR was used to reduce
spurious amplification. The SSR markers were screened
for length polymorphisms using GeneMapper 3.0 software
(Applied Biosystems, Foster City, CA). Of the 2,138 ESTSSR primer pairs tested, markers derived from 598 could
be mapped. A set of 34 SSR markers from genomic
sequences of Arachis previously screened for polymorphism between parents of the A. duranensis mapping
population (Guo Y et al: Comparative mapping in intraspecific populations uncovers a high degree of macrosynteny between A-and B-genome diploid species of peanut,
Submitted) were also mapped (Table 1).
Plant materials
Thirty-seven accessions of A. duranensis, 14 accessions
of A. stenosperma (A genome), one accession of A.
ipaensis, and eight accessions of A. batizocoi (B genome)
were obtained from the USDA or NCSU germplasm collections. Plants were then grown in greenhouses at the
University of Georgia at Athens. The accessions evaluated are shown in Figure 1. Hybrids were made between
three pairs of A. duranensis accessions, including PI
468200 × PI 468198, PI 468319 × PI 475885, and
PI 475887 × Grif 15036. The hybrid combination
PI 475887 × Grif 15036 was one of the most polymorphic as revealed by using a panel of SSR markers
as described below and thus was selected for subsequent mapping. PI 475887 was originally collected by
Krapovickas, Schinini, and Simpson near Salta,
Argentina during 1980, and Grif 15036 was originally
collected by Williams, Simpson, and Vargas near
Boqueron, Paraguay during 2002 [22]. Crosses were
made by manually emasculating flowers of the female
parent PI 475887 during the late afternoon and pollinating stigmas between 8 and 10 am the following
morning with pollen from the male parent Grif 15036.
An F2 population was developed by self-pollinating
multiple F1 individuals. The intraspecific F2 population
(n = 94) from a hybrid between two A. duranensis
accessions was then used for mapping studies.
Molecular diversity between and within A- and B-genome
diploid species
DNA was isolated from leaf samples of A. duranensis,
A. ipaensis, A. stenosperma, and A. batizocoi accessions
using a modified CTAB method [45,46]. The 60 DNA
samples were amplified using 709 different SSR primer
pairs (GM1-GM709) that had been generated from
sequences reported in the literature [6,29,47-53] and
screened for polymorphisms. SSR markers were genotyped on an ABI3730XL Capillary DNA Sequencer
(Applied Biosystems, Foster City, CA) as described in
a related paper by Guo et al. (Guo Y et al: Comparative mapping in intraspecific populations uncovers a
high degree of macrosynteny between A-and B-genome
diploid species of peanut, Submitted) using forward
primers labelled with FAM, HEX, or TAMRA fluorophores. Microsat [54] was used for construction of a
distance matrix based on the proportion of shared
bands (D = 1 - ps) from 556 primer pairs amplifying
Marker development
Simple sequence repeat (SSR) markers
Single-stranded conformational polymorphism
(SSCP) markers
SSCP markers were developed from genomic DNA templates for previously described NBS sequences isolated
by targeting conserved sequence motifs in NBS-LRR
encoding genes [56,57] and from Arachis unigenes
showing similarity to R-gene homologs identified by mining a peanut transcript assembly [43]. SSCP fragments
were amplified using touch-down PCR and detected by
silver-staining as previously described [58-60]. A total
of 380 SSCP markers were evaluated for polymorphism
between the parents PI 475887 and Grif 15036. The
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resistance gene analog markers are prefixed by either ‘GS’
or ‘RGC’ in the map. cDNA sequences for unigenes
targeted for SSCP marker development in the present
study were deposited in GenBank (Acc. No. GF100476GF100638). One additional marker, the SCAR marker
S197 linked to a root-knot nematode resistance gene in
Arachis hypogaea [43,61] was also mapped.
Development of single nucleotide polymorphism (SNP)
markers
Total RNA was isolated from roots of young seedlings
(up to four trifoliate) and from developing seeds (up to
developmental stage R6) of the two parental genotypes,
PI 475887 and Grif 15036 (alias DUR25 and DUR2,
respectively). cDNA libraries were developed using the
Mint cDNA synthesis kit (Evrogen) and normalized
using the Trimmer cDNA normalization kit (Evrogen).
cDNA sequences were generated by Sanger and 454 GSFLX sequencing methods and assembled using the tool
Mira [62]. Altogether, more than one million cDNA
sequence reads were generated from A. duranensis PI
475887 and Grif 15036. These were assembled into
81,116 unique transcripts (unigenes) (GenBank Accn.
No. HP000001-HP081116). Assemblies were searched
for single nucleotide polymorphisms (SNPs) that fulfilled
the following two criteria: (a) the SNP position is
covered at least by two reads from each genotype, and
(b) at least 80% of the reads call the SNP in the particular genotype. Using these criteria, we identified 8,478
SNPs in 3,922 unigenes. To facilitate the selection of
candidate SNPs for designing and building Illumina
GoldenGate SNP genotyping arrays, putative intron
positions were predicted by aligning Arachis contigs with
Arabidopsis and Medicago genomic DNA sequences
identified by BLAST analyses. SNPs within 60 bp of a
putative intron were eliminated, thereby reducing the
collection of candidate SNPs to 6,789 in 3,264 unigenes
from which 1,236 high-quality SNPs, each representing
separate unigenes, were selected for genotyping. SNPs
were also detected by allele re-sequencing in a subset of
768 conserved legume orthologs identified by coauthors
(R.V. Penmetsa, N. Carrasquilla-Garcia, A. D. Farmer
and D.R. Cook), and 300 of these SNPs were added to
the GoldenGate array. SNP genotyping on the GoldenGate array was conducted at the Emory Biomarker
Service Center, Emory University. The BeadStudio (Illumina) genotyping module was used for calling genotypes. Markers with GC quality scores lower than 0.25
were excluded from subsequent analysis.
Map construction
The program, MSTMap [39] was used to build a core
genetic map including all codominant markers using
the cut-off p-value of 10-12 for clustering markers into
Page 9 of 11
linkage groups. The recombinant inbred line2 (RIL2) algorithm and Kosambi function were used to calculate
genetic distances. The program Joinmap 3.0 [63] was
used to localize the dominant markers and to confirm
the marker order, a range of LOD scores of 5–16 was
used to create groups. The Kosambi mapping function
was used for map length estimations. Markers were
tested for segregation distortion by the chi-square test.
Graphic presentation of the map was drawn using Mapchart 2.0 software [64].
Gene annotation
The cDNA sequences included in the genetic map have
been used to search for homologous genes in the Medicago (www.medicago.org), Uniprot (www.uniprot.org)
and GenBank NR (http://www.ncbi.nlm.nih.gov/genbank)
databases using various blast algorithms. Gene ontology
annotations were also added by searching Medicago
Gene Atlas (http://mtgea.noble.org) and The Arabidopsis
Information Resource (TAIR, www.arabidopsis.org) databases. A significance threshold of E =1e-5 was applied in
all inquiries.
Synteny between Arachis, Medicago, and Glycine
The EST sequences used for marker-development were
compared to the whole genome sequences of Glycine
max and Medicago truncatula to establish synteny.
Sequences for the genomes G. max V5 and M. truncatula MT3.0 were obtained through www.phytozome.net.
The sequences associated with each locus on the A. duranensis peanut map (Additional file 1 and Additional file 5)
were searched against the respective whole genome
sequences using blastn and E < =1e-6. For comparison to
Medicago, only the best match was retained because diploid peanut and M. truncatula are at the same relative
ploidy level. However for Glycine, the two best matches
for each peanut sequence were retained because of the recent polyploidy within soybean and the high level of retention of duplicated genes in the species. Blast hits to
scaffolds or Bacterial Artificial Chromosomes (BACs) not
anchored to the chromosomal assembly in the target
genomes were discarded. Plotting the data and processing of blast results were performed with Visual Basic
programs written for this study.
Additional files
Additional file 1: SNP markers on Illumina GoldenGate array.
Marker ID along with sequence information for OPAs and target ESTs
are provided.
Additional file 2: Comparative genetic mapping of Arachis
duranensis using two different software programs on the same
dataset. Genetic maps were constructed by MSTMap (left) using 1673
co-dominant markers and Joinmap 3.0 (right) using 1724 markers.
Linkage group assignments, marker orders and genetic distances were
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highly consistent, except for the order among a few closely linked loci.
Marker positions determined by Joinmap 3.0 are provided in Additional
file 6.
Additional file 3: Mapped markers with segregation distortion
(p = 0.05) and their position on the map. Column A lists the marker
positions on each chromosome, column B indicates marker name,
column C is the chromosome number, columns D to H list the number
of individuals in each genotype class, and columns J and K provide χ2
and P values, respectively.
Additional file 4: Annotated loci mapped in Arachis duranensis.
Columns B to J include homologs identified in the Medicago (B-D),
GenBank-NR (E-G) and Uniprot-Sprot (H- J) databases. Columns L to S
include gene ontology (GO) terms identified in the three major GO
categories: molecular function (N, O), biological process (P,Q) and cellular
component (R,S).
Page 10 of 11
5.
6.
7.
8.
Additional file 5: Sequences associated with SSR and SSCP markers
for synteny analysis. Column A is the marker name and column B is the
Genebank ID or sequence used as query for synteny analysis.
Additional file 6: Marker positions for the linkage map.
Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
EDN developed the cDNA libraries, organized sequencing and SNP
genotyping, and constructed the initial genetic maps. YG conducted
parental genotyping and constructed final genetic maps. EDN and ST
developed the mapping population. EDN and RAO genotyped the SSR, RGC,
and RS markers. JEB performed the synteny analysis. ST, YG, SK, AFH, NK
developed the SSR markers. SK performed the genetic diversity study. CAT
and DZ assembled the ESTs and nominated SNPs for the genic sequences.
ADF, NCG, RVP and DC nominated the conserved legume ortholog SNPs.
EDN and JEB participated in manuscript writing. DC, HTS and NN
contributed to project design. POA was Co-PI of the project and finalized the
manuscript. SJK was the PI of the project, designed experiments, coordinated
the study, and participated in manuscript writing. All authors read and
approved the final manuscript.
Acknowledgements
This research was supported by funding from the USDA National Institute of
Food and Agriculture National Research Initiative Competitive Grants
Program (#2006-35604-17242) awarded to SJK and POA and by the National
Peanut Board, the Peanut Foundation, the Georgia Seed Development
Commission, and Georgia Research Alliance endowment funding awarded to
SJK.
Author details
1
Institute of Plant Breeding, Genetics and Genomics, University of Georgia,
111 Riverbend Rd, Athens, GA 30605, USA. 2National Center for Genome
Resources, 2935 Rodeo Park Drive East, Santa Fe, NM 87505, USA.
3
Department of Plant Pathology, University of California, Davis, CA 95616,
USA. 4Department of Crop Science, North Carolina State University, Raleigh,
NC 27695, USA. 5Department of Horticulture, University of Georgia, Tifton, GA
31793, USA.
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Received: 22 December 2011 Accepted: 30 August 2012
Published: 11 September 2012
18.
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doi:10.1186/1471-2164-13-469
Cite this article as: Nagy et al.: A high-density genetic map of Arachis
duranensis, a diploid ancestor of cultivated peanut. BMC Genomics 2012
13:469.