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Received: 30 September 2018    Revised: 5 February 2019    Accepted: 1 March 2019

DOI: 10.1111/pbr.12700

ORIGINAL ARTICLE

Genetic diversity, population structure and key phenotypic

traits driving variation within soyabean (Glycine max) collection
in Ghana

Nicholas Ninju Denwar1 | Frederick J. Awuku1  | Brian Diers2 |

Francisca Addae‐Frimpomaah1 | Godfree Chigeza3  | Richard Oteng‐Frimpong1  |
1 4
Doris K. Puozaa  | Michael T. Barnor

1
CSIR‐Savanna Agricultural Research
Institute, Tamale, Ghana Abstract
2
Department of Crop Sciences, University of Soybean [Glycine max (L.) Merrill] is an important oilseed crop worldwide and it has
Illinois, Urbana, Indiana
recently become the crop of interest in Ghana. In this study, 142 soybean accessions
3
International Institute of Tropical
Agriculture, Lusaka, Zambia
were genotyped with 34 SSR markers and concurrently evaluated for five quanti‐
4
West Africa Centre for Crop Improvement, tative and two qualitative phenotypic traits. Twenty‐nine of the SSR markers were
Accra, Ghana polymorphic with mean allele number of 5.3, polymorphic information content (PIC)
Correspondence of 0.51 and gene diversity of 0.55. Molecular analysis based on unweighted paired
Michael T. Barnor, West Africa Centre for group arithmetic mean (UPGMA) clustering and principal coordinate analysis (PCoA)
Crop Improvement, Accra, Ghana.
Email: teye.barnor@gmail.com was similar in explaining the extent of diversity within the accessions. Structure
analysis placed most of the accessions into two subpopulations with 18 (12.7%) as
Funding information
USAID Soybean Innovation Lab admixtures. Principal component analysis (PCA) based on phenotypic traits revealed
two clusters. Both UPGMA clustering‐based SSR data and PCA from phenotypic data
*Communicated by: Bradley Morris
showed similar results. The assembled germplasm is genetically diverse with high
variation in flowering and maturity period, and key yield components which could
be exploited in developing superior varieties well adapted to Ghana and West Africa.

KEYWORDS
gene diversity, germplasm, maturity, population structure, soybean, traits

1 |  I NTRO D U C TI O N
Soybean (Glycine max [L.] Merr.) is one of the world's most import‐
ant legume crops due to its high seed oil and protein concentration
(Satué‐Gracia, Heinonen, & Frankel, 1997; Van Ee, 2009). Although
Ghana is not a major producer of soybean, it has seen a steady rise in
production, from 50,000 MT in 2005 to 142,000 MT in 2015 (MoFA,
2016; MoFA‐SRID, 2007). However, the national average yield of
1.65 t/ha (MoFA, 2016) is far lower than the average of 3.38 t/ha for
Brazil (Statista, 2018) and the 3.34 t/ha obtained in the US (USDA‐
NAS, 2018). The production and utilization of soybean in Ghana
received a boost through the ongoing Soybean Innovation Lab
project funded by USAID’s Feed the Future Programme. Under this
project, the Council for Scientific and Industrial Research‐Savanna
Agricultural Research Institute (SARI) received soybean germplasm
as part of its collaboration with the US Department of Agriculture
and the University of Illinois as part of an effort to genetically im‐
prove the crop. SARI previously received limited amounts of germ‐
plasm from only the International Institute of Tropical Agriculture
(IITA), hence its gene pool was narrow with limited diversity. With this
new germplasm, it is important to study its genetic diversity and suit‐
ability to the poor soils in major growing areas of the country (Buri,
Iassaka, Fujii, & Wakatsuki, 2010; Goldsmith, 2017). Eujayl, Sorrells,
Baum, Wolters, and Powell (2001) indicated that the evaluation of

Plant Breeding. 2019;00:1–11. © 2019 Blackwell Verlag GmbH |  1

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2       DENWAR et al.
variation with DNA markers helps ensure the effective use of ger‐
mplasm for crop productivity improvement/conservation. The use
of new sources of genetic variation to develop improved varieties is
enhanced when the selection of parental genotypes is based on both
phenotypic and genetic dissimilarity (Bisen, Khare, Nair, & Tripathi,
2014). Hence, estimation of variation among soybean germplasm
in Ghana is important to effectively select parental genotypes for
hybridization and development of improved varieties. Dissimilarity
among genotypes can be determined with morphological markers
(Holbrook & Stalker, 2010); however, such markers are less effective
than DNA markers because they are limited in number and may be
subjective and influenced by the environment which could affect in‐
ferences from such experiments (Holbrook & Stalker, 2010).
The advent of polymerase chain reaction (PCR) (Mullis &
Faloona, 1987) has led to the development and utilization of PCR‐
based markers such as simple sequence repeats (SSRs), single nu‐
cleotide polymorphism (SNP), random amplified polymorphic DNA
(RAPDs) and amplified fragment length polymorphisms (AFLPs).
In particular, SSR markers have been widely used for genotyp‐
ing and characterizing soybean (Bisen et al., 2014; Guan et al.,
2010; Kuroda, Kaga, Tomooka, & Vaughan, 2006; Li et al., 2010;
Wang, Run‐zhi, Wan‐ming, & Wei‐jun, 2010). More recently, the
soySNP50K iSelect BeadChip has been developed for the char‐
acterization of soybean genotypes, and quantitative mapping of
desired loci (Song et al., 2013). Genotyping service providers are
moving towards the use of SNP markers but SSR markers con‐
tinue to be used because of their abundance in the plant genome,
simple equipment required, codominant nature and high level of
allelic variation (Bisen et al., 2014; Kuroda et al., 2006; Rakoczy‐
trojanowska & Bolibok, 2004). These traits make SSR markers more
efficient for estimation of dissimilarity and evaluation of genetic
distance. In this study, 142 soybean genotypes comprising released
varieties, advanced breeding lines and plant introductions showing
wide variations for maturity period and seed size were assessed for
their genetic diversity and population structure using SSR markers
and to identify major phenotypic traits contributing to variations
among the accessions.
2 |  M ATE R I A L S A N D M E TH O DS
2.1 | Plant materials
In this study, 142 soybean genotypes (Table S1) comprising 5 re‐
leased varieties, 11 plant introductions and 126 advanced breed‐
ing lines were evaluated on the field for 2 years. These genotypes
were assembled from SARI, Ghana; USA; IITA, Nigeria; Brazil; and
SEEDCO, Zimbabwe.
2.2 | Field evaluation
The accessions were field evaluated for two seasons to assess their di‐
versity in relation to qualitative and quantitative traits of importance
for crop improvement and adaptation to the local environment. The
accessions were arranged in an alpha lattice design in three replicates
at SARI's experimental field, Akokeyiri in Nyankpala for the 2016
cropping season (N09°24.388; W000°59.281) on the 27 June 2016
and 2017 cropping season (N09°23.411; W001°00.234) on the 7 July
2017. Each accession was planted in single row plot of 5 m long with a
spacing of 50 cm between rows and 5 cm between plants within rows.
Fifteen plants per plot were tagged for data collection. The accessions
were evaluated for flower colour, pubescence colour, days to 50%
flowering (plants/plot), height at flowering, days to maturity (90% of
pods turn brown), number of pods/plant, and 100 seed weight.
2.3 | DNA extraction and SSR genotyping
Fresh leaves were harvested from the first trifoliate of ten 3‐
week‐old plants for each genotype and bulked into Ziploc bags
and transported to the laboratory on ice. The leaf samples were
then dried for 3 days using silica gel and ground using bead
beaters. About 0.02 g of ground tissue was transferred into 2‐ml
Eppendorf tubes and genomic DNA extraction was carried out
using the CTAB method (Tiwari, Jadhav, & Gupta, 2012). The
quantity and quality of extracted DNA were checked on a 2%
agarose gel stained with ethidium bromide. The DNA samples
were then diluted to 50 ng/μl prior to PCR amplification. A set
of 34 SSR markers, distributed on 16 of the 20 soybean chro‐
mosomes (Cregan et al., 1999) (Table S1) were used to geno‐
type the accessions. Amplification of DNA was carried out in
a 10‐µl PCR reaction volume (5 µl of x2 VWR Red Taq DNA
Polymerase master mix, 3 µl nuclease‐free water, 1 µl of 10 mM
primer [0.5 µl each of forward and reverse], 1 µl of genomic
DNA). The reaction was amplified in an ABI thermal cycler with
the conditions of denaturation (94°C /30 s), annealing X°C [de‐
pending on primer (Table S1)]/30 s, and extension 72°C/30 s for
a cycle length of 35. PCR products were resolved on a horizon‐
tal 6% polyacrylamide gel electrophoresis system at 120V for
3 hr. Ethidium bromide staining was employed and the image
captured for analysis.
3 | DATA A N A LYS I S
3.1 | Gene diversity
Two methods of scoring bands were employed. The presence and
absence method of band scoring where the presence of bands for
each locus was scored as ‘1’ and the absence as ‘0’. In the second
approach, bands were scored by size, where two clearest bands for
each marker/sample is scored as a/b. Cluster analysis was carried out
using the Jaccard similarity test and unweighted pair group method
with arithmetic mean (UPGMA) clustering method (Durvasula &
Rao, 2018) in DARwin (Perrier & Jacquemoud, 2009). Genetic dif‐
ferentiation parameters including polymorphic information content
(PIC), major allele frequency, heterozygosity and number of alleles
per markers were generated with the aid of PowerMarker V3.2.5
(Liu & Muse, 2005).
DENWAR et al. |
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membership probability ≥ 0.6 were put into the same subpopu‐

3.2 | Population structure
Simple sequence repeat marker data were subjected to structural
analysis using data from 29 polymorphic markers. The geno‐
types were assigned population IDs (1–6) based on their origin
(Table S3). Bayesian clustering analysis was carried out in the
STRUCTURE programme (Pritchard, Stephens, & Donnelly, 2000)
to evaluate the population structure. Three independent runs
were performed for each number of assumed subpopulations (K)
ranging from 1 to 12, using the admixture model correlated for
all frequencies. A burn‐in period of 50,000 and an MCMC run
length of 100,000 for each run for all ranges of Evanno's method
(Earl & vonHoldt, 2012; Evanno, Regnaut, & Goudet, 2005) was
used to estimate the best number of subpopulations (ΔK) that
explains the structure of the genotypes. The run with the maxi‐
mum log likelihood was used to assign genotypes into subpopu‐
lations based on their membership probability. Genotypes with
lation while genotypes with membership probability less than
0.6 were put into mixed groups. The fixation index (Fst) for each
subpopulation by the best ‘K’ was estimated by the STRUCTURE
software.
3.3 | Principal coordinate analysis
The general diversity of the soybean accessions was visualized in
the principal coordinate analysis (PCoA) using the Jaccard dissimilar‐
ity matrix generated in Darwin V6 (Perrier and Jacquemoud‐Collet,
2006) based on molecular data. The two flower colour types, pur‐
ple and white, were scored for each genotype, and given numeri‐
cal identification where purple was represented by 1 and white as
0. Similarly, pod pubescence colour, grey, brown and white, were
also scored as 0, 1 and 2 for grey, brown and white respectively.
These were combined with the five quantitative traits and subjected

TA B L E 1   Genetic differentiation
Major allele
parameters on 142 soybean varieties as
Marker frequency Allele number Allele diversity Heterozygosity PIC
revealed by 29 polymorphic SSR markers
sat_280 0.88 6 0.22 0.08 0.21
satt612 0.38 8 0.71 0.73 0.66
GMES4591 0.50 3 0.59 1.00 0.51
GMES2891 0.34 9 0.80 0.54 0.77
Satt244 0.26 11 0.81 0.77 0.79
Satt545 0.32 7 0.79 0.99 0.76
GMES6224 0.50 4 0.62 0.99 0.55
GMES3177 0.94 3 0.11 0.00 0.10
GMES3757 0.89 2 0.20 0.00 0.18
GMES6842 0.68 2 0.43 0.00 0.34
GMES0676 0.96 4 0.07 0.01 0.07
CSSR18 0.40 11 0.71 0.97 0.65
CSSR438 0.50 4 0.55 0.99 0.45
GMES0707 0.73 3 0.41 0.08 0.35
GMES2038 0.50 3 0.53 1.00 0.42
GMES2036 0.99 2 0.03 0.00 0.03
GMES6270 0.43 5 0.62 0.91 0.55
GMES3941 0.82 3 0.30 0.00 0.26
Sat_210 0.36 8 0.72 0.83 0.68
AW781285 0.51 4 0.60 0.07 0.52
Sat_142 0.35 4 0.73 0.46 0.68
Sat_038 0.40 5 0.66 0.82 0.60
AW310961 0.32 5 0.75 0.38 0.70
Sat_305 0.89 2 0.19 0.00 0.17
Sat_205 0.23 9 0.83 0.70 0.80
Sat_254 0.57 5 0.59 0.11 0.54
Sat_105 0.30 7 0.78 0.80 0.75
Sat_125 0.18 9 0.86 0.54 0.84
Sat_171 0.26 5 0.79 0.85 0.75
Mean 0.53 5.3 0.55 0.50 0.51
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4       DENWAR et al.

to principal component analysis (PCA) in the software, XLSTAT
(XLSTAT, 2015).
4 |   R E S U LT S
4.1 | Gene diversity
Of the 34 SSR markers, 29 (85.3%) successfully amplified polymor‐
phic loci in all 142 soybean genotypes evaluated (Table 1). Allele
frequencies recorded ranged from 2 to 11 with an average of 5.3
alleles per marker and amplified a total of 153 alleles (Table 1). Of
the 29 polymorphic markers, 4 (13.8%) were biallelic (2 alleles/locus)
while the remaining 25 were multiallelic (>2 alleles/locus). The mean
allele frequency of 5.3 alleles per locus is relatively high. Major allele
frequency for the markers used ranged from 0.18 for Sat_125 to 0.99
for GMES2036 with an average of 0.53. Of the 29 SSR markers, 15
used recorded major allele frequency of 0.5 and above. The marker
Sat_125 had the highest allele diversity (0.86), whereas the lowest
of 0.03 was recorded for marker GMES2036 (Table 1) with a mean
gene diversity of 0.55. Polymorphic information content (PIC), a
measure of allelic diversity, ranged from 0.03 for marker GMES2036
to 0.84 for marker Sat_125 with a mean of 0.51. Eighteen SSR mark‐
ers representing 62.1% of the markers used were observed to have
PIC value of greater than 0.5. These markers were the most in‐
formative for discerning differences among the soybean accessions.
Heterogeneity ranged from 0.0 to 1 with an average of 0.50.
4.2 | Population structure
The Evanno method estimated the best ‘K’ to be 2 (Figure 1a),
thereby clustering the genotypes into two subpopulations/Clusters
(Table S3; Figure 1c). The proportion of each of the 6 groupings/
population of genotypes in each cluster is shown in Figure 1b.
Most of the accessions (87.3%) were assigned to a cluster with
the remaining 12.7% classified as mixed. An allele frequency diver‐
gence of 0.108 was estimated between the two subpopulations.
Expected heterozygosity in subpopulation 2 was 0.1942 while
subpopulation 1 recorded expected heterozygosity of 0.310.
The lines from IITA (TGX genotypes) showed great diversity from
structure analysis with the majority belonging to subpopulations

F I G U R E 1   Population structure of 142 soybean accessions. a, Delta K for varying number of sub populations (K) from 1 to 12. b,
Distribution of genotypes among the two clusters revealed by structure analysis. Blue colour represents proportion of genotypes in cluster
1, brown colour represents proportion of genotypes in cluster 2 and ash colour represents the total number of genotype in each grouping.
c, Estimated population structure of the soybean genotypes as revealed by 29 simple sequence markers for (K = 2), red indicates cluster 1,
green indicates cluster 2. 1–6 represents the source/origin of the genotypes as IITA, USA, SEEDCO, SARI, BRAZIL and MIXED respectively
DENWAR et al.
1. Genotypes from the USA were less diverse with most occurring
in subpopulation 2 (S3). 'Jenguma', the most cultivated soybean
variety in Ghana was classified as mixed. Fixation index (FST ), a
measure of genetic differentiation was higher in subpopulation 1
(0.256) which mainly contains the TGX accessions from IITA, than
that of subpopulation 2 (0.0058), which is composed of a mixture
of US and IITA accessions.
4.3 | Cluster analysis
The UPGMA clustering grouped the genotypes into two clusters
(Figure 2). Cluster I is made up of 23 accessions most of which
were from IITA. There is no clear‐cut demarcation for their group‐
ing as members of cluster I show wide variations for phenotypic
and agronomic traits evaluated in this study (Table S1). One geno‐
type (TGX2011‐6F) was found to be an outlier within this cluster;
this genotype is a TGX line of medium maturity (115 days) with 100
seed weight of 11 g (S2). Although its agronomic and phenotypic
traits are not distinct from all the lines in this collection, it was
genetically placed as an outlier (Figure 2). The remaining 119 ac‐
cession grouped into cluster II, which subdivided into several sub‐
clusters. The most distinct subcluster of cluster II was IID which
was genotypes from the USA. These genotypes were photoperiod
sensitive which resulted in their early flowering, early maturing and
shorter plant height at flowering (S1). However, these genotypes
recorded higher seed weight than most of the TGX genotypes. The
most similar genotypes among this group were LD11‐2,170 and
LG15‐2049, with a similarity coefficient of 0.97 and the most simi‐
lar among all the genotypes.
Four locally released varieties ('Quarshie', 'Songda', 'Jenguma'
and 'Salintuya1') and TGX1844‐22E, an advanced breeding line,
were all grouped in subcluster IIF. 'Jenguma', the most cultivated
soybean variety in Ghana was 76% similar to TGX1844‐22E which
has been earmarked for release. 'SoungPungun', another released
variety in Ghana which occurred in cluster I, was the most distant
among the locally released varieties. Subcluster IID is 53% similar to
subcluster IIE, and shared 51% similarity with subcluster IIIF which
harbours most of the locally released varieties. Subcluster IIIA, had
only two genotypes, TGX2004‐10F and TGX2025‐8E, and are con‐
sidered outliers as the line, TGX2011‐6F, in cluster I. Though both
genotypes were similar in days to 50% flowering and plant height at
flowering, they differed markedly in 100 seed weight and number of
days to maturity (S1). With the exception of subcluster IID in which
the genotypes were similar in 100 seed weight, days to 50% flower‐
ing, height at flowering and days to maturity (S1), clustering of other
genotypes based on markers did not show a clear relationship with
the phenotypic traits evaluated in this study.
4.4 | Principal coordinate analysis
Three groupings (I, II and III) can be seen from the PCoA (Figure 3).
Group II could be seen as a subset of group I, similar to subcluster
IID of the UPGMA clustering (Figure 2) and genotypes in group III
|
      5
also matched with those in cluster I of the UPGMA clustering. In
all, the grouping due to PCoA showed concordance with that of the
UPGMA. All accessions from the US grouped together (mostly in
II) with a few occurring within other groups just as accessions from
IITA grouped together in group I, similar to cluster II in the UPGMA
clustering (Figure 2). The locally released varieties; 'Jenguma',
'Salintuya1', 'Quarshie' and 'Songdaa' occurred in group I, similar to
subcluster IIE (Figure 2).
4.5 | Principal component analysis
The phenotypic data revealed high diversity among the genotypes.
Days to 50% flowering ranged from 28 to 63 days averaging 45 days.
The genotypes recorded a height at flowering of 0.12 m for the
shortest and 0.72 m for the tallest with a mean height of  0.31 m. The
number of pods per plant recorded a range of 16 pods to 116 pods
per plant. Days to maturity also recorded a range of 83–175 days
and averaging 102 days to maturity. 100 seed weight recorded as
low as 7.6 g and a maximum of 20.9 g with a mean value of 13.11 g.
Flower colour recorded two categories among the genotypes: white
flower type (16.2%) and purple flower type (83.8%); these were
corded as ‘1’ and ‘0’ respectively. Pod pubescence colour exhibited
three categories: grey (21.8%), brown (64.1%) and white (14.1%) and
were corded as ‘0’, ‘1’ and ‘2’ respectively. All seven phenotypic data
were used for the PCA which saw the first principal component, PC1
accounting for 41.39% of the total variation among the genotypes
(Table 2), with positive associations with some key traits (Table 3).
PC2 accounted for 13.45% of the total variation and associated pos‐
itively with 100 seed weight, number of pods per plant, height at
flowering and pubescence colour (Figure 4).
Principal component analysis placed the 142 soybean genotypes
into two groups (Figure 4). Summary of the characteristics of the
two groups as revealed by the five quantitative traits is shown in
Table 4. Group I comprised mainly of genotypes obtained from the
USA. However, four tropical glycine genotypes, TGX1990‐40F, TGX
1,448‐2E, TGX 1993‐4FN and TGX 2025‐13E were grouped with the
US lines. Genotypes in group I strongly associated with 100 seed
weight, which gave the second highest contribution (18.909) to the
observed variation in PC1 (Table 3). 100 seed weight of the lines
in Group I ranged from 12 to 23 g (Table S1). Group II contained all
the TGX genotypes obtained from IITA and five released varieties
in Ghana, including 'Jenguma', 'Quarshie', 'Songdaa', 'SoungPungun'
and 'Salintuyo1' (Figure 4).
5 | D I S CU S S I O N
Many reports showed that SSR markers were effective for gene di‐
versity studies as a result of the multiallelic and co‐dominant nature
of these markers (Khanh, Anh, Buu, & Xuan, 2013; Priolli, Mendes‐
Junior, Arantes, & Contel, 2002). Inghelandt, Melchinger, Lebreton,
and Stich (2010) and Yu et al. (2011) also noted that many SNPs
markers were required to obtain the same information as a few SSR
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6       DENWAR et al.

LD14-3698
TGX2008-4F
LD14-3702
TGX1990-57F
TGX1990-21F
LD14-6796
TGX1485-1D
SoungPungun
TGX2004-3F
TGX2011-7F
I TGX2006-3F
TGX1989-19F
BRSGO8660
TGX2007-11F
BRSGO8360
TGX1844-19F
SEEDCO23
SEEDCO6
LG14-7959
FTCristaline
NE3400
Ozark
TGX2011-6F
TGX1805-8F
TGX1989-20F
IIH TGX1988-3F
TGX1799-8F
LD00-3309
SEEDCO25
Croton
IIG Liuyuemang
11S-8205
SEEDCO4
TGX1989-11F
TGX1835-10E
Quarshie
TGX1448-2E
TGX1993-4FN
Songda
SEEDCO3
TGX1448-2E
TGX1990-57F
TGX1990-55F
TGX1990-93F
IIF TGX1844-22E
Jenguma
SEEDCO27
TGX1990-114FN
Salintuya1
TGX1448-2E
SEEDCO26
TGX1993-4FN
TGX1845-10E
TGX1903-7F
TGX1989-68FN
TGX1740-2F
TGX2010-15F
TGX1987-14F
TGX2027-1E
TGX2022-4E
TGX2025-9E
TGX2004-13F
Wamini
LD14-3090
TGX1990-46F
TGX1990-95F
TGX2009-12F
TGX1990-40F
TGX1990-52F
TGX1990-80F
TGX1834-5E
TGX1989-41F
TGX1987-62F
IIE TGX1988-5E
TGX2023-3E
TGX2016-3E
TGX1910-6E
TGX1993-4FN
TGX1989-45F
TGX1989-19F
TGX1989-53FN
TGX1989-42F
TGX1989-48FN
TGX1987-10F
TGX1485-1D-C
TGX1989-48FN
TGX1989-49FN
SEEDCO28
TGX2008-4F
TGX1989-45F
TGX1990-78F
TGX1989-40F
TGX2009-16F
TGX1990-40F
LG14-8024
LG15-2243
LG15-4486
LG14-6165
LG15-2775
LD14-1429
LD14-3340
LD14-6766
LD14-1167
LG14-6201
LD11-2170
IID
LG15-2049
LG15-4695
LD14-6444
LD13-8769
LD13-3483
LD14-3218
LD14-6190
LG15-2224
LG15-2214
LG13-3993
LG13-1257
LG15-4418
LG15-2136
TGX2010-3F
TGX2007-8F
TGX2010-12F
TGX1987-62F
TGX2025-19E
TGX2025-13E
TGX2025-6E
TGX1990-110FN
IIC BRS313
TGX2010-11F
TGX2010-4E
TGX1989-68FN
TGX2011-3F
TGX2019-1C
TGX2017-5E
TGX2017-6E
TGX2008-2F
TGX2020-1E
TGX1445-3E
IIB QinghuangzadouNo.7
BRS326
TGX2026-2E
IIA TGX2004-10F
TGX2025-8E

0.93 0.83 0.73 0.63 0.53 0.43 0.33

Similarity

F I G U R E 2   An UPGMA clustering of soybean accessions based on 29 polymorphic simple sequence repeat markers
DENWAR et al.
F I G U R E 3   Principal coordinate analysis displays of 142 soybean accessions. Green coloured lines indicate lines from the US, red coloured
lines indicate lines from IITA, blue coloured lines indicate local Ghanaian lines and black coloured lines indicated mixed accessions. The circle
is to explain the groupings as observed
markers. Consequently, genetic studies that use SSR markers tend
to record higher allelic diversity parameters than studies that use
SNP markers. The mean gene diversity of 0.55 recorded in this study
is similar to that of Huang et al. (2016) who reported a mean gene
diversity of 0.59 using 24 SSR markers to screen 150 soybean acces‐
sions. Similarly, the PIC of 0.51 observed in this study is similar to
the mean PIC of 0.49 obtained by Torres et al. (2015) in a study that
involves genotyping 191 soybean accessions with 22 SSR markers.
Bisen et al., (2014) recorded lower diversity parameters on soybean
including mean gene diversity of 0.2339, heterozygosity of 0.0469
and polymorphic information content of 0.1998 as compared to this
study. With exception of the results reported by Bisen et al., (2014),
genotyping soybean with SSR markers generally give high values of
diversity parameters.
On the other hand, Zhangxiong et al., (2017) obtained gene di‐
versity of 0.34 despite using 5,195 SNP markers to genotype 577
accessions of soybean. Despite a large number of markers and acces‐
sions used in this study, diversity parameters reported were lower
than what has been reported by most SSR marker‐based studies in‐
cluding the present study. Generally, diversity within a collection of
advanced breeding lines and released varieties are lower than among
|
      7
landraces. The 142 accessions used in this study were largely com‐
posed of advanced breeding lines and a few released varieties which
might explain the lower gene diversity as compared to other studies
(Diwan & Cregan, 1997; Li et al., 2010; Xu, Li, Yang, & Xu, 2017) that
used landraces. Gene diversity of 0.55 obtained in this study implies
that the SSR markers used were highly polymorphic and could be
used in soybean breeding programmes.
Structure analysis revealed two subpopulations for the 142 ac‐
cessions, except for 18 accession admixtures. The mean fixation
index (Fst) of 0.256 within subpopulation 1 indicates a higher ge‐
netic diversity within this subpopulation from which parental stocks
could selected to produce variable populations for selection. Very
low mean Fst value (0.0056) was recorded among genotypes in sub‐
population 2 which was dominated by the advanced breeding lines
from the US. The grouping of some IITA lines with the bulk of US
genotypes indicated their close relatedness through common an‐
cestry. Crosses between individuals in subpopulations 1 and 2 could
also adduce the needed variation to enhance genetic gain through
active selection.
Cluster analysis discerned all the accessions as distinct indi‐
viduals thus, agreeing with the suggestion that SSR markers are
 |
8       DENWAR et al.

TA B L E 2   The contribution of traits to

  PC1 PC2 PC3 PC4 PC5 PC6 PC7
the observed variation in each principal
Flower 7.968 33.457 40.219 1.321 0.941 16.074 0.02 component
colour
Pubescence 11.355 20.304 0.332 60.108 2.726 0.451 4.724
colour
Days to 50% 21.191 0.454 0.006 2.14 18.445 15.555 42.209
flowering
Height at 17.268 0.026 2.345 10.659 35.998 31.056 2.648
flowering
No. of pod/ 9.665 37.873 9.157 24.154 4.468 14.505 0.178
plant
Days to 13.645 1.573 39.244 0.672 14.354 22.235 8.278
maturity
100 seed 18.909 6.314 8.697 0.946 23.069 0.123 41.943
weight
Eigenvalue 2.897 0.942 0.874 0.73 0.617 0.543 0.398
Variability 41.39 13.452 12.48 10.43 8.814 7.751 5.682
(%)
Cumulative 41.39 54.843 67.322 77.753 86.567 94.318 100
%

most suitable at clustering germplasm into populations (Hamblin,
Warburton, & Buckler, 2007; Rosenberg, Li, Ward, & Pritchard,
2003). The three major clusters were formed at a similarity coeffi‐
cient of 35%, an indication of the high diversity between and among
accessions evaluated in this study. Largely, the placement of acces‐
sions into major clusters did not reveal a clear relationship with phe‐
notypic traits considered in this study as genotypes with clusters
showed contrasting phenotypic traits. However, placement of ac‐
cessions in subcluster IIID corresponded strongly with phenotypic
data including days to 50% flowering, 100 seed weight and days to
maturity (Table S1).
Similar genotypes were grouped in subcluster IID (Figure 2),
group II of PCoA (Figure 3) and the group I of PCA (Figure 4).
These genotypes were similar in days to 50% flowering and 100
seed weight (S2). The fact that genotypes with similar flowering
response were grouped together by SSR marker‐based cluster
analysis gives some credence to the suggestion that the origin of
genotypes is the principal factor driving differentiation in culti‐
vated soybean (Liu et al., 2017; Roberts et al., 1996). High vari‐
ation in days to flowering among the accessions studied can be
exploited to develop varieties for the different agroecologies in
Ghana, especially, the Guinea and Sudan savannah ecologies with
monomodal rainfall pattern.
Number of pods per plant and 100 seed weight are principal
yield components and key yield determinants of soybean. Zhang,
Xu, Mao, Hu, and Gong (2013) noted that soybean seed weight
was stable across environments, and concluded that most of the
observed phenotypic variation was mainly due to genetic effect.
100 seed weight of released varieties in Ghana ranges from 10 to
13 g while that of the accessions used in this study ranges from 8

TA B L E 3   Principal components for 142

  PC1 PC2 PC3 PC4 PC5 PC6 PC7
soybean accessions based on seven traits
Flower 0.48 −0.561 −0.593 0.098 0.076 −0.295 0.009 showing the loading values
colour
Pubescence 0.574 0.437 −0.054 0.662 0.13 −0.049 −0.137
colour
Days to 50% 0.784 −0.065 −0.007 0.125 −0.337 0.291 0.41
flowering
Height at 0.707 0.016 −0.143 −0.279 0.471 0.41 −0.103
flowering
No. of pod/ 0.529 0.597 −0.283 −0.42 −0.166 −0.281 0.027
plant
Days to 0.629 −0.122 0.586 −0.07 0.298 −0.347 0.181
maturity
100 seed −0.74 0.244 −0.276 0.083 0.377 −0.026 0.408
weight
DENWAR et al. |
      9

4
SEEDCO 6

Qing huangzadou No.7

FT Cristaline
TGX 2007-8F
TGX 2020-1E
TGX 1448-2E
TGX 2009-12F
2 TGX 2025-19E
SEEDCO 23 BRSGO 8660
Quarshie No. of pod/plant
Liu yuemang
LG 15-2136 SEEDCO 27 TGX 2010-3F TGX 2011-3F
TGX 2008-4F TGX 1989-42F
Pod hair colour
LD 14-1167 LG 15-2243 TGX 1835-10E
TGX 2011-7F
LG 14-8024 Croton TGX 1910-6E TGX 1990-57F
Ozark TGX 2010-12F
TGX
SEEDCO 3 2004-10F
1 Wamini TGX 2017-5ETGX 1799-8F
TGX 1485-1D TGX 1448-2E
PC2(13.45 %)

100 seed wtLG 15-2224 TGX 2025-6E

LD 11-2170 TGX 2025-13E TGX 2017-6E TGX 1903-7F
LG 14-6201 NE 3400 TGX 1993-4FN TGX 2022-4E
TGX 1485-1D-C Salintuya 1
TGX 2008-4F
LG 15-2049 LD 14-3218 11S-8205 TGX 2010-4E
TGX1990-40F TGX 2004-3F
LG 14-6165 TGX 2008-2F
BRS 313 TGX 1990-80F TGX 1990-55F
LD 14-3090
Songda TGX 1834-5ETGX 1989-68FNSEEDCO 26 Height at flowering

0
LD 14-6190 BRSGOTGX8360
2010-15F Jenguma
LG 15-2214 LD 13-3483 LD 14-6444
LD… LD 00-3309 TGX TGX 1989-45F
2026-2E TGX 1805-8F Days to 50% flowering
SEEDCO 25 TGX 1988-5E TGX 1990-93F
LG 13-3993 TGX 1990-52F SEEDCO Days to maturity
4 1844-22E
TGX 1989-41F TGX
LD 14-3702 TGX 2025-8E TGX 1990-78F SEEDCO 28
TGX 1990-40F TGX 2004-13F
LD 14-3340 TGX 1448-2E TGX 1990-46F TGX 1989-20F
TGX 1990-95F TGX 1989-19F
LD 14-6766 TGX 1989-53FN
–1 BRS 326
TGX 1989-11F TGX 1990-114FN TGX 1993-4FN
TGX 1845-10E
LG 13-1257 LG 14-7959
LG 15-4695 TGX 2016-3E
TGX TGX
2023-3E
2007-11F TGX…
LG
LD 13-8769 15-4418
LG 15-4486 TGX 1993-4FN TGX 1987-62F
TGX 2027-1E
LD 14-6796 TGX 2025-9E TGX 1989-49FNTGX 1989-68FN
TGX 1988-3F
LG 15-2775 TGX 1989-48FN TGX 1989-45F
TGX 2019-1C Flower colour
TGX 2011-6F TGX 1987-62F
LD 14-1429 TGX 1990-110FN TGX 1740-2F
–2 TGXTGXTGX 1990-57F
… 2006-3F
SoungTGX TGX
Pungun 1987-14F
2010-11F
TGX 1987-10F
TGX 1990-21F

TGX 1989-48FN
–3
–3 –2.5 –2 –1.5 –1 –0.5 0 0.5 1 1.5 2 2.5
PC1 (41.39 %)

F I G U R E 4   Biplot of the first two principal components showing 142 accessions and their associations with phenotypic traits

TA B L E 4   Summary statistics for two

Minimum Maximum Mean
groups revealed by principal component
Traits GRP1 GRP 2 GRP1 GRP 2 GRP1 GRP 2 analysis

Days to 50% flower‐ 28 40 50 63 35 51

ing (days)
Height at flowering 12.00 18.55 38.75 72.80 21.86 35.67
(cm)
No. of pods per plant 16.00 18.00 103.30 116.00 36.43 56.38
Days to maturity 85.00 83.00 129.00 175.00 92.64 105.99
100 seed weight (g) 10.00 7.60 20.90 19.40 15.90 11.82

to 23 g (Table S1). The wide variation in 100 seed weight among
the current collection could be used to improve existing varieties
for yield and other yield components (Han et al., 2012; Hao et al.,
2012; Zhou et al., 2015).
6 | CO N C LU S I O N
The SSR markers used in this study were very informative and
could be useful in future studies including Quantitative trait loci
(QTL) mapping of desired traits. Structure, cluster and PCoAs gave
similar patterns of grouping indicating that the assembled germ‐
plasm largely belonged to two subpopulations. Structure analysis
also showed that a significant proportion of the observed genetic
variation among 142 genotypes was due to population struc‐
ture. PCA of phenotypic traits was useful in elucidating the vital
phenotypic traits driving variation in the assembled germplasm.
Combining molecular characterization with phenotypic charac‐
terization is more informative than either of the two alone, espe‐
cially, for breeders who do not only need knowledge of population
structure and relatedness among genotypes but also need to iden‐
tify useful phenotypic traits to drive hybridization and selection.
More sources of germplasm must be targeted to increase diversity
beyond the two main groups identified in the study to improve
cultivar development at CSIR‐SARI.
AC K N OW L E D G E M E N T
The funding for this work was provided by USAID Soybean
Innovation Lab. We are grateful to Ofosu Asante Aning, Mwintuana
Yelkuro Masuum and Emmanuella Adams of CSIR‐SARI for their as‐
sistance with the laboratory analysis.
 |
10       DENWAR et al.
C O N FL I C T O F I N T E R E S T
The authors declare no conflict of interest.
AU T H O R C O N T R I B U T I O N S
The funding for the work was received through Nicholas Ninju
Denwar. Nicholas Ninju Denwar, Frederick Justice Awuku and
Michael Teye Barnor conceptualized the initial work and the planned
activities of this work. Francisca Addae‐Frimpomaah, Richard Oteng‐
Frimpong and Michael Teye Barnor carried out the field experiment
and collection of the data. Frederick Justice Awuku and Doris K.
Puoza performed the laboratory experiments and organized the
data. Frederick Justice Awuku and Michael Teye Barnor analysis and
interpreted the data generated for both field and laboratory work.
Frederick Justice Awuku and Michael Teye Barnor drafted the initial
manuscript. Nicholas Ninju Denwar and Godfree Chigeza carried out
the initial review of the manuscript. Brian Diers finally reviewed and
shaped the manuscript. All the authors read and approved the final
draft of the manuscript.
ORCID
Frederick J. Awuku  https://orcid.org/0000-0003-2567-5699
Godfree Chigeza  https://orcid.org/0000-0002-9235-0694
Richard Oteng‐Frimpong  https://orcid.
org/0000-0001-6083-1461
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S U P P O R T I N G I N FO R M AT I O N
Additional supporting information may be found online in the
Supporting Information section at the end of the article.   
How to cite this article: Denwar NN, Awuku FJ, Diers B, et al.
Genetic diversity, population structure and key phenotypic
traits driving variation within soyabean (Glycine max)
collection in Ghana. Plant Breed. 2019;00:1–11. https​://doi.
org/10.1111/pbr.12700​