ORIGINAL PAPER
Integrated consensus genetic and physical maps of flax(Linum usitatissimum L.)
Sylvie Cloutier • Raja Ragupathy • Evelyn Miranda •
Natasa Radovanovic • Elsa Reimer • Andrzej Walichnowski •
Kerry Ward • Gordon Rowland • Scott Duguid • Mitali Banik
Received: 28 March 2012 / Accepted: 21 July 2012 / Published online: 14 August 2012
� The Author(s) 2012. This article is published with open access at Springerlink.com
Abstract Three linkage maps of flax (Linum usitatissi-
mum L.) were constructed from populations CDC Bethune/
Macbeth, E1747/Viking and SP2047/UGG5-5 containing
between 385 and 469 mapped markers each. The first
consensus map of flax was constructed incorporating 770
markers based on 371 shared markers including 114 that
were shared by all three populations and 257 shared
between any two populations. The 15 linkage group map
corresponds to the haploid number of chromosomes of this
species. The marker order of the consensus map was lar-
gely collinear in all three individual maps but a few local
inversions and marker rearrangements spanning short
intervals were observed. Segregation distortion was present
in all linkage groups which contained 1–52 markers
displaying non-Mendelian segregation. The total length of
the consensus genetic map is 1,551 cM with a mean marker
density of 2.0 cM. A total of 670 markers were anchored to
204 of the 416 fingerprinted contigs of the physical map
corresponding to *274 Mb or 74 % of the estimated flax
genome size of 370 Mb. This high resolution consensus
map will be a resource for comparative genomics, genome
organization, evolution studies and anchoring of the whole
genome shotgun sequence.
Introduction
Flax (Linum usitatissimum L., 2n = 2x = 30), is an annual
self-pollinated crop that is commercially grown as a source
of stem fibre and seed oil. Flax seed oil is utilized for the
fabrication of various biodegradable products such as high
quality drying oil, paints, varnishes and linoleum flooring.
Flax oil is a rich source of omega-3 fatty acids used as
nutraceuticals and also as a functional food for both humans
and animals. Fibre and oilseed flax belong to the same
species but are morphologically different. Oilseed type flax
plants (linseed) are more branched and shorter than the fibre
type (Gill 1987). Fibre flax is grown mainly in Northern
Europe, Russia and China but linseed is the primary type
grown in Canada, USA, Argentina and India as well as
Russia and China (Gill 1987; Marchenkov et al. 2003).
Development and characterization of flax genetic
resources and assessment of genetic variability are essen-
tial for germplasm conservation and breeding. Flax germ-
plasm collections contain thousands of accessions of
L. usitatissimum and related species, of which, subsets
were assessed for the extent of genetic diversity for mor-
phological characteristics (Diederichsen and Hammer
1995; Diederichsen 2001; Diederichsen and Raney 2006;
Communicated by R. Visser.
Electronic supplementary material The online version of thisarticle (doi:10.1007/s00122-012-1953-0) contains supplementarymaterial, which is available to authorized users.
S. Cloutier (&) � R. Ragupathy � E. Miranda �N. Radovanovic � E. Reimer � A. Walichnowski �K. Ward � M. Banik
Cereal Research Centre, Agriculture and Agri-Food Canada,
195 Dafoe Road, Winnipeg, MB R3T 2M9, Canada
e-mail: [email protected]
S. Cloutier
Department of Plant Science, University of Manitoba,
66 Dafoe Road, Winnipeg, MB R3T 2N2, Canada
G. Rowland
Crop Development Centre, University of Saskatchewan,
51 Campus Drive, Saskatoon, SK S7N 5A8, Canada
S. Duguid
Morden Research Station, Agriculture and Agri-Food Canada,
101 Route 100, Unit 100, Morden, MB R6M 1Y5, Canada
123
Theor Appl Genet (2012) 125:1783–1795
DOI 10.1007/s00122-012-1953-0
Saeidi 2008). A variety of molecular markers including
random amplified polymorphic DNA (RAPD), restriction
fragment length polymorphism (RFLP), amplified fragment
length polymorphism (AFLP) and simple sequence repeat
(SSR) have been developed and used in assessing flax
genetic diversity (Spielmeyer et al. 1998; Oh et al. 2000;
Wiesner et al. 2001; Fu et al. 2003; Adugna et al. 2006; Fu
2006, Roose-Amsaleg et al. 2006; Cloutier et al. 2009,
2012; Uysal et al. 2010; Deng et al. 2010, 2011; Bickel et al.
2011; Kale et al. 2012; Rachinskaya et al. 2011; Soto-Cerda
et al. 2011a, 2011b). While the reports are numerous, the
number of informative markers in each of the studies is
somewhat limited with the majority reporting between 9
and 60 markers only (Cloutier et al. 2012).
SSRs are stretches of DNA consisting of a variable
number of short tandem repeats that are generally co-
dominant, highly polymorphic, multi-allelic, relatively
abundant, heritable, reproducible and reliable (Powell et al.
1996; Hwang et al. 2009). They also show cross-species
usefulness and can be used in closely related species
(Powell et al. 1996; Collard et al. 2005; Varshney et al.
2005). SSRs have been developed from genomic sequences
or Expressed Sequence Tags (ESTs). In flax, Ragupathy
et al. (2011) identified 4,064 putative SSRs from bacterial
artificial chromosome (BAC) end sequences (BES). SSR
markers have also been developed from various flax EST
libraries (Cloutier et al. 2009; Soto-Cerda et al. 2011b) and
from SSR-enriched genomic libraries or other genomic
sequences (Roose-Amsaleg et al. 2006; Deng et al. 2010,
2011; Bickel et al. 2011; Kale et al. 2012; Rachinskaya et al.
2011). There are currently 1,326 SSR markers published in
flax (Cloutier et al. 2012). SSR markers have been used for
the construction of genetic maps of many plant species and
provide dependable landmarks throughout the genome
(Cheng et al. 2009; Studer et al. 2010). In flax, genetic maps
and genetic diversity assessment were achieved with this
type of marker (Fu and Peterson 2010; Cloutier et al. 2011;
Soto-Cerda et al. 2011b). Genetic maps are useful for
evolutionary and comparative studies as they provide both
intra- and inter-species genome wide insights on recombi-
nation rates and gene rearrangements within or across
chromosomes (Ball et al. 2010; Wang et al. 2011).
Only three individual flax linkage maps (Spielmeyer et al.
1998; Oh et al. 2000; Cloutier et al. 2011) have been pub-
lished to date. The linkage map developed by Cloutier et al.
(2011) had 113 markers, mostly SSRs, grouped into 24
linkage groups, while those of Spielmeyer et al. (1998) and
Oh et al. (2000) were based on 213 AFLP markers forming
18 linkage groups and 94 RFLP/RAPD markers grouped into
15 linkage groups, respectively. The limitations of these
maps reside in either or both the type and limited number of
markers. Hence the need exists for a reliable, high density
genetic map of flax that would serve as reference for a wide
variety of applications such as QTL mapping, map based
gene cloning, marker assisted crop improvement, linkage
disequilibrium (LD) mapping, phylogenetic analysis and
anchoring of the whole genome shotgun sequence assembly.
Consensus genetic linkage maps have been constructed for
various plant species including Arabidopsis (Hauge et al.
1993), Brassica (Xu et al. 2010, Wang et al. 2011), barley
(Langridge et al. 1995; Varshney et al. 2007), sorghum
(Mace et al. 2009), wheat (Somers et al. 2004), rice (Antonio
et al. 1996), maize (Cone et al. 2002), red clover (Isobe et al.
2009), lettuce (Truco et al. 2007), rye (Gustafson et al. 2009),
soybean (Hwang et al. 2009), melon (Diaz et al. 2011),
grapevine (Vezzulli et al. 2008), cowpea (Muchero et al.
2009), chickpea (Millan et al. 2010), potato (Danan et al.
2011), eucalyptus (Brondani et al. 2006), Cucurbita pepo
(Zraidi et al. 2007) and Zoysia species (Li et al. 2010). High
density consensus maps are well suited as references for the
incorporation of information from genetically diverse indi-
viduals or multiple populations thus facilitating comparative
analyses across germplasm.
Whole genome physical maps have been constructed for
maize (Messing et al. 2004), Brachypodium (Gu et al.
2009), melon (Gonzalez et al. 2010), grapevine (Scalabrin
et al. 2010), Arabidopsis (Mozo et al. 1999), Brassica rapa
(Mun et al. 2008), soybean (Wu et al. 2004), apple (Han
et al. 2011) and flax (Ragupathy et al. 2011). BAC-based
physical maps have been anchored to genetic maps in a
number of plants such as rice (Chen et al. 2002), maize
(Wei et al. 2009), papaya (Yu et al. 2009), Medicago (Mun
et al. 2006), bean (Cordoba et al. 2010), poplar (Kelleher
et al. 2007), grapevine (Scalabrin et al. 2010) and melon
(Gonzalez et al. 2010), where they were used to order
physical maps and provide a framework for genome
sequence assemblies.
The physical map of the flax genome cv. CDC Bethune,
an oilseed flax variety, consists of 416 fingerprinted contigs
(FPC) spanning *368 Mb, very close to the estimated
genome size of CDC Bethune of approximately 370 Mb
(Ragupathy et al. 2011). The present study was intended to:
(1) construct three independent genetic maps, (2) create an
integrated consensus genetic map and (3) anchor the con-
sensus genetic and physical maps of flax to provide the
backbone information for ordering the whole genome
shotgun (WGS) sequence assembly.
Materials and methods
Plant material, DNA extraction and marker
amplification
Three segregating populations were used for mapping.
CDC Bethune/Macbeth (BM) is comprised of 243 F6-
1784 Theor Appl Genet (2012) 125:1783–1795
123
derived recombinant inbred lines (RILs). The two parents
are current varieties termed ‘conventional’ oilseed types
because they contain 55–57 % linolenic acid, a ‘‘standard’’
amount for oilseed flax varieties. E1747/Viking (EV)
received from S. Knapp (University of Georgia, USA)
consists of 90 F6-derived RILs generated from a cross
between the low linolenic acid line E1747 and the Euro-
pean fibre flax variety Viking. SP2047/UGG5-5 (SU) is an
F1-derived doubled haploid (DH) population of 78 indi-
viduals. SP2047 is a solin breeding line characterized by its
2–4 % linolenic acid content and yellow seeds while
UGG5-5 is a ‘‘high-lin’’ line with 65–70 % linolenic acid
(Banik et al. 2011; Cloutier et al. 2011).
Genomic DNA was extracted from lyophilized leaf tis-
sue (*100 mg fresh) of individual seedlings of all the
segregating and parental lines of the three mapping popu-
lations using the DNeasy 96 plant kit according to manu-
facturer’s instructions (Qiagen Inc, Toronto, ON, Canada).
The genomic DNA was quantified by fluorometer and re-
suspended to a final concentration of 6 ng/ll. Amplification
of template DNA with SSR primers was performed in 384–
well plates in a final volume of 10 ll. The amplification
products were resolved on an ABI 3130xl Genetic analyzer
(Applied Biosystems, Foster City, CA, USA) and scored for
segregation of parental alleles at each SSR locus. A total of
five SNPs and seven genes (fad2A, fad2B, fad3A, fad3B,
dgatA, dgatB and ysc1) were also positioned on the maps
(Cloutier et al. 2011). Protocols and primer information for
SSR markers Lu4 to Lu1193, Lu2097 to Lu2300 and
Lu2331 to Lu3291 were previously described (Cloutier
et al. 2009, 2011, 2012). In addition, SSR markers Lu2001
to Lu2096 were from previously published reports (Roose-
Amsaleg et al. 2006; Deng et al. 2010, 2011) while Lu2301
to Lu2330 were designed from scaffold 505 of the flax
WGS sequence assembly (http://www.phytozome.net), as
previously described (Cloutier et al. 2009, 2012). Refer-
ences for individual markers of the consensus map are listed
(Supplementary Table S1).
Anchoring the genetic and physical maps
Genetic and physical map anchoring was performed using
complementary strategies. First, the CDC Bethune BAC
library was screened with a subset of the SSR primers
positioned on the genetic maps to identify BAC addresses
and their corresponding FPC contigs (Ragupathy et al.
2011). SSR markers Lu2097-Lu2300 and Lu2331-Lu3291
were derived directly from BESs and, as such, were
directly assigned BAC addresses and corresponding FPC
contigs. These BES anchors were confirmed by performing
BLASTn searches of the SSR primer sequences against the
flax WGS sequence assembly (http://www.phytozome.net).
Only perfect matches of both primer sequences to the same
scaffold were considered for anchoring. BLASTn searches
were also performed using the BESs from which the SSRs
were derived and matches with an e value \1e-25 were
considered scaffold anchors. Similarly, primer BLAST and
BLASTn were performed for SSR markers Lu4 to Lu1193
derived from ESTs (Cloutier et al. 2009, 2012). The marker
name convention ‘‘marker name_FPC contig number’’ (e.g.
Lu3156_405) was adopted to indicate positioning of the
markers on the physical map. Markers not anchored to an
FPC contig were labelled ‘0’ (e.g. Lu2312_0). Markers
amplifying two or three loci were labelled with a single
contig number but likely belong to paralogous contigs.
Map construction and linkage analysis
Linkage analysis was carried out independently for each
mapping population using JoinMap 4.0 (Van Ooijen 2006)
with a LOD of 4.0 and a maximum recombination fre-
quency of 40. Marker segregation was tested against the
expected Mendelian ratio of 1:1 using the Chi-square
goodness-of-fit test. For conversion of recombination fre-
quency into map distances expressed in centiMorgans
(cM), the Kosambi mapping function which accounts for
genetic interference from double cross over was used
(Kosambi 1944).
The consensus linkage map was constructed based on
the principle illustrated by Stam (1993) using JoinMap 4.0
(Van Ooijen 2006). LOD scores and pairwise recombina-
tion frequencies were computed for all linkage groups
(LGs) of individual populations. They were then combined
into a single group node in the navigation tree using the
‘grouping node’ command. Consensus LGs were obtained
using the ‘combine groups for map integration’ function
that is based on the presence of a minimum of two marker
loci common to at least two populations on the basis of the
mean recombination frequencies and combined LOD
scores of pairwise data from the three segregating popu-
lations. The consensus map was constructed according to
Alheit et al. (2011) and Gong et al. (2008) using the fol-
lowing parameters: Kosambi mapping function, regression
mapping option, maximum recombination frequency of 40,
LOD[1.0, ripple = 1, third round = yes and goodness-of-
fit jump threshold for removal of loci = 5.0. Comparative
analyses of marker distance and marker order were per-
formed across individual maps and with the consensus map
by visualization of the four final maps obtained as descri-
bed above. The individual homologous LGs from the three
populations were integrated using commonly mapped
markers based on 411 segregating lines (243 RILs from
BM, 90 RILs from EV and 78 DHs from SU). The 15
consensus LGs were numbered LG1 to LG15, in decreas-
ing size order (cM). The population specific LGs were
described using the population acronyms and the same
Theor Appl Genet (2012) 125:1783–1795 1785
123
number as in the consensus map (BMLG1-BMLG16,
EVLG1-EVLG18 and SULG1-SULG15).
Results
Individual genetic linkage maps
CDC Bethune/Macbeth
A total of 389 segregating marker loci were polymorphic in
the BM population, of which, 385 marker loci assembled
into 16 LGs spanning 2,007 cM, for an average density of
one marker locus every 5.2 cM, and 4 marker loci
remained unlinked (Table 1). LGs ranged from 27 to
187 cM and contained 9–40 markers. The dgatA and dgatB
genes were mapped in this population. The population
showed segregation distortion for 56 loci (P \ 0.05) with
equal numbers of loci skewed towards each parent (Sup-
plementary Table S2).
E1747/Viking
The EV population was assayed with 443 polymorphic
marker loci, 442 of which grouped into 18 LGs leaving a
single marker unlinked (Table 1). The length of the LGs
and number of marker loci per LG varied from 10 to
168 cM and 2–46, respectively. The total length of the map
was 1,731 cM with a mean marker density of 3.9 cM
between loci. 77 of the 442 loci diverged significantly from
the expected 1:1 segregation ratio with 33 skewed towards
E1747 and 44 skewed towards Viking (Supplementary
Table S2).
SP2047/UGG5-5
The previously published SU map was based on 125
marker loci assembled in 24 linkage groups spanning
834 cM (Cloutier et al. 2011). The SU map constructed
herein is more saturated and comprehensive with 477
polymorphic marker loci, of which 8 remained unlinked
and the remaining 469 formed 15 LGs totalling 3,044 cM
(Table 1). The length of the LGs and the number of loci
per LG varied from 136 to 362 cM and 20–47, respec-
tively. The approximate average marker density was one
every 6.5 cM. A total of 168 loci showed segregation
distortion (Supplementary Table S2). All LGs contained
distorted markers except SULG13 which contained a
single polymorphic marker in the SU population and
SULG14 (Supplementary Table S2). A total of seven
genes including six from fatty acid biosynthetic pathways
(fad2A, fad2B, fad3A, fad3B, dgatA and dgatB) were also
mapped.
Consensus map
A total of 795 markers generated 821 loci for a locus per
marker ratio of 1.03 because 18 markers identified two loci
and four markers identified three loci. Of the 821 loci
scored in the three populations, 114 were common to all
three populations and another 257 were common to two of
the three populations (Supplementary Table S3). A total of
770 marker loci were assembled into the 15 LGs consti-
tuting the consensus map (Fig. 1; Table 1). Four additional
LGs contained 19 marker loci ordered based on a single
mapping population and 32 markers remained unlinked
(Table 1; Supplementary Table S2). The total length of the
consensus genetic linkage map was 1,551 cM and LGs
ranged from 60 to 170 cM. The consensus map had an
average marker density of one per 2.0 cM. Assuming a
genome size of 370 Mb for CDC Bethune (Ragupathy et al.
2011), the genome wide ratio of physical to genetic dis-
tance averaged 239 Kb/cM, equivalent to an average of one
marker per 481 Kb.
The length of the consensus map was shorter than the
individual population specific maps (Table 1). All LGs of
Table 1 Mapping statistics for the three individual and the consensus genetic maps of flax
Populations No.
individuals
Total no.
marker loci
No. marker
loci in LGsaLength
(cM)
No.
LGsaNo. unlinked
marker loci
No. marker loci in
the consensus map
Percent marker loci
in the consensus map
CDC Bethune/
Macbeth (BM)
243 389 385 2,007 16 4 373 95.9
E1747/Viking (EV) 90 443 442 1,731 18 1 419 94.6
SP2047/UGG5-5 (SU) 78 477 469 3,044 15 8 463 97.1
Consensus 411 821 770 (19) 1,551 15 (4) 32 770 93.8
a Numbers in brackets represent marker loci and LGs belonging to a single population and that did not incorporate in the 15 LGs of the
consensus map
1786 Theor Appl Genet (2012) 125:1783–1795
123
*Lu2857_00.0*Lu25_18720.8*Lu2089_18721.6*Lu3235_53722.2*Lu2026_56022.9*Lu2853_18724.5*Lu2861_18726.6*Lu2091_027.3*Lu2858_18732.1*Lu1066_161_28536.1*Lu49B_036.6*Lu3020_28537.2*Lu427_16139.0*Lu2390_5140.8*Lu2807_16141.8*Lu2808_16142.4*Lu987_044.8*Lu2803_16146.2*Lu2802_161 *Lu955_046.9*Lu1160b_58647.6*Lu56_051.1*fad2A_32551.2*Lu869_052.4*Lu3283_88753.7*Lu2392a_5158.4*Lu2392b_5158.8*Lu2387_5160.5*Lu299_51 *Lu866_063.2*Lu796_5163.3*Lu2393_5163.9*Lu2388_5164.2*Lu2053_5165.4*Lu3220_51270.2*Lu3222_51272.8*Lu1160a_58674.5*Lu2589_9376.8*Lu2597_9478.3*Lu2592_9381.6*Lu2895b_20391.2*Lu2055_11992.0*Lu998_27592.9*Lu2712_12694.4*Lu2698_11998.6*Lu868_119100.9*Lu2374b_45106.2*Lu870_0109.7*Lu2010c_0111.3*Lu2184a_12113.3*Lu2183a_12118.8*Lu3053_297121.9*Lu2010b_0123.5*Lu999_550125.4*Lu1148_550129.8*Lu3231_550130.8*Lu3279_856131.3*Lu46_0132.3*Lu47_0132.8*Lu2687_112136.8*Lu114_181147.1*Lu981_222148.8*Lu943_222150.0*Lu2681_0170.0
LG1
*Lu747b_00.0*Lu2799_1600.1*Lu2794_1601.1*Lu2795_16011.0*Lu2796_16012.7*Lu2800_16022.1*Lu2113_128.5*Lu129_135.1*Lu906_138.9*Lu2115_143.4*Lu2250_2846.6*Lu2247_2850.9*Lu2469_10853.5*Lu344_2855.7*Lu3291_110856.7*Lu2137_4 *Lu2370_4456.8*Lu2188_1457.1*Lu859_1057.4*Lu2135_4 *Lu257_23958.0*Lu2139_458.4*Lu910_458.7*Lu3238_56858.9*Lu3023_28659.0*Lu2144_4 *Lu3022_286*Lu840_286 *Lu3256_67659.1
*Lu3269_77159.3*Lu2067a_059.7*Lu2145_4 *Lu532_459.8*Lu2457a_6660.5*Lu824_460.6*Lu2366_4460.8*Lu2959_23761.7*Lu926Ba_062.9*Lu128_20667.0*Lu209_20668.3*Lu2907_20670.0*Lu2908_20670.5*Lu2909_20671.3*Lu2340_3972.4*Lu2351_3974.3*Lu2349_3978.0*Lu2347_3982.0*Lu2341_3982.5*Lu2346_3983.5*Lu125_3985.9*Lu3068_31386.4*Lu3276_81387.8*Lu2344_3989.1*Lu2352_3992.5*Lu900_3998.1*Lu1028_9099.2*Lu2027_475 *Lu2007_475*Lu2021a_475105.0
*Lu3206_475105.8*Lu324_51106.5*Lu2718_134109.2*Lu2720_134113.3*Lu3205_475122.9*Lu1115_134137.5
LG2
*Lu342_1010.0*Lu445_5425.1*Lu318_4139.2*Lu1039_010.5*Lu3024_28712.3*Lu1161_10115.4*Lu2628_10115.6*Lu452_018.6*Lu2625a_10120.4*Lu899_15038.0*Lu774_15048.1*Lu2764_15049.0*Lu2767_15049.4*Lu821_15051.0*Lu3262_72955.7*Lu64_060.9*Lu2777b_15662.7*Lu373_1664.4*Lu139_1666.0*Lu787a_069.3*Lu2194_1672.3*Lu2689_11373.3*Lu2161_773.6*Lu2047_0 *Lu2044_074.3*Lu2040_0 *Lu2049_0*Lu3117_37474.4
*Lu2063_0 *Lu2163_775.2*Lu3223_51975.7*Lu2164_776.5*Lu3199a_444 *Lu2635_10279.1*Lu106_10282.2*Lu105_10282.5*Lu2633_10283.2*Lu2631_10284.8*Lu3195_44487.7*Lu104_10291.4*Lu3111_36992.5*Lu3151_40197.0*Lu3153_40198.3*Lu3152_40198.8*Lu933_099.5*Lu638_401 *Lu639_401100.1*Lu1144_0103.3*Lu2706_119107.0*Lu3290_1078107.1*dgatA_400108.1*Lu2038_0108.9*Lu658_400110.7*Lu3148_400111.6*Lu3150_400113.4*Lu3146_400117.4*Lu3144_400120.4*Lu2838b_177121.1*Lu509_118122.0*Lu558_118122.4*Lu2693_115127.4*Lu2775b_156127.5*Lu450_115127.8*Lu422a_115128.7
LG3
*Lu3281_8650.0*Lu2966_2439.1*Lu996_24317.4*Lu2006_0 *Lu2004_32*Lu2002_24327.3
*Lu2968_24327.4*Lu2073_67332.4*Lu722B_67334.4*Lu2025_32441.8*Lu2008_32441.9*Lu2059_32442.5*Lu2207_48747.2*Lu3228_54549.2*Lu3229_54550.2*Lu2399_5550.9*Lu2396_5553.8*Lu3252_65953.9*Lu3213_48754.1*Lu2087_48754.2*Lu717_23157.8*Lu2397_5558.4*Lu2942_23158.8*Lu2944_23158.9*Lu2940_23159.3*Lu2943_23166.4*Lu989_67568.7*Lu207_67570.2*Lu3113_37175.5*Lu3116_37176.5*Lu2983_26178.2*Lu2980_26181.2*Lu587_082.8*Lu833_26183.4*Lu1049_084.5*Lu2984_26186.6*Lu851_26187.8*Lu2981_26188.9*Lu2239_2489.5*Lu2043_26191.8*Lu2054_26192.0*Lu2237_2494.6*Lu2031_2494.9*Lu2233_2497.8*Lu2230_24102.2*Lu919_0109.0*Lu2235_24110.4*Lu2076_24111.2*Lu2286_35119.4*Lu2287_35121.2*Lu2009_0 *Lu2011_0121.7*Lu2024_0121.8
LG4
*s18B10_00.0*Lu227_1307.4*Lu643_1309.1*Lu361_13010.7*Lu274B_13013.5*Lu3201_44724.1*Lu223_44724.2*Lu2037b_025.9*Lu2014_028.2*Lu2295_3628.9*Lu505_030.9*Lu176_3634.2*Lu1182_037.1*Lu2288_3642.0*Lu2291_3643.1*Lu2411_5844.1*Lu2292_3645.7*Lu744_3646.9*Lu2297_3648.3*Lu2293_3649.3*Lu2704_11950.2*dgatB_11951.2*Lu2086_051.6*Lu2304_0 *Lu2305_051.8*Lu2752_14651.9*Lu3225_53752.2*Lu922_052.4*Lu2362_4352.5*Lu2365_4352.8*Lu2364_4353.2*Lu2017_053.5*Lu2466_7053.9*Lu2700_11954.7*Lu3132_38554.9*Lu2884_202 *Lu2889_20255.2*Lu2318_0 *Lu2886_20255.3*Lu2885_20255.4*Lu2883_20255.9*Lu2887_20256.1*Lu623_20256.3*Lu3248_59656.4*Lu2890_20256.6*Lu41_20257.1*Lu164_3057.6*Lu2255_3058.0*Lu266_3058.5*Lu2403_5870.4*Lu2408_5873.5*Lu738_5876.2*Lu652_5879.1*Lu2409_5880.7*Lu2405_5885.7*Lu2512_7698.8*Lu968_55101.6*Lu330_76104.8*Lu2509_76108.0*Lu2516a_76108.7*Lu682_76110.4*Lu2068_76112.4
LG5
*Lu1006_590.0*Lu1177_00.9*Lu3287_9982.5*Lu2420_596.5*Lu2418_598.7*Lu2917b_2149.6*Lu2084_5910.4*Lu1179_5910.8*Lu502_8616.3*Lu699_38026.3*Lu1094_38026.6*Lu1107_197 *Lu944_19730.7*Lu2608_9732.3*Lu3014_28037.1*Lu3013_28039.2*Lu2545_8139.8*Lu442a_044.3*Lu2169_945.8*Lu2544_8146.6*Lu2548_8148.1*Lu2972_24848.2*Lu1178_2748.6*Lu456_775 *Lu2550_8149.2*Lu457_775 *Lu2242_2749.4*Lu2072_2749.8*Lu2039_2749.9*Lu2613_9750.8*Lu918_051.0*Lu3267_75751.3*Lu728b_2751.5*Lu69_0 *Lu2542_81*Lu2549_81 *Lu2543_8152.2
*Lu2071_81 *Lu2064_8152.6*Lu836_053.4*Lu2975_24853.7*Lu2974_24854.9*Lu2971_24857.5*Lu2556_8262.5*Lu1002B_8262.6*Lu2561b_8263.2*Lu2560_8263.4*Lu2564_8264.1*Lu2565_8267.5*Lu2553_8268.6*Lu2554_8269.4*Lu2555_8272.0*Lu2557_8272.6*Lu3057a_30373.0*Lu60_073.7*Lu861_59184.1*Lu1112_17184.8*Lu3091_32996.6*Lu2078_329_10107.3
LG6
*Lu260_790.0*Lu2532_792.7*Lu2535_793.6*Lu2536_795.7*Lu2534_795.8*Lu2083_796.7*Lu2540_797.2*Lu402_797.9*Lu675_7910.6*Lu2533_7910.8*Lu511_7910.9*Lu2032_011.6*Lu741_7913.3*Lu1146a_7913.7*Lu2825b_17518.3*Lu2832_17521.1*Lu146_17523.9*Lu138_175 *Lu151_17524.0*Lu235_23024.9*Lu2810_16627.3*Lu2827_17528.8*Lu1055_7738.2*Lu3181_43640.2*Lu296_43640.8*Lu1022_43642.3*Lu3180_43642.7*Lu672_43643.4*Lu3184_43644.1*Lu3178b_43646.0*Lu2815_17051.7*Lu2812a_17052.5*Lu585B_10853.7*Lu2065_054.7*Lu2651_10855.6*Lu2652_10856.2*Lu2654_10856.3*Lu557_45460.8*Lu3100a_35366.8*Lu1124_8471.2*Lu2571_8476.6*Lu2658_10880.0*Lu2648_10880.9*fad3A_8483.8*Lu44E4_8484.7*Lu2003_28190.0*Lu566_28191.6*Lu449_28192.1*Lu3016_28192.5*Lu3017_28196.6*Lu3266_73399.6*Lu58a_257104.3
LG7
*Lu595_1080.0*Lu2561c_8212.0*Lu2659_10814.8*Lu2649_10818.9*Lu2840_17824.8*Lu339_17825.7*Lu1171_17827.6*Lu2561a_8229.0*Lu265_73637.1*Lu2030_8238.2*Lu3057b_30340.1*Lu3059_30344.0*Lu2563_8245.4*Lu3157_40547.9*Lu3156_40551.9*Lu633_053.2*Lu2428_6053.4*Lu2957_235 *Lu2268_3154.7*Lu2056_21756.0*Lu2923_21756.3*Lu2578_8856.6*Lu2203_1857.1*Lu2103_4664.9*Lu857_4665.3*Lu2918_21566.1*Lu2921_21567.6*Lu2082_4670.2*Lu2098_4670.8*Lu2105_4672.1*Lu2102_4672.7*Lu2306_4673.0*Lu2313_073.5*Lu2312_0 *ysc1_0*Lu2307_073.6
*Lu2316_4674.2*Lu928_0 *Lu2106_4674.3*Lu2317_074.8*Lu2320_075.1*Lu2326_075.3*Lu2330_076.4*Lu447_21576.5*Lu2329_077.2*Lu178_4678.5*Lu2424_6079.8*Lu2618_9880.8*Lu2101_4683.5*Lu2431_6084.5*Lu2430_6084.6*Lu2156_584.7*Lu2429_6085.2*Lu3189_44187.9*Lu625_6088.7*Lu1077_17488.8*Lu2425_6090.0*Lu2820_17491.0*Lu2823_17493.0*Lu2822_17493.8*Lu3280_86494.2*Lu2745_14194.7*Lu2587_9195.7*Lu2714_13098.0*Lu963_9199.0*Lu2515_76100.4*Lu1103_60103.0*Lu2338a_38103.9
LG8
Fig. 1 Consensus genetic map
of flax integrated from three
mapping populations. Numbers
to the left of each linkage group
represent Kosambi map units
(cM). Locus names followed by
their FPC contig anchor
separated by an underscore are
on the right. Linkage groups are
in decreasing size order
Theor Appl Genet (2012) 125:1783–1795 1787
123
the consensus map were constructed based on markers
shared among the three populations except LG13 which
was constructed mostly with markers from the BM and EV
populations because all markers, with the exception of
Lu850, were monomorphic in the SU population. The
consensus map displayed a few gaps that were mostly
smaller than 10 cM. The largest gap of 20.8 cM is in LG1
(Fig. 1; Supplementary Table S1). The marker orders were
consistent between the three independent genetic maps
with some local inversions.
*Lu2917a_2140.0*Lu628_786.5*Lu14_08.0*Lu927_7812.0*Lu891_7812.7*Lu140_267 *Lu1041_013.4*Lu756_26715.1*Lu2991_26716.6*Lu2041_018.0*Lu2042_018.2*Lu2694_11518.7*Lu2997_26719.9*Lu2992_3623.9*Lu2996_26727.0*Lu220_26727.3*Lu2779_15630.0*Lu2780_15630.6*Lu2096_031.6*Lu867_031.7*Lu926Bb_032.4*Lu2774_15633.4*Lu2880_19734.7*Lu2778_15637.0*Lu2775a_15637.7*Lu2985_038.3*Lu2773_15639.7*Lu2090_040.2*Lu2081_7340.5*Lu2482_7340.9*Lu2486_7341.5*Lu2600_9741.7*Lu2787_15641.9*Lu2485_7342.4*Lu2127a_242.7*Lu2183b_1243.0*Lu2183c_1243.3*Lu1135_7344.1*Lu263_9744.5*Lu896_044.8*Lu439_73 *Lu728a_16145.5*Lu2612_9746.8*Lu787b_15654.7*fad3B_20757.2*Lu58b_25758.8*Lu2914_20761.7*Lu2913_20764.9*Lu275_068.6*Lu206b_30669.7*Lu1052_070.0*Lu203b_30670.7*Lu765Bb_30672.9*Lu3063_30673.3*Lu2574_8475.3*Lu2911_207 *Lu803_076.6*Lu3064_30677.4*Lu1151_079.0*Lu381_20183.4*Lu3289_106484.4
LG12
*Lu2046_380.0*Lu2333_382.1*Lu2332_383.5*Lu2331_386.7*Lu2867_1889.5*Lu11_010.6*Lu2019_11111.0*Lu2676_11113.5*Lu2679a_11120.7*Lu2673_11121.0*Lu83_11121.4*Lu291_023.9*Lu575_11127.5
*Lu3218_49745.2*Lu935_32148.6*Lu3217_49750.6*Lu2850_18751.4*Lu568_052.0*Lu512_32155.1*Lu785_32155.8*Lu13_32157.9*Lu3078_32159.1*Lu325_32160.1*Lu934_061.1*Lu292_061.2*Lu2580_8969.1*Lu3003_26971.5
*Lu2127b_282.2*Lu2118_284.2*Lu2123_285.6*Lu1165_286.2
LG11
*Lu2472_710.0*Lu1158_712.1*Lu668_713.7*Lu2149_510.0*Lu685_512.9*Lu2155_515.1*Lu2157_517.2*Lu2154a_518.8*Lu2162_727.5*Lu2901_20430.5*Lu3099_353 *Lu2052_3231.5*Lu3100b_35331.7*Lu273_3234.4*Lu3007_26934.8*Lu657_3234.9*Lu1168_10435.8*Lu3120_37936.4*Lu2725_13740.1*Lu2728_13742.1*Lu2732_13742.5*Lu458_13742.7*Lu2731_13746.0*Lu2051_13747.1*Lu2929_22153.0*Lu2928_22155.0*Lu2926_22155.2*Lu483_22156.0*Lu37b_22158.6*Lu1117_059.4*Lu1116_059.6*Lu1176_22160.6*Lu2092b_062.3*Lu2265_3164.9*Lu2016_067.5*Lu2050_068.5*Lu2264a_3169.7*Lu2270_3173.0*Lu1043_78074.2*Lu1136_3174.4*Lu1042_78075.4*Lu2272_3176.2*Lu371B_3781.7*Lu2360_4282.6*Lu804_14184.6*Lu2746_14187.7
LG10
*Lu2437_610.0*Lu2451_611.5*Lu213_6110.8*Lu2262_3114.4*Lu2438_6117.4*Lu2453_6118.7*Lu2443_6120.0*Lu2447_2825.3*Lu2758_14925.4*Lu2936_23026.1*Lu2450_6127.2*Lu2448_2827.5*Lu2058_029.4*Lu801_6129.6*Lu181_3130.1*Lu2446_2831.6*Lu519_77 *Lu526_7731.7*Lu2523_7732.4*Lu3097_33832.8*Lu2524_7733.0*Lu2739_14036.0*Lu2741_14136.5*Lu3082_32339.1*Lu3083_32339.3*Lu144b_0 *Lu3085_32339.6*Lu2809a_16641.9*Lu2361_4242.4*Lu2449_7743.0*Lu3199a_44447.7*Lu2828_175 *Lu2824_17552.1*Lu757_230 *Lu798_23056.2*Lu2939a_23056.5*Lu895_061.5*Lu2168_964.0*Lu2878_19664.9*Lu1146b_19670.0*Lu2538_7985.8*Lu3244_59287.5*Lu932_490 *Lu1125_84190.6*Lu283_84192.0*Lu3216_49093.4*Lu91_49094.6
LG9
*Lu2621_990.0*Lu897_02.1*Lu3251_6305.0*Lu3209_4825.6*Lu3210_4825.8*Lu3212_4826.1*Lu2045_08.1*Lu2373_4513.0*Lu2377_4514.9*Lu808_015.3*Lu3043_29017.2*Lu3040_29020.5*Lu850_29025.0*Lu793_29026.2*Lu3033_29026.7*Lu514_29026.9*Lu3046_29027.9*Lu3219a_49728.8*Lu9_029.8*Lu813_29030.1*Lu3038_29032.2*Lu3036_29032.4*Lu786_10133.1*Lu3103_35533.4*Lu225_35540.3*Lu2679b_11141.0*Lu601b_35543.0*Lu2625b_10143.6*Lu444_18847.1*Lu476_18847.9*Lu461_18848.3*Lu2863_18851.0*Lu684_18851.6*Lu2865_18853.0*Lu613_18853.3*s7F06_056.7*Lu2862_18861.6*Lu2020_8964.1*s16E2_067.8*s19C1c3_068.5*Lu701_8968.7*Lu1044_8970.1*Lu1044B_8970.2*Lu959_8970.8*s19C1c1_076.0
LG14
*Lu2219_210.0
*Lu2223_2111.8
*Lu2216_2122.0*Lu2074_0 *Lu197_2125.9*Lu2468a_7027.4*Lu2639_10327.6*Lu2638_10328.1*Lu650_2028.9*Lu2459_70 *Lu2467_7029.3*Lu2196_1629.5*Lu2463_7030.8*Lu485_032.6*Lu2279_3237.2*Lu2771_15442.1*Lu805_1146.5*Lu2468b_7049.4
*Lu2176_1166.0
*Lu2012_90 *Lu2021b_9076.2
LG13
*Lu2697b_1180.0*fad2B_4052.1*Lu113_1186.6*Lu2695_1187.4*Lu2696_1187.8*Lu462a_21210.9*Lu838_21211.0*Lu1172_11811.7*Lu1007_4013.5*Lu2354_4017.1*Lu359_017.9
*Lu2010a_19031.9*Lu2001_034.3*Lu1127_12134.9*Lu2965_24335.4*Lu2382_5035.9*Lu357_4236.0*Lu2057_69236.8*Lu1163_12137.7*Lu2383_5039.0*Lu2707_12140.7*Lu3028_28742.6*Lu3185_44045.2*Lu3026_28746.4*Lu3186_44048.0*Lu510_44051.2*Lu2931_22652.8*Lu271_22653.6*Lu637_22657.2*Lu1001_7457.9*Lu451_058.4*Lu2497_7459.9
LG15
Fig. 1 continued
1788 Theor Appl Genet (2012) 125:1783–1795
123
The percentages of distorted loci in BM, EV and SU
populations were 15, 17 and 36 %, respectively (Supple-
mentary Table S2). Out of the 770 loci of the consensus
map, 292 (38 %) loci exhibited segregation distortion in at
least one of the three mapping populations (Supplemen-
tary Table S2). The presence of a large number of dis-
torted loci seems to have caused the large gap in LG11.
Although LG8 carried the most distorted loci (76 %),
the length of that LG was not affected as compared to the
bi-parental populations where marker segregation was
Mendelian, although some local rearrangements of closely
linked markers were observed. Overall, only a few
ambiguities were identified with respect to marker posi-
tion compared to individual maps. Markers Lu359_0 and
Lu2354_40, common to all three populations, mapped at
the proximal end of each individual population LG15.
However, these markers were placed at internal positions
(41.977 and 42.829 cM) in the consensus LG15 (Supple-
mentary Table S1).
Anchoring genetic and physical maps
Of the 770 loci in the consensus genetic map, 670 were
anchored to 204 of the 416 FPC contigs of the physical
map (Ragupathy et al. 2011) corresponding to 274 Mb or
74 % of the flax genome (Fig. 2). Twenty-one of the 204
FPC contigs were anchored at more than one map location
because some markers amplified two or three polymorphic
loci (labelled with a small a, b or c suffix in Fig. 1). FPC
contigs were anchored with 1–14 markers. Examples of
FPC contigs anchored with 14 markers include FPC
contigs 79 and 82 estimated at 2.836 Mb with 205 BAC
clones and 3.192 Mb with 265 BAC clones, respectively.
The largest FPC contig (21), estimated at 5.562 Mb,
consisted of 437 BAC clones and was anchored with four
markers. Sixty-seven FPC contigs contained a single
marker.
Discussion
Comparison of individual and consensus maps
Three genetic maps of flax have been published to date: a
213 AFLP marker-based map of 18 LGs covering
1,400 cM (Spielmeyer et al. 1998), a 1,000 cM RFLP/
RAPD map with 94 markers assembled into 15 LGs (Oh
et al. 2000) and a 113 marker map containing EST-SSRs,
SNPs, genes and one phenotypic trait grouped into 24 LGs
and spanning 834 cM (Cloutier et al. 2011). Major QTL for
fusarium wilt (Spielmeyer et al. 1998), for fatty acid
composition and seed coat colour (Cloutier et al. 2011)
were identified using these genetic maps. The increased
marker density of the SU map from 113 (Cloutier et al.
2011) to 469 (this publication) significantly improved the
map by bridging gaps and thereby reducing the number of
LGs from 24 to 15 while increasing the map coverage from
834 to 3,044 cM which promises to enhance QTL detec-
tion. Collinearity with two additional genetic maps (BM
and EV) further confirmed the accuracy of the groupings.
The three maps were largely collinear with few marker
inversions. Although some LGs or portions thereof were
fixed in some of the individual populations (e.g. LG4,
LG13 and LG15 in SU; LG7, LG9 and LG15 in BM; LG15
in EV), the consensus map successfully bridged LGs and
resulted in good coverage across the genome with few
gaps. Some local inconsistencies of marker order such as
small inversions or local rearrangements between individ-
ual and consensus maps were observed, particularly in
closely linked markers and markers located at the distal
ends of LGs as previously reported for rye (Studer et al.
2010), cotton (Xu et al. 2008), Zoysia (Li et al. 2010),
grapevine (Vezzulli et al. 2008) and Eucalyptus (Brondani
et al. 2006). The single most striking discrepancy resided in
the total length of the SU map which exceeded the size of
the BM and EV maps by more than 1,000 cM. The higher
Fig. 2 Distribution of the genetic markers of the consensus map
across the FPC contigs of the physical map (Ragupathy et al. 2011). A
total of 204 of the 416 FPC contigs (x axis) were anchored by 670
marker loci (dots). The length of the contigs (Mb) is on the left y axis.
From 1 to 14 marker loci (dots) were anchored onto each FPC contig
represented on the right y axis
Theor Appl Genet (2012) 125:1783–1795 1789
123
percentage (36 %) of distorted markers of this DH popu-
lation, i.e. at least twice as high as the other two popula-
tions, may be responsible for the artifactual expansion of
the genetic map length (Garcia-Dorado and Gallego 1992;
Zhu et al. 2007; Li et al. 2011).
Comparative and consensus mapping are advantageous
to obtain an unbiased linkage map representing the genome
under investigation. As discussed above, mapping of
markers employing multiple populations provides
increased genome coverage because it is unlikely that
multiple parents would all be fixed (monomorphic) in the
same genomic regions. Also, overall population size
afforded by multiple populations increases the chances of
capturing recombination events, the foundation of genetic
mapping. Reports of overrepresentation of localized
crossover promoting 13mer motifs (Myers et al. 2008) in
recombination hot spots of 1–2 kb (Ptak et al. 2005), and
the influence of highly polymorphic trans-acting loci such
as PRDM9 on the activation of those recombination hot-
spots in human (Baudat et al. 2010; Paigen and Petkov
2010) indicates the importance of the genomic background
in crossover frequencies. Considering crossing over as a
fundamental cellular process conserved across eukaryotes,
variability for distribution of recombination hot spots and
its genetic determinants can be determined using multiple
populations. Comparative mapping can also offer evidence
for duplications or chromosomal rearrangements (Sewell
et al. 1999). As a consequence of merging of datasets from
three populations, the consensus map had fewer and
smaller gaps compared to the individual genetic maps,
hence it was more comprehensive. Fatty acid desaturase
genes fad2A, fad2B, fad3A and fad3B, diacyl glycerol
transferase genes dgatA and dgatB and seed coat color gene
ysc1 were positioned to seven different LGs of the con-
sensus map. The majority were polymorphic in a single
population but common neighbouring polymorphic mark-
ers permitted their integration in the consensus map,
illustrating another advantage of consensus mapping. Flax
has a relatively low level of genetic polymorphism, indi-
cating a lower degree of genome divergence (Deng et al.
2010; Cloutier et al. 2011; Kale et al. 2012), unlike crops
like maize where extensive molecular variation has been
reported, primarily due to the activity of transposable ele-
ments (Llaca et al. 2011). The use of multiple populations
followed by consensus mapping greatly increases marker
saturation, a valuable feature for all mapping applications,
for understanding the LD structure across genomes and
germplasm characterization by association mapping (Soto-
Cerda and Cloutier 2012).
The present consensus map of 770 SSR markers repre-
sents a major improvement over the low resolution phy-
logenetic analyses published to date with other marker
types (McDill et al. 2009; Fu and Allaby 2010) and those
published with few SSR markers within Linum usitatissi-
mum (Wiesner et al. 2001; Cloutier et al. 2009) and across
Linum species (Fu and Peterson 2010; Soto-Cerda et al.
2011b). Pale flax (Linum bienne Mill, L. angustifolia Huds)
is the wild progenitor of cultivated flax. Both have similar
karyotypes bearing equal numbers of chromosomes
(2n = 2x = 30) (Muravenko et al. 2003) and interspecific
crosses between them produce fertile progeny (Gill and
Yermanos 1967; Diederichsen and Hammer 1995). Pale
and cultivated flax have been inferred to differ by a single
translocation event (Gill and Yermanos 1967). The
exceptionally high transferability (97 %) of EST-SSRs
from cultivated flax to L. bienne supports the assignment of
pale flax to the primary gene pool (Diederichsen 2007; Fu
and Peterson 2010; Soto-Cerda et al. 2011b). A genetic
map for pale flax or an interspecific cross map has yet to be
produced. The current availability of SSR markers com-
bined with their cross applicability should allow for an in-
depth analysis of genetic diversity in L. bienne which
should be useful to explore its potential to widen the gene
pool of cultivated flax to meet breeding objectives.
Linum usitatissimum is a self-pollinated diploid species
which, like a number of crop genomes, is an ancient
polyploid (Blanc and Wolfe 2004; Paterson et al. 2004;
Pfeil et al. 2005; Sterck et al. 2005; Gong et al. 2008; Soltis
et al. 2009; Schmutz et al. 2010; Jiao et al. 2011; Lin and
Paterson 2011). The remnant of ancestral whole genome
duplication is reflected by the fact that a subset of SSR
markers amplified two paralogous loci, although in most
cases, only one was polymorphic (Cloutier et al. 2009,
2012). Mapping of the markers that amplified multiple
polymorphic loci revealed ancestral chromosomal rear-
rangements resulting from paleopolyploidization events as
noticed in LG6 and LG8. The existence of duplicated
regions in consensus linkage groups LG6 and LG8
delimited by Lu2561 and Lu3057 markers indicate signa-
tures of ancient duplication (Supplementary Figure 1).
Analyses of a large collection of flax ESTs also corroborate
the ancient duplication of flax (Venglat et al. 2011), as
exemplified by the duplicate nature of the genes of the fatty
acid biosynthetic pathway (Cloutier et al. 2011). Global
comparative analysis of the scaffolds of the WGS sequence
assembly promises a more comprehensive picture of the
events that have shaped the flax genome through evolution.
Distorted markers
The segregation distortion in the three populations ranged
from 15 to 36 %, an intermediate level, comparable to
extent of distorted markers reported in common bean
(37.3 %, de Campos et al. 2011), maize (19–36 %, Lu et al.
2002), red clover (5.8–45 %, Isobe et al. 2009), Medicago
truncatula (27 %, Thoquet et al. 2002) and peanut
1790 Theor Appl Genet (2012) 125:1783–1795
123
(8.5–22.8 %, Hong et al. 2010) but higher than C. pepo
(3.7 %, Zraidi et al. 2007; Gong et al. 2008), Brassica rapa
(2.6 %, Song et al. 1991), grapevine (7–11 %, Doligez
et al. 2006) and globe artichoke (13 %, Portis et al. 2009).
Species such as Arabidopsis (43.0 %, Reiter et al. 1992),
cotton (71 %, Lacape et al. 2009), tomato (68 %, Paterson
et al. 1988), perennial ryegrass (32–63 %, Anhalt et al.
2008), Zoysia (43.7 %, Li et al. 2010) and cowpea (41 %,
Muchero et al. 2009), all displayed substantially higher
percentages of markers deviating from the expected seg-
regation ratios.
Non-Mendelian segregation ratios arise from chromo-
somal rearrangements, gametic competition, embryo via-
bility and various physiological causes (Xu et al. 1997;
Gonzalo et al. 2005; Portis et al. 2009) and, inadvertently,
also from sampling errors (Lorieux et al. 2000). Segrega-
tion distortion has been associated more strongly with
genetic effects as opposed to population structure or mar-
ker type (Anhalt et al. 2008).
Among the three flax populations reported here, the DH
SU population had the highest proportion of distorted loci,
similar to DH populations of rice (31.8 %, Xu et al. 1997)
and Brassica (22–49 %, Wang et al. 2011), but lower than
alfalfa (68 %, Li et al. 2011). The higher proportion of non-
Mendelian markers in DH populations may be attributed to
selection for tissue culture responsiveness loci (Xu et al.
1997; Alheit et al. 2011). Even though they were located on
all LGs, distorted markers were not randomly distributed
but were clustered within LGs (Cloutier et al. 2011;
Cordoba et al. 2010; Li et al. 2011) supporting the cause of
selection rather than experimental errors (Li et al. 2011), a
view further emphasized by higher proportions of distorted
markers on specific LGs such as LG2 (36/65), LG8 (52/68)
and LG10 (32/46). Chromosome specific uneven distribu-
tion of markers has been reported for triticale chromosome
2A and 1R (Alheit et al. 2011); LG1, LG2 and LG3 of
Medicago truncatula (Studer et al. 2010) and M20 and M32
of Zoysia (Li et al. 2010). Clustering of distorted markers
was also documented in lettuce (Truco et al. 2007), Euca-
lyptus (Brondani et al. 2006) and peanut (Hong et al. 2010).
Map distance and map order can both be affected by
segregation distortion (Lyttle 1991; Zhu et al. 2007) as was
observed in the SU population where large gaps were
observed between blocks of non-Mendelian markers adja-
cent to blocks of markers with non-skewed segregation,
which accounted in part for the overestimation of the map
length for this population. Elimination of non-Mendelian
marker loci was suggested to improve mapping accuracies
(Zhu et al. 2007; Xu 2008). However, such an approach
would decrease the number of markers available and
reduce coverage of some genomic regions, hence dimin-
ishing the map saturation (Zhu et al. 2007; Xu 2008). Here,
we clearly demonstrated that consensus mapping was a
powerful way to correct for mapping inaccuracies caused
by non-Mendelian markers because consensus mapping
takes into account segregation data from multiple popula-
tions including common markers with Mendelian segre-
gation in at least one population.
Anchoring genetic and physical maps
The physical map of flax is comprised of 416 FPC contigs
spanning *368 Mb (Ragupathy et al. 2011). A total of 670
markers were anchored to 204 FPC contigs representing
*274 Mb, i.e. 74 % of the estimated 370 Mb genome of
CDC Bethune, comparable to papaya (72.4 %, Yu et al.
2009), apple (60 %, Han et al. 2011) and grapevine (72 %,
Scalabrin et al. 2010) genomes and exceeding the extent of
anchoring reported in Medicago truncatula (32 %, Mun
et al. 2006), Populus trichocarpa (22 %, Kelleher et al.
2007), Prunus (15.5 %, Zhebentyayeva et al. 2008), bean
(8 %, Cordoba et al. 2010) and melon (12 %, Gonzalez
et al. 2010). Although sufficient to provide initial ordering
of the WGS sequence assembly into bins, the level of
anchoring of the physical and genetic maps of flax pre-
sented herein falls short of the requirement for high accu-
racy ordering and orienting of the scaffolds of genomic
sequence as was shown in maize (93 %, Wei et al. 2009)
and rice (91 %, Chen et al. 2002). Tens of thousands of
genome-wide SNPs currently being developed in our lab
from the three mapping populations, using the state of the
art ‘genotyping by sequencing (GBS)’ approach (Davey
et al. 2011) will likely provide the degree of saturation
necessary for the task of obtaining an accurate physical
map ordering and orientation, a prerequisite for a high
quality genome sequence assembly.
In conclusion, we reported on the construction of the
first consensus genetic map of flax using 411 individuals
from three populations and grouping and ordering 770
markers in 15 LGs spanning 1,551 cM. The vast majority
of the markers are SSRs, a highly reproducible marker
system which should prove its usefulness as an important
resource for the flax research community, especially flax
breeders. The overall map density averaged one marker
every 2.0 cM. The consensus genetic map has been
anchored to the flax physical map, a first step in the
ordering of the scaffolds that currently make up the WGS
sequence assembly of the flax genome. This integrated map
will enable structural and functional genomic studies
including fine mapping of genes of interest, marker-assis-
ted flax breeding, map-based gene cloning, comparative/
synteny mapping, QTL analysis and association mapping
in flax and other related species.
Acknowledgments We sincerely thank Rae-Ann Trudeau for
technical assistance, Joanne Schiavoni for manuscript editing and
Theor Appl Genet (2012) 125:1783–1795 1791
123
Michael Shillinglaw for figure preparation. This research is part of the
Total Utilization Flax GENomics (TUFGEN) project funded by
Genome Canada and co-funded in part by the Agriculture Develop-
ment Fund of Saskatchewan, the Governments of Manitoba and
Saskatchewan, the Flax Council of Canada and the Manitoba Flax
Growers Association. Project management and support by Genome
Prairie are also acknowledged.
Open Access This article is distributed under the terms of the
Creative Commons Attribution License which permits any use, dis-
tribution, and reproduction in any medium, provided the original
author(s) and the source are credited.
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