Genetic mapping and identification of QTL for earliness in the globe artichoke/cultivated cardoon

16 Pages·2012·1.7 MB·English

by  Ezio Portis

Genetic mapping and identification of QTL for
earliness in the globe artichoke/cultivated
cardoon complex
Portis et al.
Portisetal.BMCResearchNotes2012,5:252
http://www.biomedcentral.com/1756-0500/5/252(23May2012)
Portisetal.BMCResearchNotes2012,5:252
http://www.biomedcentral.com/1756-0500/5/252
RESEARCH ARTICLE Open Access
Genetic mapping and identification of QTL for
earliness in the globe artichoke/cultivated
cardoon complex
Ezio Portis1, Davide Scaglione1, Alberto Acquadro1, Giovanni Mauromicale2, Rosario Mauro2, Steven J Knapp3,4
and Sergio Lanteri1*
Abstract
Background: The Asteraceae species Cynaracardunculus(2n=2x=34) includesthetwo fully cross-compatible
domesticated taxa globe artichoke (var. scolymusL.) and cultivated cardoon (var. altilis DC). As both are out-
pollinators and suffer from marked inbreeding depression, linkage analysis has focussed ontheuse of a two way
pseudo-test cross approach.
Results: A set of 172microsatellite (SSR) lociderived from expressed sequence tag DNA sequence were integrated
intothe reference C. cardunculus geneticmaps,based on segregation among theF progeny ofa cross between a
1
globe artichoke and a cultivated cardoon. The resulting maps each detected 17 major linkage groups,
corresponding to thespecies’haploid chromosome number. A consensus map based on 66 co-dominant shared
loci(64 SSRs and two SNPs) assembled 694 loci, with a meaninter-markerspacing of 2.5cM. When the maps were
used to elucidatethe pattern of inheritance of head production earliness, a key commercial trait, seven regions
were shown to harbour relevant quantitative trait loci (QTL). Together, these QTL accounted for up to 74% ofthe
overall phenotypic variance.
Conclusion: The newly developed consensus as well as the parental geneticmapscan accelerate the process of
tagging and eventually isolating the genes underlying earliness in both the domesticated C. cardunculus forms. The
largest single effect mappedto thesame linkage group in each parental maps, and explained about one half ofthe
phenotypic variance, thus representing a goodcandidate for markerassisted selection.
Keywords: Cynara cardunculus, Linkage map, Microsatellite, QTL, Earliness
Background maintained by its cross-pollinating habit [1]. The domesti-
The Asteraceae (ex Compositae) species Cynara cardun- cated forms produce a variety of nutraceuticals and
culus L. comprises three taxa, namely the two domesti- pharmaceutically active compounds like inulin, mono- and
cated form globe artichoke (var. scolymus) and cultivated di-caffeoylquinic acids [2–6] and sesquiterpene lac-
cardoon (var. altilis), along with their common ancestor tones, which are responsible for its characteristic bit-
the wild cardoon (var. sylvestris). While the globe arti- terness [7–9]. Globe artichoke contributes significantly to
choke was selected for its large immature inflorescences, the Mediterranean agricultural economy in the form of an
the cardoon was selected for its fleshy leaves and stalks. annualproductionof~750MtworthoverUS$500Mannu-
Thethreetaxaremainfullycross-compatiblewithonean- ally. It is also cultivated in the Americas, North Africa and
other, and their F hybrids are fertile. The species complex China(http://faostat.fao.org).
1
has a highly heterozygous diploid genome (2n=2x=34), Most of the Mediterranean globe artichoke germplasm
is vegetatively propagated, and a number of varietal
*Correspondence:[email protected] groups have been defined on the basis of the appearance
1Di.Va.P.R.A.PlantGeneticsandBreeding,UniversityofTorino,viaL.daVinci of the inflorescence and harvesting time of the head
44,I-10095,Grugliasco,Torino,Italy
(capitula) Flowering can be induced between autumn
Fulllistofauthorinformationisavailableattheendofthearticle
and spring in early flowering types by watering dormant
©2012Portisetal.;licenseeBioMedCentralLtd.ThisisanOpenAccessarticledistributedunderthetermsoftheCreative
CommonsAttributionLicense(http://creativecommons.org/licenses/by/2.0),whichpermitsunrestricteduse,distribution,and
reproductioninanymedium,providedtheoriginalworkisproperlycited.
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underground shoots, whereas late flowering types flower cultivated cardoon maps of a large number of these
only during spring and early summer. Acommon breed- EST-SSR loci, and show that they can be used as bridg-
ing target for both vegetatively and seed-propagated var- ing markers to merge the two maps. The resulting dense
ieties is the promotion of earliness since inflorescences maps was then used to identify a number of quantitative
produced intheearly part ofthe yearcommandahigher trait loci (QTL) underlying early head production in C.
price than those produced in the summer. Unlike globe cardunculus.
artichoke, the cultivated cardoon is exclusively seed-
propagated, and is generally handled as an annual crop. Results and discussion
Oflateithasbeenpromotedasasourceoflignocellulosic Genotyping
biomass [10–12] and the evidence suggests that it Six of the 178 informative Cynara Expressed Microsatel-
should be possible to derive types able to flower lite (CyEM) markers, identified by Scaglione et al. [22],
early, to produce stems with a high lignin content wereexcludedfrom theanalysisonthebasisofexcessive
and to generate biomass with a good level of energy missing values. Of the remaining 172, 54 segregated in
efficiency [13,14]. Earliness is therefore an important both parents(46as1:1:1:1,eightas1:2:1)and118injust
trait in both domesticated forms. one of the parents (85 in globe artichoke ‘Romanesco
The first generation of C. cardunculus marker-based C3’, 33 in cultivated cardoon ‘Altilis 41’). On the whole
genetic maps [15–17] have resulted in a cultivated car- 228 microsatellite markers were available for map con-
doonmapcomposedofnearly200loci(17majorlinkage struction (Table 1). Eleven of these loci suffered from
groups, LGs) spanning just over 10 M, and a globe arti- mild segregation distortion (χα2=0.05<χ2≤χα2=0.01) but just
choke one featuring 326 loci (20 major LGs) spanning one (CyEM_58) from severe distortion (χ2>χα2=0.01).
about 15 M. The two maps have since been integrated Since CyEM_58 was excluded from the mapping ana-
on the basis of common loci with the inclusion of a lysis, this left a total of 227 SSR loci. Co-dominant mar-
number ofgenesinvolvedinthesynthesisofcaffeoylqui- kers appear to be less affected by segregation distortion
nic acids [18,19]. More recently crosses between globe than dominant ones [23,24], and this certainly was
artichoke and its ancestor wild cardoon have generated the case for C. cardunculus, where ~13% of AFLP
highly segregating F populations exploitable as orna- and S-SAP loci [17], but only ~5% of SSRs and SNPs
1
mentals [20]aswell asformappingstudies[21]. are distorted. Segregation distortion has been associated
The multi-allelism of many microsatellite (SSR) loci with statistical bias and/or with errors in genotyping, but
makes them particularly well suited as bridging markers they can also stem from a number of biological phenom-
to link independent maps. The design of SSR assays ena affectingmeiosis, fertilization and embryogenesis [25]
requires DNA sequence, which in globe artichoke exists as well as the presence of null alleles. Null alleles at SSR
at present largely in the form of expressed sequence tag loci are not uncommon, as they can arise where either
(EST) sequence (http://compgenomics.ucdavis.edu/). one (or both) of the primers fail to anneal because of se-
Over 4,000 potential EST-SSR loci have been identified quence mismatch or the deletion of the whole locus, and
from this sequence resource, and the experimental test- causeanhigherapparentnumberofhomozygotesbecause
ing of a sample of 300 loci showed that more than one they can no longer be distinguished from the heterozy-
half were informative between the parents of our two gotes[26].Inthissituation,theoptionsareeithertodisre-
mapping populations [22]. In the present report, we de- gard the affected loci, to score segregation in the same
scribe the integration into the globe artichoke and wayasforadominantmarker[27],toattempttoredesign
Table1PolymorphismandsegregationpatternsfortheSSRlociusedformapconstruction
SR Number Markertype Segregationtype<RomanescoC3xAltilis41> References
prefix ofloci <abxaa><abxcc> <aaxab><aaxbc> <abxab> <abxac><abxcd>
CDAT 1 Genic 1 - - - Acquadroetal.[63]
CLIB 3 Inter-genic 2 - - 1 Acquadroetal.[63]
CMAL 5 Inter-genic 4 1 - - Acquadroetal.[64]
CMAFLP 2 Inter-genic 1 - - 1 Acquadroetal.[65]
CsPal 1 Genic 1 - - - Sonnanteetal.[66]
CsLib 1 Inter-genic 1 - - - Sonnanteetal.[66]
CELMS 43 Genicandinter-genic1 19 4 1 19 Acquadroetal.[16]
CyEM 172 Genic 85 33 8 46 Scaglioneetal.[22]
Total 228 114 38 9 67
127outofthe61CELMSmicrosatellitesarereportedtobegenicSSRastheycontainatleastoneORF.
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the primers [28,29], or to adjust allele frequencies on the before [17] and is thought to be a consequence of the
basis of a global estimate of the frequency of null alleles. sustained vegetative propagation used in globe arti-
As recently described byLanteriet al. [20] the null alleles choke, in contrast to the seed propagation applied to
segregating in a Mendelian fashion were identified, thus the cultivated cardoon, which led to a certain degree
limiting the segregation distortion in our populations. As of purifying selection aimed at stabilizing production.
noted previously [15,17], although the inclusion of loci
distorted at the 1% level and above increases the fre- Globeartichokemap
quency of type I errors, it does help to maintain marker The globe artichoke ‘Romanesco C3’ map (LOD 6.0)
densitythroughoutthemap. consisted of 473loci fallinginto 20 LGs,each containing
The updated ‘Romanesco C3’ map was built from 574 at least eight loci (Figure 1). The number of mapped
loci (359 AFLPs, 19 S-SAPs, 189 SSRs and seven SNPs), SSRs has now risen from 46 to 185. The largest LG con-
and the ‘Altilis 41’ one from 373 loci (246 AFLPs, tained 73 loci, and the range in genetic length of the in-
8 S-SAPs, 114 SSRs and five SNPs); of these, 78 (76 dividual LGs was 34.5-140.9 cM. CyEM loci (139
SSRs and two SNPs) were in common between the markers) were mapped to all the major LGs, and their
two parental genotypes. The CyEM SSRs were less in- inclusion allowed the integration of six AFLP loci which
formative in the cultivated cardoon than in the globe previouslyhadremainedunlinked[17].TwoLGs(C3_13
artichoke. Of the 228 assayed SSR loci 189 (83%) seg- and C3_18) which were previously separated have now
regated in ‘Romanesco C3’, but only 114 (50%) in been merged, while LG C3_4 has been split into C3_4a
‘Altilis 41’ (Table 1). The difference in level of hetero- and C3_4b as a consequence of more stringent LOD ap-
zygosity between these parents has been remarked on plied (Figure 1). LG C3_17 has increased in genetic
C3_10 C3_14 C3_1 C3_3 C3_2 C3_13+18 C3_7 C3_6 C3_4a C3_17
1256734577241244556781342344613456712235555667887688413744910806645931745971693541192034802100,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,166807473678453881745500565450497424790508034000 LOD 5CepCeeCrrererCepeeprereepeeeeppeeeprrerrecrpeeeeCCCCCCCCCCCCy33333341343434133334133333333131yyyyrEEELyyyyyyy96835823925565375585398835556353EEEE3eEEEEEEEI////////////////////////////////LLLBMMM5MtttttttttttmmmmmmmmmmmmmmmmmmmmmMMMMMMM88888888888MMM_/-____t5454554445566446455660290002201918_______SSS1331800700777991277179022-----------259261226943325312261---3267---------------------443-7033561352121651131541311328848037148665218397612480418991485936541000040620254008400005600020440805** 2233524211124553322234444455667922234444455667923444568234445680394789075891500990099668800882585035685807202585035685807200024746100247461,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,0091909193395453942281280244779469472674337899469472674337890834557608345576LOD 5eCeeCeeeereeeerrCCrCpCeppepeeeeecpprrCCrCpCeppepeeeeecppreereeeerreereeeerCCCCCCCCCCCCCC33333333yy33333333311113333411111333341333333433433yEyyEEyyEErryEEEEyyyyyy22yy55583247825525523225864522322586452663838245245EEEEEeeLLLLLEEEEEEEEE3/////////////////////////////////////////////////LLLLMMMMM55ttttttttMttttttttttMMMMmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmmMMMMMMM88MM8888888888888888MMMM//_____SttSSSS55401991291_56464455465646445546554554551191190202_88________SSSS52222007--------9072879071907287907199008008-17711111122123154211233----------8-------89279272242475074933321321161623030----66----------------------------8513513002121--090494494803838060026111121522111112152213323123172723319194000820822020800078098480007809840094894822005042088000504208806680680600**** 11111111111111111111111001200013000000011112241447911233378999956788889111122444555666778993938247400123567016914033638575138651379242234994899180180581350484668340M,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,1585902651273118496563933770825386879821390741973639692353323833769576890inopCerCCrrrpreeerpppreCpCCppeerCeeCrerpCCrrrCeeeeeeeecereeArprepeeCCpeeeeeCerCCCCCCCCCCCCCCCC1y33333333333133111431433133333333333333141cyEyyyyEyEEEyyE 2r-EEDyyyyyyyyyyyy22228252389285552285389232593575346653352yEEEEEEEEePLLLLLLgEEEEEEEEEEEE/l/////////////////////////////////////////LLAmMMMMMMtMM5MtttMttMMMtttttttttttttMttttmmmmmmmmmmmmmmmmmmmArMMMMMMMMMMMM8888888888888888888888MMT/a4______r__LSSSSSSe55555455654655455562021229290102101900019n_-___________SS716212122030000070009920079992----------------------0------os5125111829112826222815131963411116--015250-055923533-------------------1f40475644704216333322343121412211422221379675066834397492568629906444_-80u9717643603545312264304716126646002640446080088004850040600422406602008-sp0***np 1111112122225943445556667882345677880300139553497631013938803751850579126754134050,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,8027152439689490820545488663090164533030C3eeeCCCCerCeperCCrepprpCprpeeeeeerreeeerp_HCCCCCCC3333313311111333333333313EyyEEyy865CEyyyyyyEE882583233398325378662HEEL2LLEEEEEE////////////////////////LTMMtMmm-MMMMttttttttmmmmmmmmmmmmm8sMMMMMM88888888Ms0__S559__nSS654666466554622009010n______S2109-21p-2070017010981----------p222122232652242244-01--69-------------942388026919322421181216210658725841009751249906048823976333094002040242020052564000 11110023112331122333444444458961222223333344444444555566947013831905151702800234454245612458446690233677818893790928,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,05000543013320783323601769993655786270279116338270525580186CLOD 53crCeepeepeereeCreApCpeeerCCeCpCrepeCpCeerrrpeeepeeerprrCeeCC_CCCCCCCCCCy33133333333333313341331114441333133cy4ELEyyyyyyry2535EEyyyyyyyy8322283822838552852235222846374EyEEEEEEHIeLL1EEEEEEEEEB////l///////////////////////////////LLM5ttMMMMMMmmmttttttttttMM-mmmmmmmmmmmmmmmmmmmmmrM-8MMMMMMMM8888888888MMs/a90_e______5442SS5545665564655565446640901200021n_n________SS223171221-079097000002709000098119----------p--1s2522892722314142393--4175279588------------------------f410253963264092332632211115111382511396452928273-_8002353174112362402589932091197102271228000040020004*60000500660606682208-*s***np 1101112332341122233344555678891122333678889248459080003147500794690258948977295265914788457774037066,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,6047184053897306725220587526290251973728485456898787 C3errprCrrrrCrpeeeeCCeCerrrrrpeeppepepeeeerCeCrrCrrCCCeCCCCCCCCCCCCCCCCCC_343333344334333333341143EyyyyyyyyyyELyyyyyyYyyyyyyyMy758232228222532568553258EEEEEEEEEELEEEEEEEEEEEEEE1I///////E/L////////////////BMAMMMMMMMMMtttttMtttttttmmmmmmmmmmmmMMMMMMMMMMMMMM88888M8888888M-L__________1S5456554566552291200010900______________S-_1111112741070009790209-------------4237211111252128215424123114-025558304337------------127539218693848331791112131883053767241426*59563803363191058*434687180297820080002840605*42260008** 10356478814126791122222333444135470353361699361321413469468238800027,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,240222524520613021122944149090372038LOD 5C3eCeeeprrCeeeeprCprcerereeeeppeeCprpeCCCCCCC33333133311y_3333333443431yyyyyyM25592568522rEyyyEEE8555535255535EeEEEEE///////////1/////////////LAMMMttmmmmmMmmmm5tttttttmmmmmmMM88MMM8888888MF/___95465426564_t65665569010119__8L___S851-97207-20089911000-------2432P71211619121051----------3------17315216342110-7411122216688579-22*14232372532676261304524608488314000020580284080** 12234679134112233344455530941262004238132274570180,,,,,,,,,,,,,,,,,,,,,,,,,,91729124007549572901403810C3eepeereppCpCerCrerCCrCprrpC33343311C_CCCCC13341yyyyyyy28555832Myyyyy36652EEEEEEE////////1EEEEE/////ttmmmmmmMMMMMMAMm88mmmmMMMMM46545600L5______5_4456_____--900792-42222108792122111241------8481315-----1641118415842211212061778942166220257454682044600840 111223334441221426725123364567039581580791270068491949091261217202,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,580894374753460043139508402310266305CLOD 53eepeppprCreeeerrreerCCpCpeeeCCperCereC_CCCCCCCC3333341113333133331133yEyyyyysyyyy2333553328857Eyyy298233338EEEEE9EEEE-L3/////////////EEE/////////LMttttmmmmmmmmmMMMMMMttttttmmmMMMM8888MMM888888I_B_4644555452909_____S665200100_______2----707709980-12361220-------23171221121592421134---------600440---4062633524552422142243141505274080403224525127042493b643460240b882000422480*** 74513445566781223345555668512345112222333344555706403280548734397932369160057042340375802682578170586,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,02415192178549990365383983586051582879298798099223311 CLOD 53rCrrerrerrCrrerrpeeereeCrHepeeeeeerprereeeeeCeeeeceppCCCCCCCCCCCCCCCC_33333331431y33333333334313333311EQyyyEEEryyyyyyyyyM58322533263EEy95525885952627459522EEELeTEEEEEEEEE1E///////////LLL////////////////////LLAMMtttttttMM5tttmmmmtttttttmmmmmmmmmm-MMMMMMMMM8888888MMMM8888888888MMs/L__2S_90001905446m54644455640912122901n_________SSS-_SS232-------07810829990017-----------2p194122211413421525---16542662261--251--------------4332453703973204919867921637016221363529462796208758*5644*1381276-4064020282446447729038000262460*3*92800*20588002035*2*
p13/m50-520 34,5 p45/m47-162 46,7 e35/m49-164 52,5 e39/t80-90 55,2 e37/m61-262 63,2 e33/t80-322 7891,,44 rCCyyEEMM__27301** 82,3 p12/m47-305
Figure1Geneticmapofglobeartichoke‘RomanescoC3’.MarkernamesareshowntotherightofeachLG,withmapdistances(incM)to
theleft.OneLGcontaining<4lociislabelledas‘minorgroup’.Locishowingsignificantlevelsofsegregationdistortionareidentifiedbyasterisks
(0.1>*P≥0.05;0.05>**P≥0.01).SegmentsshadedinredindicatewhereapairofLGshasmergedasaresultofreducingthestringencyto
LOD5.
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length by 36 cM (86%), while that of LG C3_3 and C3_8 confirming the desirability of including co-dominant
has increased by ~30% and ~20%, respectively. The map markerstoobtainreliable marker placement.
spanned 1543.8 cM, with a mean inter-marker distance
of 3.40 cM, corresponding to a 3.8% increase in length Cultivatedcardoonmap
over the earlier map [17], but in a ~28% decrease in the The genetic map of the cultivated cardoon ‘Altilis 41’
mean inter-marker distance. The proportion of intervals parent (LOD 6.0) was constructed from 373 segregating
shorter than 7 cM is now 88% (previously 77%), and loci (82 CyEM loci), of which 273 were ordered into 21
only six gaps of >15 cM remain. The SSRs appeared to major LGs, whose length ranged from 27.1 to 125.2 cM,
be rather quite uniformly dispersed, although some clus- with the largest LG consisting of 29 loci. The result of
tering is present in the distal regions of C3_8, C3_2 and integrating the CyEM loci was an increase in the num-
C3_17, and around the putative centromeric region, of ber of major LGs from 17 to 21. This involved the rec-
C3_3, C3_15 and C3_20. These chromosomal regions ognition of four new LGs (Alt_18 to _21), the splitting
are typically enriched for SSRs [30–35]. The relatively of Alt_1 into two (Alt_1a and Alt_1b) and the merging
low marker saturation present in the distal regions of of Alt_16 and Alt_1b (Figure 3). The updated ‘Altilis 41’
C3_3 and C3_14 presumably reflects a localized reduced map included 107 SSR loci distributed across all but one
levelofpolymorphismbetweenthemappingparents. (Alt_13) of the major LGs, with a total genetic length of
Some segregation distortion was present at five of the 1485.7 cM and a mean inter-marker distance of
CyEM loci (CyEM_19, _47, _70, _73 and _231; three at 5.44 cM. This represents a marked increase in both
α=0.05 and two at α=0.01, Figure 1) which affected a length (+42%) and number of loci (+50%), together with
cluster of loci on both C3_17 and C3_9. In both cases a minor decrease in the mean inter-marker distance
thedistortionwasduetoanexcessofthebanddetectedin (−5%). The proportion of intervals smaller than 10 cM
thefemaleparent,thusitislikelytohaveabiologicalbasis, (about 80%) was not significantly reduced. Some of the
rather than being due to either scoring error or chance LGs recorded large increases in their genetic length –
[36].Biologicalmechanismscausingsegregationdistortions for example, that of Alt_17 by 64.7 cM, Alt_14 by
have been extensively studied in Drosophila [37], and are 58.9 cM and Alt_18 by 57.8 cM. The only LG which
known to occur in many plant species [38–42]. On the recorded a reduction in length was Alt_5. There was
otherhand,theother18distortedlociwerescatteredacross some clustering of CyEM loci in the distal region of
thegenome,acommonfeatureinthegeneticmapsofboth Alt_2 and around the putative centromeric region of
plantandanimalspecies[43]. Alt_1b and Alt_15. Three CyEM loci (CyEM_73, _3 and
By lowering the LOD threshold from 6.0 to 5.0, three _231; Figure 3) showed a degree of segregation distor-
pairsofLGsweremerged: C3_10withC3_5,C3_14with tion (two at α=0.05 and one at α=0.01), but none of
C3_8 and C3_4a with C3_4b, resulting in the formation these were linked to other distorted loci, similar to the
of 17 LGs (corresponding to the haploid chromosome other nine markers showing segregation distortion. The
number, Figure 1). It also allowed the inclusion of two additionofthenewSSR markersdecreasedthe mean in-
unlinked pairs of loci (one into C3_2 and the other into ter-marker distance on Alt_11 by ~60%, and some gaps
C3_12) and the singlet AFLP locus e35/m46-156 (into in the previous map have been filled; but ten gaps of
C3_7). This generated an increase in the genetic length >15 cM remained, perhaps reflecting regions of genetic
of the map of ~60 cM; one doublet still remains fixation which have arisen during cultivated cardoon
unlinked(Figure 1). domestication.
Both the goodness-of-fit of marker placement (mean Lowering the LOD threshold to 5.0 led to the merging
χ2 contribution) and nearest neighbour fit (cM) were of four pairs of LG: Alt_1a with Alt_1b, Alt_11 with
evaluated. Compared to the earlier ‘Romanesco C3’ map Alt_2, Alt_7 with Alt_10, and Alt_19 with Alt_13. At
[17], the average mean χ2 contribution of markers across this level of stringency the number of LGs corresponded
the LGs has been significantly reduced from 5.38 to 4.42 to the haploid chromosome (Figure 3). The lowered
(t test at α=0.005), highlighting the improvement in ro- stringency also allowed the incorporation of two groups
bustness. The variation in this parameter for each LG is of three linked loci into LGs Alt_20, and Alt_5, and of
illustrated in Figure 2, which confirms that LGs C3_1, onedoubletintoLGAlt_6.Asaresult,theoveralllength
C3_2, C3_5, C3_8, C3_10, C3_17 and C3_20 have all of the map was increased by 133.5 cM; one triplet and
shown an improved goodness-of-fit. C3_12 remained fourdoubletsstill remainunlinked(Figure 3).
largely composed of AFLP loci (only two CyEM loci
were integrated) and thus its robustness was hardly TheC.Cardunculusconsensusmap
improved. The mean nearest neighbour fit of the CyEM The number of informative shared co-dominant markers
loci (24.3±3.7 cM) was markedly lower (t test at was raised from 19 to 66 (64 SSRs, two SNPs), repre-
α=0.005) than that of the AFLP loci (51.0±4.6 cM), senting from one to 15 bridging markers per LG. As a
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Figure2Variationinthemeangoodness-of-fitofmarkersforeach‘RomanescoC3’LG.Variationdetectedbycomparingthecurrent
‘RomanescoC3’mapwiththatpublishedbyPortisetal.[17].LGsC3_13andC3_18havebeenmerged.
result,19ofthe‘RomanescoC3’LGswerealignablewith shared markers with both Alt_11 and Alt_2 (Table 2).
20 of the ‘Altilis 41’ ones (Table 2). There was a one to C3_4b remained non-aligned, but did harbour a number
one correspondence between 18 LG pairs, but C3_1 of SSR loci which were informative for the second step
Alt_1a Alt_11 Alt_7 Alt_3 Alt_19 Alt_20 Alt_5 Alt_14 Alt_9 Alt_4
0,0 e34/m49-380* 0,0 p13/m62-138 0,0 e32/t80-76 0,0 e32/t81-200 0,0 e35/m62-280 0,0 p45/m60-420 0,0 aCyEM_197 0,0 e38/m47-248 0,0 CyEM_225 0,0 CyEM_128
1442462364684767543108907,,,,,,,,,,,,,,29850024958107 LOD 5epeeeCpaCpeeceAy313331C1333yyr2338532972EEye//////////lE5MMttttmmmmmm8888tM/__t46565600021913_720201----02321327------b-326112203166133938002024+5806645**16 3321111452129216243509440968,,,,,,,,,,,,,,,,,71898777352110906 LOD 5CCaeeeaCACCpeaaCaAC333C43CCCcEyyyyy44558yEEEEEyyyyyLl/////EEEEEtlMMMMMtMmmmmr8MMMMMta_____S05565n_11262_____-0010-s30550921131----f511612509663402_89966434682-snp 22345411331111119309984473600014014,,,,,,,,,,,,,,,,,,,2982895175100899899 LOD 5peCCeeCCeappeCCeaeeCC33333314433AEyyyyy25255225557EEEEEyML///////////EMMMMMlMttttmmmmmmmA8888Mt_____S66422F125464__222252277--209L---511133233-----P--3822154121660022-185844800448852060788 13344577781682589293557625910962,,,,,,,,,,,,,,,,,48164322073705933 aeeapCpeeeeCeCpeC33333CC443334EEyy29822555775EEyyLL///////////EEMMttMMtttmmmmmm88888MM__SS10211564666__21908110-------11333015269------62025426221109409846820552407280802* 5261134406250,,,,,,,3511620 LOD 5eCeeeep333A331y822462E//////Mltttmmm888t_541621_9072---1783---424682179050053 12345790092553,,,,,,,2240711 LOD 5eeaCeep33C331y22663Ey/////EMttmmm88M_224562_--8921199---311563024984600 61312244844564303790356,,,,,,,,,,,,139565892721 LOD 5paeeeeCpCepe133331313Cyy259992337EEE/////////MMLttttmmmmm8888M__00096555481S----00009272510------082114191120401160648004*a 66458122233577351176467588079,,,,,,,,,,,,,,,717870097602095 CCCepeCCCeeeeCC3333313yyyyyyyy8552525EEEEEEEE///////MMMMMMMMttttmmm8888________210266411411127----228485230355843---0642115*56593*608410440 4723688601878,,,,,,,1191631 aCeeepC3C331Ey8593EyL////EMtMmmm8M_S6256_32-00-1125---32287294327000 4331463333112233568234678935910707917384656062630456,,,,,,,,,,,,,,,,,,,,,,,,,,,154020670482901063203298729 CCeepapeCepeCCeeaCeaeAeeeCe331C1331333C3C33333cLEEyyyy2333253438825825yEEEEIEyyLLBl////////////////EEtMMMMLttttttttMMttmmmmmmr-8888888888MMMa0____SS2914559252561002n21722__S---790--9-02------s245461329941975812-------f346099323121430022300228_610237890084085242502020-snp
12,0 CyEM_36 96,6 e37/m49-336 96,1 CyEM_183
424572266133960465879957,,,,,,,,,,,,379808160125 aaCaeaepeeeeCCC3C313333y2322828EEyyy///////EEEMLtttttmm88888MMMM_55102121___S00-----226257391---2650444558919080809850 74372346675649465922991187742056,,,,,,,,,,,,,,,,9912327517736436 pCCeepCCeCCCpeCe13313133EEEEEyyy25524238EEELLLLL////////MMMttMMMMMmmmmmm88___SSSSS90466454221--722907-----1210552352------475112116982644442965510048025*** M554464463323334452333445776926928852220265222026i,,,,,,,,,,,,,,,,,,,,,,,,223350298646075029864607nor CCpepeepeepCCpepeepeepppg33333343431443431444EEEE25525558553558553555rLLLL////////////////////MMtttMMtttttmmmmmmmmmmmmo88888888SSSS45654456546600099099u707707909000------------112322323030------------24212421216212162111p7272508735087362262288220284028422s 111021053,,,725 Ape13c35yl//tmmra54n99s--f42_50306-snp 2494,,20 Alept31_32//1tm86922-1-38900** 110,3 Alet3_5/1m457-686 1023113201881340,,,,,,,,68180510 AleteCCCCae3C33_yyyy847EEEEy///E8MMMMmmmM____4544222_70988131---32261755373422 10123097009,,,,,,437802 AlCCeCaCtC3y_yyyE4EEEy/EM1MMMmM____471_631971440-184088 2343910,,,,6300Alteapa_CC3485yy1//EEmmMM846__7211--431947600 1223333122223443340701286246930606081,,,,,,,,,,,,,,,,,,,,85901168037364172005Alt_eepCceeeeeaeeeeaeeae6y331333333333C33C3Cyr692446585325358EEyye///////////////EEM5Lttttttmmmmmmmmm888888MM/M_m0544055020965552__S-097-00----20994511144112----------72204574919122135860-7203047795448027410808428060
889939,,,438 pep131362///mmm646072---112055886** 603033030,,,,,,,,,000000000 eeeeeaeee333C3333345725576E////////mmmLttmmm88M56619454021S--70921-------231211661199799848244286** 1790,,,630 AltpCa1_Cy3EE/2MLmM_512S97--117015 2212144705,,,,,,051902 eCCppe3311Ey9523EL////MtMtmm88_S5600102---3805--9246463500 221200340,,,,,39190 pCeCC13Eyy35EEL//MMMmm__S541208-344--119482048 545456844317,,,,,,877009 eCeCep3331yy2332EE////MMtttm888__209521---051141-17930708010** 6467574026,,,,,40546 CCpCp11Eyy33EEL//MMMmm__S462172-600--410083704 67565075,,,,4772 a4paCC4C5Lyy/EE-msMMn6p__126-159560 5556884930,,,,,13252 LOD 5eaeae3C33C529Ey///ELttm88MM905_S--0312--35141262990
0,0 p45/m61-154 21,0 aCyEM_3* 76,8 p13/m62-170
3,0 p45/m61-328** 27,1 Myb12-snp 39,5 e39/m50-360 40,2 p12/m47-162 81,0 e38/m50-170 82,1 CyEM_231* 87,9 e36/m48-410 91,7 e39/m50-490
Figure3Geneticmapofcultivatedcardoon‘Altilis41’.MarkernamesareshowntotherightofeachLG,withmapdistances(incM)tothe
left.LGscontaining<4lociarelabelledas‘minorgroups’.Locishowingsignificantlevelsofsegregationdistortionareidentifiedbyasterisks
(0.1>*P≥0.05;0.05>**P≥0.01).SegmentsshadedinredindicatewhereapairofLGshasmergedasaresultofreducingthestringencyto
LOD5.
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Table2CharacteristicsandalignmentoftheconsensusC.cardunculuslinkagemap
Linkagegroupname Shared Size Total Marker Alignmentwithglobeartichokexwildcardoonmap1
markers (cM) markers density
Consensus Romanesco Altilis AlignedLGs Sharedmarkers
LG C3 41
LG_I C3_1 Alt_2+_11 15 143.6 92 1.6 LG_I 21
LG_II C3_2 Alt_4 10 144.5 72 2.0 LG_II 10
LG_III C3_3 Alt_3 4 140 50 2.9 LG_III 7
LG_IV C3_4a+_4b Alt_20 1 107.8 27 4.1 LG_VI 4
LG_V C3_5+_10 Alt_1a+_1b 4 132.5 81 1.7 LG_VandLG_X 14
LG_VI C3_6 Alt_18 1 105 19 5.8 Mola-18 3
LG_VII C3_7 Alt_5 2 106.1 28 3.9 LG_VII+LG_XVII 5
LG_VIII C3_8+_14 Alt_10+_7 6 117.8 68 1.8 LG_VIII+LG_XV+tripl. 11
LG_IX C3_9 Alt_17 7 83.5 27 3.2 LG_IX 5
LG_X C3_11 Alt_12 2 61.3 19 3.4 LG_XI 6
LG_XI C3_12 Alt_6 1 97.6 49 2.0 LG_XII 4
LG_XII C3_(13+18) Alt_14 9 123.2 54 2.3 LG_IV 13
LG_XIII C3_15 Alt_8 6 66.5 26 2.7 LG_XIV 6
LG_XIV C3_16 Alt_19 1 54.6 23 2.5 LG_XIII 2
LG_XV C3_17 Alt_9 3 92.4 36 2.6 LG_XVI 9
LG_XVI C3_19 Alt_21 1 66.7 12 6.1 LG_XIV 2
LG_XVII C3_20 Alt_15 3 44.5 11 4.5 LG_XI 3
Total 76 1687.6 694 125
Average 4.05 99.3 40.8 2.5 7.4
1AlignmentwiththemappublishedbySonnanteetal.[21]andbasedonacrossbetweenaglobeartichoke(“Mola”)withawildcardoon(“Tolfa”)genotypes.
oftheanalysis;thiswasnotthecaseforAlt_13(Table2). comparable, with some exceptions. It has been estab-
The alignment was followed by the construction of a lished that wild cardoon is more divergent from the two
consensus map based on a LOD threshold of above 5.0 cultivated forms (globe artichoke and cultivated car-
(Figure 4), which succeeded in capturing 694 loci, 227 doon) than are the two cultivated forms with respect to
(217 SSRs, ten SNPs) of which involved co-dominant one another [44,45]. Somewhat surprisingly, therefore,
markers. The map generated 17 LGs with a total genetic over 100 SSR loci featured in the consensus map but ap-
length of 1687.6 cM and a mean inter-marker spacing of parently were either non-informative or remained as
2.5 cM; consensus LG numbers (from LG I to LG XVII) singletloci inthe‘Mola’/‘Tolfa’ population.
have been assigned (Table 2, Figure 4). The length of
each individual LG varied from 44.5 to 144.5 cM (mean EST-SSRsasfunctionalmarkers
99.3 cM), with the largest containing 92 loci. Only three Putative functions can be deduced for markers derived
of the CyEM loci (the intercross locus CyEM_134 and from ESTs using homology searches with public protein
the two ‘Altilis 41’ testcross loci CyEM_167 and _79) databases. Annotation of mapped loci was performed via
remainedunlinked. BlastX search as well as InterPro scan and GO categor-
The consensus map, obtained from the domesticated isationmadeitpossible totagsome biological functions.
C. cardunculus forms, was compared with the Sonnante A set of 17 CyEM markers were annotated with GO
et al. [21] map constructed from a cross between the terms involved in the ‘response to stimulus’ (Table 3),
var. scolymus cultivar ‘Mola’ and the var. sylvestris (wild five of which were derived from transcripts related to
cardoon) accession ‘Tolfa’, by considering 125 (117 SSRs, ‘response to cold stress’ and eight to ‘response to salt
eight SNPs) common markers. The common markers stress’ terms. In particular, the marker CyEM-42, devel-
identified each of the 17 LGs on the consensus map, oped from the contig CL4773Contig1 (1281 bp, 267
with between two and 21 present on each LG (Table 2). aminoacids) [22] and mapped on LG_12 of “Romanesco
Ten of the LGs aligned readily; LGs V and VII aligned C3” map, showed high amino acid similarity (81%) with
with two ‘Mola’/‘Tolfa’ LGs, and LG VIII with two major the Arabidopsis protein kinase PBS1 (NP_196820.1, uni-
groups and a triplet of markers. LGs X/XVII, and XIII/ gene At.23518). To consider reliable orthology, a recip-
XVI each aligned with only a single ‘Mola’/‘Tolfa’ LG. In rocal tblastx analysis against the whole EST collection,
general, marker order and genetic separation were currently available for C. cardunculus, was performed
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I II III IV V VI VII VIII
C3-1 / Alt-2+11 C3-2 / Alt-4 C3-3 / Alt-3 C3-4 / Alt-20 C3-5+-10 / Alt-1a+-1b C3-6 / Alt-18 C3-7 / Alt-5 C3-8+-14 / Alt-7+-10
0,0 a-p13/m62-138 0,0 a-e33/t89-310 0,0 a-p45/m60-258 0,0 a-p45/m60-420 0,0 a-e38/m50-600 0,0 a-p45/m62-370 0,0 r-e38/m47-530 0,0 a-p45/m50-590
1111111111111111111111111111111222233333444444555555566666677777888889999999999000000000000011111111125670346678892378023470124790123568115678045671267801335578991123345667789012345789,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,05922473850676237142943124853465273183379364230007387391814060964687475938338005438 rrCarCCrArarrrrarrrararrrrrrarararaCrCrCarrrrararCCrrraCaCrrCrrrrCrrrrCarrCrrarrrar--------------------------------------------------------------------cCCCCCCCCCCCCCCcCCeeeeeeeeeeeepeeeeeeeepppppeppppepeEEEEEEyyyyyyyyCCCCCpeeeeeeepeppyEEEEEEEEy3333333333331333333331111431414313LLLLLLsDEEyyyyyyyyyyyy333333334111lyyyyyr3545392225822823582522233593525825tEEEEEEEEEEEEMMMMMMMM-MMMMMM584454582522LLeEEEEEArP//////////////////////////////////aMMMMMMMMMMMM////////////5MMttttttttttttttttttmmmmmmmmmmmmmmmm--------SSSSSSTMMMMMttmmmmmmmmmmA888888888888888888n1621222188/-------------------SSe5551902021021219102925455556555566Ls-----05593526555444566099551821211110001255211313000------------------0709000909022f0--9064441000970722--1470425663471596682-16128216162943122804634023----------------3134----------a4371607692422203613813497963223214312343248043-611121155141-37510084640600080263212264356464s66991509845656460060042200080n2682805040***p*** MMFFSSMMFFOOOOOOHHHHHHHHHH----0000------8989000000898989 11111111112233333344444555555556666666666666666666666777777788888999900111222093725688922578122357890111234445556667778999113449912368236934377169,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,920322126256023539338040058931351463450291068381595683806639475317823 rarrrrArraCrrrCararrraCCrrrrrrarrrCrCrrCraaaraCararararCrrraaaarararrC-----------------------------------------------------------cCCCCCCCCCCeepeepeppppeepepeepeeeeeeeeeeEE4LyyyyyyCCCCeepeeeeeeeeeepepyEEEEEE33133141411344313313333333333HILLEEyyyyyyyy3333333333333111BElyyy88356222523525824838322238822tEEEEEEEEMMMMMMMM-3325458232583832LLEEEr-Ls/////////////////////////////a0MMMMMMMM////////////////MMttttttttttmmmmmmmmmmmmmmmmmmm------SSMMMttttttttttnMmmmmmm8888888888n17122128888888888--p------SS--55554656546556564460901100022s---279858S551451029202956225822927411120090800297000000971----------f--8020390-70--------92-9662340-23-231342349101628-----5-----3-99422-6-4------122------4112139862952412328351111216213323522494009633013232145-b0028990191279312572280004000s00008827643891*568600066600600202n052200***p* MFSOOHHH-0--90099 111111112223333344455555566666667777888899999000111249101302567689014569012456755670567012390681895,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,48825653648802504628358988907388511935942894390 rraaaaCrrrarararrarrarrrCrrraCrrAararrrHarrCara-----------------------------------------cCCCCCCeepeeeeeeeeeeeepppppECEyyCCepeeeeeeeepepyEE13333331111133333333LLEyyyyyT3333333333414lyy27856863322388958233tEEEEEMMMM2577982225535L-EEr////////////////////saMMMMM/////////////Mttttttttmmmmmmmmmmmm--SSMMttttttmmmmmmmn88888888n21888888-------Sp22010090665554466666s--694664556102111222220125--------210097800101f-098119990------9860110-26522422494213313------------3-------969010625785312212684221576222441020265-40002402433838062939s054250004688206420000052n2020086p* 111222223334444555567888890021145600702460128040688127,,,,,,,,,,,,,,,,,,,,,,,,,,01615417418141377422545568 rarrrrrrrrrrrCrarrarrrrrrr-------------------------CCCCCeeppeepppeeeeeeeeyCeeE33443311133333333yyyyL33y52552333288338572EEEEMI26EB/////////////////MMMM//tttttttmmmmmmmmmm-Mtm-8888888281----46644555450290092-9422317497007709980-------38-406221252110----------1-4032445524221663435635223542512764402402888202004226*** 11111111111111112222222223333333344444444555555555566666666677777778888999900011233359114577891446678890023468924457799011234678811224788800458892359155815727402,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,928285315358174575785675190981790507958046710279458145735294542287303324686235 rrrarraarrrrraararrrrarrarrrarrrarraarararCararrrCrarraaraCrarrararraraCrararaaa----------------------------------------------------------------------------cCCCCCCCCCCCppeeeeepeeeepepeeeeeeeeepeeepeeepeyyyycCCCCCepeeeepeppeppeeeeeeeeeeeEEEEy4433333133334313333334331333133314EEELyyyyyyyy333333333333343333111111Eyyyyr5555587355335838658582453699389632EEEEEEEMMMMrI238228322552927232363545eLLLEEEEBeL//////////////////////////////////MMMMMMM////////////////////////5MMMtttttttttttmmmmmmmmmmmmmmmmmmmmmmm----MMMM5tttttttttttMmmmmmmmmmmmmm-88888888888133188888888888/1/-------tSSS0190029202064546465564645544455545----7623tS20211000299565464656665482961262293917-----------17072719927270077800970---56-----------01072920202079063735-263253124134430891662522232122-----------------------5--------------791635113112534211311112588440861873813-8404543021451211231211263291911849190009410822450243680560642008802024280853839305935420400865000804000480400568605628848055******* 111223344455666790724394901605028505,,,,,,,,,,,,,,,,,,514740263269155611 aarrrrrrra4aaaarrr----------C-------CpeeeeeppeCCCCeep433333113Ly334yyyy552258328-E865EEEEs/////////M///tttmmmmmmnMMMMmmm888p-645564000----44611126090927---78152129159---------88046141116644149266122241520464000 11112223344445556667778890373895673734993571390361626,,,,,,,,,,,,,,,,,,,,,,,,,,,543223273371288545556546631 arCararrrararaararrrarrCrrr-------------------------CCCCeeeeeepepepeyyCCeeeppeeEE333333131313yMyy3333311Ey655292252225EEEMM3939259EAL////////////MMM///////ttttmmmmmmmm--MtttMtmmmF8888818888---566152596244L-01S56509002451922-0-0-0-78P2000----751-42319--------75221----*4324211616014179418120432376275862610110404020050880*** 111112222222223333333334444445555556666666667777777778888888999901241770023456781234667790035780136680023557780112244570111467045987,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,969091813770970632073753547534054901900481292031723306089043938938 araarCaarrarrrrrCrCCarraarrarrCarCaaarraarrarrarrrAarrararrrarrrrrr------------------------------------------------------------ccCCCCCCCeeeeeeeeeeepeeppepeeeeeeeeeeppEEEyyyCCepeeepppeepeepppeeeeyEEEy333333333431331131333333333341LLLEEyyyyy33333333333341441444lMyr847555232222823282535548646552tEEEEEMMMMMM22522555525535558755eLLEr//////////////////////////////AaMMMMM5////////////////////MMtttttttttttttmmmmmmmmmmmmmmmmm---SSSMttttttttmmmmmmmmmmmm8888888888888n522F/88888888-----t---SS555546644545554426991091112109s-2302120099665454465446811121L13010009702780790879-1------------f--6--------229907700770792827P272-4315122136211211-----------------735121321-------------51803942214111231521230747353489341-190605424212211232121863-0090980000748098804060820600822s84868224504815273882740800005402086860n82260502840*p* MMHH--0089
112203,,10 ar--ee3333//tt8890--127742 135,6 r-CyEM-120
111122334603,,,,6916 rrrr----eeee33336789////ttmm88440079--51--2517600804 140,0 r-p13/m47-175
113483,,46 ar--ee3368//mt8529--515102 144,5 r-e37/m61-510
IX X XI XII XIII XIV XV XVI XVII
C3-9 / Alt-17 C3-11 / Alt-12 C3-12 / Alt-6 C3-13+18 / Alt-14 C3-15 / Alt-8 C3-16 / Alt-19 C3-17 / Alt-9 C3-19 / Alt-21 C3-20 / Alt-15
002,,,037 rar---epp311233///tmm8264-272--51444380 0,0 r-e37/m50-340 0,0 r-e36/m48-640 0,0 r-p45/m59-172 01,,08 ar--ee3367//mm4580--125302 02,,05 rr--cCyyrEeM5/-t8177-2230 0,0 r-e33/t80-280 0,0 Myb12-snp 0,0 r-p45/m47-162MMFOHHH--00-8908
112222223344555556666784801677989020157924891,,,,,,,,,,,,,,,,,,,,,,,40711835296647227651544 rCrCrrCraaCrCararrCrarC----------------CCCCeepeepeEyyyyyyCeeppEEEEEE3313313Lyyyy3311y8933238EEEEMMMMMMM2433E///////MMMM////tttttmm------SMtmmm888886131228-----0050160-4040034662271561--0--2-801922-202402411342--*---*93b5072441811150634260607704840 MMFFOOHHHH--00--008989111122233344445661238266158126931,,,,,,,,,,,,,,,,,63844889833783743 rrrrrrCarrCrraaara----------------CCCCCCCCepeEyppeeeE343LELyyyyyy33311459EEEEEEMIM23959LB///MMMMMM/////Mtmm-Stt-mmm81880-------S4405560052131710497--602--0317952492--83---33362300923645353600500**1122222233333344444555555666666666677757957034569014579046891345670112334567012,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,14072334994241284869086642470128475125085 rrrrrrrrrrrrrrrCrrrraarararaaraaaaaaaaara----------------------------------------CcCCeeeeeeeeepeeeeeeeeepyCCeeeeeeeeeeeepeEy43333333331333333341EEy33333333333331Eyr22999558725855455522EM53295344685625eLLEL////////////////////M//////////////5MMttttttttttmmmmmmmmmm-MttttttMmmmmmmmm88888888882888888/-SSm55446445569200121291-1S00200954455556110099297001------------8------0970099247-6421265626326----------1115445--------949632211315420976692813457049191222217-0927473063060220640448002304777293008586220084208852 MMFFSSOOOOHHHHHH--00----8900008989 1111112222222223333334444455555666776014667134566789013478447890335906901,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,,4477161434317831109908140640456004316 rraraarrrrrarrrCCrrCraraCrCrrrrararra--------------------------------CCCCCpeeepeeeppeeeeeeeeyyyyyCeeeepeeeEEEEE143414333333333333yMyYy33333331y228535846852222833EEEMMMMM25758852EEA//////////////////MMM////////tttttttttmmmmmmmmm-----MMttttmmmmL888888888111118888----655655544200201099--5580312602644251222209000997---------5965--2--297838121112345120---------8452----0111179311182786733465111260422476898518802020041494608**04806424468* 111122233333444444555666255913524468335678033056,,,,,,,,,,,,,,,,,,,,,,,,916901844988353510858155 CararCraCrrCraCrarraCrra------------------CCCCCCepppeyyyyyyCeeeeepEEEEEE31413yyyyMy333331y63523EEEEEMMMMMM323482EA/////MMMMM//////tmmmm------MtttmmmL8222124888------04556-8813144550291121411-70926177090---042582135151-------2570221123217193244687718008040200**11111222223334444455613699135781460126904,,,,,,,,,,,,,,,,,,,,,969903795631298099396 rrrrarrrrrrrrrrrCrrar--------------------CCCeeppeeepeeeepeeyeeE334433313333433Eyy33355525538552557EEM25L///////////////MM//Mttttttttmmmmmmm-tm8888888898--S90661016515059631612--11---09-0-0-1--2141619123138--------1666218851172129222260408443850286882804020 111222333344444455555566666740371393457026889133468024680,,,,,,,,,,,,,,,,,,,,,,,,,,,,,52921262245101348867878294685 rCraarrraarrrrrarrrCrHrrrrrrar---------------------------CCCCCCCCCCCpeeeeeeeepEQyCeeeepE1333334331LEEEyyyMyyyy33331Ty3235252683EEEEEEEMM82853LLLEsA//////////MMMMMMM/////MMMnttttttmmmm-SMttmmmL888888288p--------SSS-4009195406-222566341911121227-----08-1---5--0205957340774153524324211-------*98671*1601792693644783532**824002426424093*8200000**12233455679518593196,,,,,,,,,,,19054930137 aarraCrrrrr----------cCeepeeyCCpEy31333y1Eyr53552EM3eEL/////M/5tmmmm-MMm82/-e25444-7S5133-0799794-2*5----1-125319-117362640000445111122345901683444,,,,,,,,,,4530080755 CCCCararra------eeeE3yyeppEE333HL311568MMM-532s//////tmm--Snmmm821p-55944454390-8480792-----83211121402624842FSSOOOHHH---000998
74,0 a-CyEM-204 7779,,33 Cr-CyEyEMM-7-31*6*9 7767,,56 aC-yeE3M9/-m3550-240
79,9 a-e38/m50-350 81,5 rr--CCyyEEMM--218485 80,4 r-CyEM-20
83,5 a-cyre5/m47-90 888256,,,633 Cra--CyeEy3EM2M/-t28-4083-338 83,8 r-CyEM-232
8888,,28 aa--cey3r5e/5t8/m9-4373-6160 8868,,54 rr--Cp4y5E/Mm-6209-1200
9923,,14 aa--Ce3y2E/Mt8-02-11952 999013,,,852 CCa-yyeEE3MM5/--m1473488-500 92,4 a-e37/m49-336
97,6 a-CELMS-49 98,8 r-e32/t81-400
101,6 r-e32/t80-410
110,5 r-e32/t82-258
123,2 a-e35/m50-650
Figure4(Seelegendonnextpage.)
Portisetal.BMCResearchNotes2012,5:252 Page8of15
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(Seefigureonpreviouspage.)
Figure4ConsensusgeneticmapofC.cardunculus.MarkernamesappeartotherightofeachLG,withmapdistancesincMtotheleft;'r-'
and'a-'indicatemarkerssegregatingonlyin,respectively,‘RomanescoC3’(C3)and‘Altilis41’(Alt41).Arrowsindicatethepositionsofearliness
QTL,namedasfollows:traitabbreviation(MH:maininflorescence;FOH:firstorderinflorescence;SOH:secondorderinflorescence)andharvest
season(08:“2008”,09:“2009”).
and no better alignment than that of contig CL4773 was The mean eMH, eFOH and eSOH lay substantially above
detected. PBS1 was found to work as R gene against the themid-parentvalue,suggestingsemi-dominanceforlate-
bacterial pathogen Pseudomonas syringae, where its ness. The low global level of heterozygosity characteristic
cleavage, operated by the pathogens’ effector AvrPphB, of the cultivated cardoon makes it possible that one or
triggers the signalling cascade, generating the host re- moreoftheearlinessQTL areinthehomozygousstatein
sponse (HR) [46]. Pseudomonas spp. together with other ‘Altilis41’,sothatthepresenceofdominantallelesforlate-
endophytic bacteria may affect globe artichoke plants ness may contribute to later flowering across the whole
both in field and during micropropagation [47] and the mapping population. The inter-trait correlations were
CyEM-42 may be likely considered a reliable marker for similar in both seasons, with the strongest correlation
taggingabacterial resistance traitinthespecies. linking eMH and eFOH (r>0.80, P<0.0001). The corre-
Our EST-SSR markers may be defined as functional lationsbetweenthe two seasonswerealsostrong, ranging
markers with the potential to target polymorphisms in from 0.64 (P<0.0001) for eMH to 0.49 (p<0.001) for
gene responsible for traits of interest and they can be eSOH (Table 5). Flowering and head harvesting time was
also particularly useful for constructing comparative a little earlier in “2009” than in “2008” (7–8 days on
frameworkmaps with other Asteraceae, giving the possi- average), while performance was somewhat more
bility toamplify ortholog genesandprovideanchorloci. variable in “2009” (Table 4), probably reflecting the
difference between re-awakened and newly sown ma-
Thegeneticbasisofearliness terial. The broad sense heritability for eMH of 0.76
An evaluation of the variance for the three earliness- (Table 4) indicated the trait to be predominantly
related traits established significant genotypic differences under genetic control, but the rather lower heritabilities
(P<0.05) between ‘Romanesco C3’ and ‘Altilis 41’ shown by the traits eFOH and eSOH suggested that
(Table 4). Thus, eMH in the former was 162 days in the environment is quite influential in their
“2009” and 178 days in “2008”, while in the latter the re- determination.
spective times were 218 and 223 days. All three traits The KW test and SIM procedure identified, at first, six
varied continuously among the F1 progeny (the distribu- QTL regions stable across years in the developed con-
tion for eMH is shown in Figure 5); no progeny was as sensus map (Figure 4). Those on LGs I, XI and XVII
early flowering as ‘Romanesco C3’, but a few were later involved all three traits, those on LGs I and IX only
floweringthan‘Altilis41’, due totransgressivesegregation. eMH and eFOH, and the one on LG VII solely eMH.
Table3CyEMmarkerswithGeneOntologyannotationforstimuliresponse-relatedterms
GOID Term Level N°ofloci LocusName
GO:0050896 responsetostimulus 2 17 CyEM-008,CyEM-030,CyEM-42,CyEM-054,CyEM-057,CyEM-070,
CyEM-072,CyEM-093,CyEM-120,CyEM-135,CyEM-145,CyEM-150,
CyEM-152,CyEM-218,CyEM-229,CyEM-259,CyEM-266
GO:0009628 responsetoabioticstimulus 3 13 CyEM-008,CyEM-030,CyEM-054,CyEM-070,CyEM-093,CyEM-120,
CyEM-135,CyEM-145,CyEM-150,CyEM-152,CyEM-218,CyEM-229,CyEM-259
GO:0042221 responsetochemicalstimulus 3 4 CyEM-093,CyEM-218,CyEM-229,CyEM-266
GO:0006950 responsetostress 3 15 CyEM-008,CyEM-030,CyEM-054,CyEM-057,CyEM-070,CyEM-072,
CyEM-093,CyEM-120,CyEM-135,CyEM-145,CyEM-150,CyEM-152,
CyEM-229,CyEM-259,CyEM-266
GO:0009266 responsetotemperaturestimulus 4 5 CyEM-008,CyEM-054,CyEM-093,CyEM-145,CyEM-150
GO:0006970 responsetoosmoticstress 4 8 CyEM-030,CyEM-070,CyEM-093,CyEM-120,CyEM-135,CyEM-152,
CyEM-229,CyEM-259
GO:0010033 responsetoorganicsubstance 4 3 CyEM-093,CyEM-229,CyEM-266
GO:0009409 responsetocold 4 5 CyEM-008,CyEM-054,CyEM-093,CyEM-145,CyEM-150
GO:0009651 responsetosaltstress 5 8 CyEM-030,CyEM-070,CyEM-093,CyEM-120,CyEM-135,CyEM-152,
CyEM-229,CyEM-259
RedundancyoccurssinceeachavailableGO-levelofannotationwasconsidered.
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Table4Earlinessoftheparentallines(‘C3’:‘RomanescoC3’,‘Alt41’:‘Altilis41’)andtheirF1progenyin“2008”and
“2009”
Precocitytrait Year Parents1 F population2
1
C3 A41 Mean Range s.e. h2
B
Mainhead(eMH) 2008 178a 223b 212.9 202-238 0.518 0.76
2009 162a 218b 206.1 190-235 0.694
Firstorderheads(eFOH) 2008 185a 229b 221.3 209-248 0.456 0.61
2009 180a 224b 214.0 198-239 0.828
Secondorderheads(eSOH) 2008 198a 239b 232.6 214-267 0.695 0.54
2009 192a 237b 225.7 207-246 0.897
1MeansfollowedbydifferentletterswithinthesamerowaresignificantlydifferentatP<0.05.
2s.e.:standarderrors;hB2:broadsenseheritabilitybasedontwoyears’data.
The seventh QTL cluster on LG II involved all three ‘Romanesco C3’ and ‘Altilis 41’ maps were used separ-
traits, but was only expressed in “2009” (Figure 4). On ately for QTL validation, the percentage of phenotypic
the whole, seven chromosomalregionsscattered over six variance explained by some of the QTL differed from
LGs of the consensus map were identified. When the that predicted by the analysis based on the consensus
50
Mainhead 2008
(eMH-08)
40
s
e
p
y
ot
n 30
e
g
1
F
of 20 A41
s
r
e
b
m
Nu 10 C3
0
8 2 5 8 1 4 7 0 3 6 9 2 5 8
7 0 0 0 1 1 1 2 2 2 2 3 3 3
1 2 2 2 2 2 2 2 2 2 2 2 2 2
40
Mainhead 2009
(eMH-09)
s
e
p 30
y
ot
n
e
g
1
F 20
of A41
s
r
e
b
m
u 10 C3
N
0
2 0 3 6 9 2 5 8 1 4 7 0 3 6 9 2 5
6 9 9 9 9 0 0 0 1 1 1 2 2 2 2 3 3
1 1 1 1 1 2 2 2 2 2 2 2 2 2 2 2 2
Earliness of head production (days)
Figure5Distributionoftheharvestingtimeofthemainheadamong154F individualsin“2008”and“2009”.Parentalmeans(‘C3’:
1
‘RomanescoC3’,A41:‘Altilis41’)indicatedbyarrows.