Botanical Journal of the Linnean Society

, 2004,

146

, 385–398. With 1 figure

© 2004 The Linnean Society of London,

Botanical Journal of the Linnean Society,

2004,

146

, 385–398

385

Blackwell Science, LtdOxford, UKBOJBotanical Journal of the Linnean Society0024-4074The Linnean Society of London, 2004? 20041464385398Original Article

AGE OF MONOCOT GROUPST. JANSSEN and K. BREMER

*Corresponding author. Postal address from 01/2005: Albrecht-von-Haller-Institute für Pflanzenwissenschaften, Abt. Spezielle Botanik, Untere Karspüle 2, 37073 Göttingen, Germany. E-mail: janssen@mnhn.fr

The age of major monocot groups inferred from 800

+

rbcL

sequences

THOMAS JANSSEN

1

* and KÅRE BREMER

2

1

Museum National d’Histoire Naturelle, Département de Systématique et Evolution, USM 0602 Taxinomie et collections, 16 rue Buffon, 75005 Paris, France

2

Department of Systematic Botany, Evolutionary Biology Centre, Norbyvägen 18D, SE-752 36 Uppsala, Sweden

Received August 2003; accepted for publication June 2004

Phylogenetic research on monocots has been extraordinarily active over the past years. With the familial interre-lationships being sufficiently understood, the question of divergence times and crown node ages of major lineagescomes into focus. In this study we present the first attempt to estimate crown and stem node ages for most ordersand families of monocots, based on

rbcL

sequence data and comprehensive taxon sampling. From our analysis it isobvious that considerable monocot diversification took place during the Early Cretaceous, with most families alreadypresent at the Cretaceous–Tertiary boundary. Araceae, Arecaceae and Orchidaceae are among the oldest familieswith crown node ages reaching back into the Early Cretaceous. We comment on possible error sources and the neces-sity for methodological improvement in molecular dating. © 2004 The Linnean Society of London,

Botanical Jour-nal of the Linnean Society

, 2004,

146

, 385–398.

ADDITIONAL KEYWORDS:

Cretaceous – dating – fossils – NPRS – phylogeny – Tertiary.

INTRODUCTION

Monocots constitute an angiosperm clade of out-standing economic and ecological importance. TheAngiosperm Phylogeny Group (APG II, 2003) recog-nizes 81 families in ten orders (with two familiesunplaced to order). Distribution of monocots is world-wide, with some families predominantly in open tem-perate habitats (e.g. grasses, Poaceae) and others withimportant species diversity in the tropics (e.g. orchids,Orchidaceae and palms, Arecaceae).

Phylogenetic research on monocots has receivedmuch interest recently, fostered by three internationalcongresses. Chase

et al

. (2000) proposed a first phylo-genetic tree, including all orders, based on a large dataset comprising three DNA regions (

rbcL

,

atpB

and 18SrDNA) but with comparatively limited taxon sam-pling. Meanwhile, a detailed treatment is available forseveral orders (Les, Cleland & Waycott, 1997; Fay

et al

., 2000; Kress

et al

., 2001; Vinnersten & Bremer,2001; Bremer, 2002; Caddick

et al

., 2002a, b). With theresults of the available studies taken together, wehave at present a rather well supported phylogenetichypothesis for the whole clade that is fairly wellresolved down to family level. This provides us with asolid basis for research on age, biogeography andevolution of this group.

Only a few studies are available to date on ageinference including monocots. Wikström, Savolainen& Chase (2001) analysed a data set of 560angiosperms (

rbcL

,

atpB

and 18S rDNA) usingnonparametric rate smoothing (Sanderson, 1997).Bremer (2000) determined the ages of major monocotlineages, calculating mean branch lengths in a phylog-eny of

rbcL

sequences. Both studies also addressedthe issue of age calibration using evidence from thefossil record. Givnish

et al

. (2000) estimated ages forseveral groups of commelinids using

ndhF

sequences.Recently, some detailed studies comparing differentdating methods have become available for Liliales(Vinnersten & Bremer, 2001) and Poales (Bremer,2002). These studies notwithstanding, the node ages

386

T. JANSSEN and K. BREMER

© 2004 The Linnean Society of London,

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2004,

146

, 385–398

of most parts of the monocot tree are still ratherimperfectly known.

Rate heterogeneities, age calibration, the underlyingphylogenetic hypothesis and taxon sampling are majorsources of error in phylogenetic dating with DNAsequences. Error sources related to sequence evolutionhave been extensively discussed (review by Sanderson,1998). It is well known that increased taxon samplingmay increase the accuracy of phylogenetic reconstruc-tion (Källersjö

et al

., 1998; Rydin & Källersjö, 2002;Zwickl & Hillis, 2002). The influence of taxon samplingon dating has not received much interest (but seeBaldwin & Sanderson, 1998) and is still insufficientlyknown, but improved taxon sampling has been shownto reduce error in branch length estimates (Sanderson,1990) and to circumvent putative biases introduced bythe overrepresentation of certain groups of herbaceousplants, which are prone to accelerated evolutionaryrates (Wikström

et al

., 2001).Herein, we present an extended study on the age of

monocots. We use all monocot

rbcL

sequences of suffi-cient quality currently available from GenBank toinfer divergence times of major lineages and crownnode ages for most monocot families. Sequences fromall families (

sensu

APG II, 2003) of monocots, with theexception of the achlorophyllous Corsiaceae and Triu-ridaceae, have been included. Within the now enlarged(APG II, 2003) families Asparagaceae, Alliaceae andXanthorrhoeaceae, all of the formerly recognizedAsparagales families (

sensu

APG, 1998) are repre-sented by at least one sequence. Dating analysisundertaken in this study is thus based on an extensivedata set comprising more than 800

rbcL

sequences.Our age estimates are based on a well supported con-sensus phylogeny derived from several detailed stud-ies. We provide the largest taxon sampling currentlyavailable under the assumption that this is as benefi-cial for dating attempts as it has been shown to be forphylogenetic reconstruction.

Nonparametric rate smoothing (NPRS; Sanderson,1997) may cope with rate heterogeneities, which aremore likely to occur in trees with an extensive anddiverse taxon sampling. Furthermore, rate changesamong adjacent parts of the tree are presumablysmaller if more taxa are sampled. This agrees with theunderlying assumption of NPRS that no abrupt ratechanges occur in the tree. For calibration, we followBremer (2000) who used eight reference fossils to esti-mate the split between

Acorus

and the other monocotsat 134 Mya.

MATERIAL AND METHODS

D

ATA

SET

We downloaded all Liliopsida (monocot)

rbcL

sequences available from GenBank in August 2002

with the aim of sampling as many genera as possible.Each genus is represented by a single sequence in thedata matrix. If multiple sequences of the same qualitywere available for one genus, the most recent was cho-sen unless there was an older version of higher qualitywith respect to sequence length and percentage ofambiguous bases. Sequences with species names havebeen preferred to sequences of ambiguously deter-mined or undetermined species. If these selection cri-teria failed, one sequence was chosen arbitrarily, thatis the ‘most common’ species or simply the first item inthe list. Following selection, sequences with long poly-N stretches (more than 150 ambiguous bases persequence) and sequences of insufficient length (lessthan 75% of the total length of the data matrix) werediscarded.

The resulting data set consists of the

rbcL

sequences of 878 genera from 77 families represent-ing all ten orders of monocots (Table 1). Familyassignments have been adopted from APG II (2003).The matrix comprises a total of 1343 includedsequence positions. Of these, 760 are parsimony-informative and 24 are single or double gap charac-ters resulting from putative errors in some sequences.The alignment was carried out by hand and involvedno indels.

rbcL

VERSUS

THREE

GENES

Our dating is based on

rbcL

sequences only. To assessthe historical information provided by this single genewe compared it with that of the data matrix compris-ing three genes (including

rbcL

) by Chase

et al

. (2000).Their tree was published with parsimony branchlengths. We also obtained parsimony branch lengthsfor a

rbcL

-gene-only tree comprising exactly the sametaxa as the three-gene-tree using PAUP (Swofford,2001), with settings as described in Chase

et al

. (2000)and their tree as a topological constraint. Dating oftheir tree was then performed with the three-gene-tree’s branch lengths as well as with the

rbcL

-only-tree’s branch lengths. For this comparative purpose,no correction was made for multiple substitutions.Dating was performed using nonparametric ratesmoothing (see below; Sanderson, 1997, 1999).

Ages estimated with

rbcL

only were 0–20 Myr (insix instances 20–24 Myr) older or 0–10 Myr (in twocases 13–14 Myr) younger than with the three-genedata set. Compared with the three-gene data set therewas no overall tendency for generally younger orgenerally older age estimates with

rbcL

only.Our data set with its single gene comprises fewer

characters per taxon compared with a multigene dataset, making it less suitable for phylogenetic recon-struction. However, considering the similarity of agesobtained with all three genes and with

rbcL

only, we

AGE OF MONOCOT GROUPS

387

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2004,

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, 385–398

Table 1.

Sampling and age estimates for families and orders of monocots. Orders are arranged according to major groupsrecognized within monocots, viz. core monocots and commelinids. Orders are in the same sequence as in Fig. 1. Numbersin parentheses after orders indicate the number of sampled families in relation to the number of families recognized inthis order according to APG II (2003). The first column gives the number of genera per family or order for which

rbcL

sequences were sampled. Crown and stem node age estimates are found in the second and third columns. Crown node ageestimates are absent for monogeneric families and for families for which data for only one genus were available

Taxon

Numberof generasampled

Crownnode age(Mya)

Stem nodeage (Mya)

Acoraceae 1 – 134(calibration)

Alismatales (14/14) 78 128 131Alismataceae 8 55 57Aponogetonaceae 1 – 98Araceae 29 117 128Butomaceae 1 – 88Cymodoceaceae 5 61 67Hydrocharitaceae 16 75 88Juncaginaceae 3 52 82Limnocharitaceae 2 44 57Posidoniaceae 1 – 67Potamogetonaceae 5 23 47Ruppiaceae 1 – 65Scheuchzeriaceae 1 – 92Tofieldiaceae 2 100 124Zosteraceae 3 17 47

CORE

MONOCOTS

674 126 131Petrosaviaceae 2 123 126Dioscoreales (3/3) 14 123 124Burmanniaceae 3 93 116Dioscoreaceae 8 80 116Nartheciaceae 3 76 123Pandanales (4/5) 12 114 124Cyclanthaceae 4 77 98Pandanaceae 2 51 98Stemonaceae 4 84 108Velloziaceae 2 14 108Liliales (9/10) 47 117 124Alstroemeriaceae 3 30 76Campynemataceae 2 73 117Colchicaceae 9 44 76Liliaceae 17 80 91Luzuriagaceae 2 56 79Melanthiaceae 10 97 109Philesiaceae 2 60 76Rhipogonaceae 1 – 76Smilacaceae 1 – 90Asparagales (14/14) 371 119 122Alliaceae 55 87 91Asparagaceae 63 89 91Asteliaceae 3 92 104Blandfordiaceae 1 – 100Boryaceae 2 54 109Doryanthaceae 1 – 107

Hypoxidaceae 6 78 100Iridaceae 57 96 103Ixioliriaceae 1 – 112Lanariaceae 1 – 113Orchidaceae 145 111 119Tecophilaeaceae 8 87 108Xanthorrhoeaceae 27 90 93Xeronemataceae 1 – 100

COMMELINIDS

353 120 122Arecaceae 120 110 120Dasypogonaceae 3 100 119Commelinales (5/5) 37 110 114Commelinaceae 27 62 89Haemodoraceae 2 81 98Hanguanaceae 1 – 104Philydraceae 2 47 110Pontederiaceae 5 39 89Zingiberales (8/8) 21 88 114Cannaceae 1 – 68Costaceae 4 47 79Heliconiaceae 1 – 88Lowiaceae 1 – 78Marantaceae 4 57 68Musaceae 3 61 87Strelitziaceae 3 59 78Zingiberaceae 4 26 79Poales (18/18) 172 113 117Anarthriaceae 3 55 96Bromeliaceae 7 96 112Centrolepidaceae 1 – 97Cyperaceae 48 76 88Ecdeiocoleaceae 2 73 89Eriocaulaceae 6 58 105Flagellariaceae 1 – 108Hydatellaceae 1 – 87Joinvilleaceae 1 – 90Juncaceae 6 74 88Mayacaceae 1 – 87Poaceae 52 83 89Rapateaceae 6 79 112Restionaceae 29 74 96Sparganiaceae 1 – 89Thurniaceae 2 33 98Typhaceae 1 – 89Xyridaceae 4 87 105

Taxon

Numberof generasampled

Crownnode age(Mya)

Stem nodeage (Mya)

388

T. JANSSEN and K. BREMER

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2004,

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, 385–398

assume that the information content is still sufficientfor age inference. Restricting the data set to a singlegene does not markedly alter the historical informa-tion in the data set, makes more taxa available, andenables us to infer crown node ages for most familiesas well as to address potential problems caused by alimited taxon sampling.

O

BTAINING

A

TREE

WITH

BRANCH

LENGTHS

FOR

DATING

Phylogenetic reconstruction is not attempted in thisstudy. A topological backbone constraint tree was con-structed, combining well supported clades from earlierstudies focused on phylogenetic reconstruction of var-ious monocot groups and based on data sets includingtwo or more genes.

First, all orders and families were constrained tobe monophyletic. No constraints were applied withinfamilies. Second, ordinal interrelationships wereadopted from APG II (2003). Third, interrelation-ships of families within orders were constrainedusing sufficiently supported nodes, that is withbootstrap or jackknife frequencies of 85% or higher,from Les

et al

. (1997; Alismatales), Fay

et al

. (2000;Asparagales), Vinnersten & Bremer (2001; Liliales),Caddick

et al

. (2002a, b; Dioscoreales, Pandanales),Kress

et al

. (2001; Zingiberales) and Bremer (2002;Poales). Branches with low support in those studieswere collapsed into polytomies in the constraint tree.For Commelinales, there is no detailed study avail-able to date. Some well supported nodes for thisorder were adopted from Chase

et al

. (2000). Theconstraint tree topology is available from the authorsupon request.

One completely resolved tree was then obtained by aheuristic PAUP search (Swofford, 2001) with topolog-ical constraints enforced as specified above, 1000 ran-dom addition sequences holding five trees at eachaddition step, using tree bisection reconnection (TBR)branch swapping, and the MulTrees option not ineffect and hence, keeping only one tree per replicate.One single most parsimonious tree, 13 533 steps long,was found and saved as an input tree for subsequentanalyses.

Parsimony branch lengths corresponding to thetotal number of observed substitutions per branchtend to underestimate the number of changes, espe-cially on long branches. Therefore, corrected branchlengths were obtained with PAUP using a maximumlikelihood optimization under the GTR

+

G

model ofsequence evolution (

a

=

0.5). With our data set, esti-mation of the shape parameter from the sequences isnot computationally feasible in a reasonable amountof time. We chose, therefore, to adopt the defaultparameter value from PAUP, arguing that the influ-

ence of the error in branch lengths thus introduced issmall compared with calibration uncertainty (Bremer,Friis & Bremer, 2004). The branch length calculation(yielding the number of expected substitutions persite) resulted in some zero-length branches, which arenot tolerated by the algorithms implemented in ther8s programme (Sanderson, 1999). Hence, the lengthsof 58 such branches in the tree were set to 0.000001substitutions per site. Considering the number ofbranches in the tree and the average number of sub-stitutions per site, this modification should not influ-ence the outcome of the analysis. The input tree withbranch lengths is available from the authors uponrequest.

D

ATING

Using eight reference fossils, Bremer (2000) esti-mated the age of the split between

Acorus

and allother monocot lineages at 134 Mya. Later, it has beenshown that one of Bremer’s reference fossils (fossil B)is not a

Pistia

of Araceae as assumed, but a different,unknown plant (Stockey, 2003). Recalculation of themean change rates used by Bremer, and recalibrationwithout that fossil gives, however, the same result. Weused this dating of the crown group age of monocots tocalibrate our tree. Further age constraints usingreference fossils were not included, but consistency ofthe inferred node ages with putative positions of allreference fossils used by Bremer (2000) has beenchecked.

The input tree with corrected branch lengths (seeabove) was subjected to nonparametric rate smoothingas implemented in the r8s programme (NPRS;Sanderson, 1997, 1999). Searches were restartedthree times with a perturbation of the initial param-eters to ensure the solution reached a stable optimum.To check for the presence of multiple optima, optimi-zations were also restarted three times from differentinitial divergence time estimates.

We also tried the mean path length method usingthe PATH program (MPL; Britton, 2002; Britton

et al

.,2002). MPL tolerates zero-length branches butrequires a strictly bifurcating tree with branchlengths in terms of whole numbers of substitutions,observed or expected (corrected). Hence, we multipliedthe maximum likelihood-derived expected branchlengths per site by the number of sites included inthe analysis to obtain (corrected) parsimony branchlengths. Polytomies within the families were arbi-trarily resolved, assigning zero-lengths to thebranches by branch-swapping using the tree itself as atopological constraint and saving only one tree. TheMPL clock tests (see Britton

et al

., 2002) reportedsignificant deviations from the clock at about one thirdof the nodes, including many nodes at the base of the

AGE OF MONOCOT GROUPS

389

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2004,

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, 385–398

tree. Hence, we proceeded with NPRS dating, whichpermits rate changes throughout the tree. Penalizedlikelihood (Sanderson, 2002) was not available as analternative, parametric dating approach because thesoftware failed to run with a data set of this size. Nec-essary computation time for Bayesian estimation ofdivergence times (Kishino, Thorne & Bruno, 2001;Thorne & Kishino, 2002) with our data is not currentlyfeasible.

No confidence intervals are provided with our NPRSanalysis because calculating confidence intervals bybootstrapping with r8s (NPRS) is not computationallyfeasible with a data matrix of this size. However, ourMPL analysis reported confidence intervals less than

±

16 Myr. Earlier studies (Vinnersten & Bremer, 2001)indicated that confidence intervals calculated by boot-strapping with r8s and by the MPL procedure are sim-ilar, so confidence intervals in this range may beenvisaged in interpreting the ages obtained by our r8sanalysis.

RESULTS

The

rbcL

matrix yielded a single most parsimonioustree that was not fully resolved but showed some poly-tomies, especially within clades that were representedby a high number of rather closely related speciessuch as Arecaceae, Orchidaceae and some families inAsparagales. At a deeper level, Dioscoreales andPandanales were in a trichotomy with the Liliales–Asparagales–commelinids clade.

Taking into account the large taxon sampling, it isnot surprising to find considerable variation in totalbranch lengths from the root to the terminals of thetree, indicating different substitution rates. Notewor-thy are comparably slow rates of molecular evolutionin Arecaceae (Wilson, Gaut & Clegg, 1990). Some taxashow particularly long-terminal branches, attribut-able to putative phylogenetic misplacement in thecase of

Xyris

,

Mayaca

,

Trithuria

and

Centrolepis

(seealso Bremer, 2002), and in

Burmannia

possiblyrelated to a specialized life history (Soltis

et al

., 2000).Within Asparagaceae (Amaryllidaceae),

Hessea

,

Geth-yllis

,

Strumaria

and

Traubia

display comparably longbranches. Meerow

et al

. (1999) also found elevatedterminal branch lengths for these taxa.

Xerolirion

(Asparagaceae, Laxmanniaceae) is another taxon withan anomalously long branch.

Age estimates for crown and stem nodes at thefamily level and above are shown in Figure 1 and inTable 1. Disregarding a few younger clades inAlismatales, all divergences are older than 65 Myr.With the exception of Zingiberales (88 Myr), all ordersare older than 100 Myr. Most families divergedbetween 65 Mya and 100 Mya. A crown group from theEarly Cretaceous (> 100 Mya) has been found for

Araceae, Petrosaviaceae, Orchidaceae, Arecaceae andDasypogonaceae.

DISCUSSION

P

HYLOGENY

In terms of taxon numbers, our tree is so far the mostcomprehensive of monocots. However, phylogeneticreconstruction was not a major goal of this study, andresolution obtained for all unconstrained nodes has tobe considered with caution because no extensivesearch has been conducted. Below we will discusstopological deviations of our tree from in-depth studiesof limited groups. It is not our intention to discussalternative phylogenetic hypotheses, but to point outpossibly problematic nodes for which dating resultsmay change with the availability of better (multigene)global phylogenetic trees. Chase

et al

. (2000) also pre-sented an extensive family level hypothesis of monocotphylogeny. In their tree, Petrosaviacae are sister toPandanales, although with low support, whereas it isthe sister group of all other core monocots in our anal-ysis. Although Chase et al. (2000) did not sample Alis-matales extensively, they also found Araceae andTofieldiaceae to be sister to Alismatales in that order.Furthermore, they obtained a different resolutionwithin Zingiberales, although with low support andincongruent with the extensive analysis by Kress et al.(2001). Otherwise, there is no important conflictbetween their tree and ours. Notably, we find exactlythe same topology for all unconstrained nodes inCommelinales, Dioscoreales and Pandanales.

Except for the position of Ruppiaceae and Scheuchz-eriaceae, our topology is congruent with that of Leset al. (1997). However, deviations concern familiesthat have been placed with a low bootstrap supportonly (40 and 11%, respectively).

The topology adopted by Bremer (2000) is alsolargely congruent with our tree used for dating. As inour analysis, and unlike Vinnersten & Bremer (2001),Campynemataceae are sister to the rest of Liliales.Orchids, however, cluster within the Boryaceae–Lanariaceae clade, whereas this clade is sister toOrchidaceae and the rest of Asparagales in our anal-ysis. Bromeliaceae are not sister to Rapateaceae inour tree, but to the graminoid clade including Erio-caulaceae and Xyridaceae. Mayacaceae are sister toEriocaulaceae and Xyridaceae, whereas in our treethis family is sister to Hydatellaceae and close toCyperaceae. The latter conflict may probably beattributed to a branch attraction phenomenon(Bremer, 2002).

We find the same interrelationship among the threefamilies of Dioscoreales as Caddick et al. (2002a, b).However, in Pandanales, these authors have Vellozi-aceae as sister of the rest and not just to Stemonaceae.

390 T. JANSSEN and K. BREMER

© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 385–398

*

**

*

* **

**

*

*

**

*

*

*

* **

**

*

***

*

**

*

*

*

* *

**

*

*

*

*

*

AGE OF MONOCOT GROUPS 391

© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 385–398

Vinnersten & Bremer (2001) presented a detailedstudy of Liliales. In their tree, Campynemataceae aresister to Melanthiaceae–Smilacaceae–Liliaceae, andthe Alstroemeriaceae–Luzuriagaceae–Colchicaceaeclade is sister to these. However, parsimony branchlengths supporting the deep topology in Liliales arecomparably short. Disregarding the relationshipswithin the enlarged Alliaceae and Asparagaceae inAsparagales (APG II, 2003), our tree is not in conflictwith that of Fay et al. (2000). Unconstrained nodes inour tree correspond to polytomies in their consensustree. Relationships between Boryaceae and relatedfamilies received low support in the analysis by Fayet al. (2000), and it is not surprising that we finda different topology. The position at three basalnodes in Zingiberales (Musaceae, Heliconiaceae andStrelitziaceae–Lowiaceae) differs from that in the treeby Kress et al. (2001), who only obtained a Bremersupport of one for this alternative topology. Finally,comparing our Poales tree with that of Bremer (2002),some unconstrained taxa end up in different positions.This is true for Flagellariaceae and Centrolepidaceae,although they remain within Bremer’s graminoidclade. Restionaceae are monophyletic in our analysis,not including Centrolepidaceae which are placed as asister group of Anarthriaceae–Restionaceae. Bremerfound a Bromeliaceae–Sparganiaceae–Typhaceaeclade sister to the cyperoid clade. However, the nodesupporting this deep level relationship within Poales,as well as the relationships within that clade of threefamilies, received low jackknife support and are hence,not in strong conflict with Bromeliaceae being sisterto Xyridaceae–Eriocaulaceae and Sparganiaceae–Typhaceae being sister to Hydatellaceae–Mayacaceaeand the cyperoid clade. Hydatellaceae and Maya-caceae were excluded from Bremer’s analysis becauseof their unstable positions. In our reconstruction,those two families end up as sister groups, whichprobably is an artefact.

INCREASED TAXON SAMPLING IN PHYLOGENETIC INFERENCE AND DATING

It has been argued that phylogenetic reconstructionprofits, in terms of accuracy and support, from theinclusion of more taxa in the analysis (Källersjö et al.,1998; Soltis et al., 2000; Rydin & Källersjö, 2002;Zwickl & Hillis, 2002). However, few indications are

available as to whether the same is true for moleculardating. Extended sampling may level out biases intro-duced by an overrepresentation of groups with highsubstitution rates, such as grasses. Their exaggeratingeffect on overall age estimates (especially at deeperlevels) should be cancelled out by the inclusion of slowgroups, such as palms. In dating, adding more genes tothe data set is not likely to be helpful because ratevariation appears to be systematic and lineage-specific, such that different molecular markers areaffected in the same way and will thus not alleviatedifferences among lineages (Sanderson & Doyle,2001). Adding more taxa seems to remain the onlyoption to improve data sets for age inference. Hence,the purpose of increased taxon sampling in this studyis not only to estimate crown node ages, but we alsoexpect it to increase precision of age estimates.

Even though the effect of uneven rates causingdifferent branch lengths, for example in groups withan atypical life history such as aquatics or parasites,cannot be entirely removed by adding taxa to the tree,excessively long branches may be broken up and theremay be a ‘smoothing’ of rate differences between adja-cent regions of the tree. This should, in turn, increasecorrespondence with the underlying assumption innonparametric rate smoothing (Sanderson, 1997),namely that rates are autocorrelated between lineagesand abrupt changes do not occur.

Sanderson (1990) noted that reduced taxon sam-pling results in reduced branch length estimates,since hidden homoplasy may give rise to additionalcharacter state transformations when more taxa areadded to the phylogeny. Hence, overall branch lengthswould become longer, and optimization methods wouldbe likely to find older ages at deeper nodes. Indeed,there seems to be a general tendency towards pushingdivergence times back in time with increased taxonsampling.

DIVERGENCE TIMES

In calibrating our tree we followed Bremer (2000)who, based on eight reference fossils, estimated thesplit between Acorus and all other monocots at134 Mya, the only reliable calibration available to thisdate. According to our dating (Fig. 1), much diversifi-cation took place before 100 Mya during the EarlyCretaceous. With a few exceptions, all family stem

Figure 1. Dated phylogenetic tree of monocot families obtained from analysis of 878 rbcL sequences. Thick bars showinferred crown node ages (absent in families represented by a single sequence only). The number of rbcL sequences andthe stem and crown node ages for each family are given in Table 1. Asterisks represent nodes not included in the enforcedtopological constraints compiled from well-supported clades in various published analyses (listed in the text); all otherbranches represent well supported clades included in a constraint tree used in the analysis. The two vertical lines showthe mid-Cretaceous 100 Mya and the Cretaceous–Tertiary boundary 65 Mya.

�

392 T. JANSSEN and K. BREMER

© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 385–398

lineages were present by 65 Mya at the Cretaceous–Tertiary boundary. Moreover, a considerable portion ofthe family crown nodes dates back to the LateCretaceous.

Bremer (2000) generally obtained younger ages. Wefound about three times as many Early Cretaceousmonocot lineages as did Bremer. He used parsimonybranch lengths, which tend to underestimate changeand may result in younger ages compared with maxi-mum likelihood estimates. Furthermore, there may beunknown methodological effects leading to youngerage estimates with mean-path-lengths or older ageestimates with nonparametric rate smoothing. Evenmore importantly, sampling was much more limitedin Bremer’s study compared with this paper and,as noted above, increased sampling may lead toincreased age estimates.

Our age estimates for Liliales are 10–50 Myr olderthan those presented by Vinnersten & Bremer (2001).This is probably related to the difference between ourstudy and that of Bremer (2000), since Vinnersten &Bremer calibrated the Liliales crown node at 82 Myafollowing Bremer (2000). We estimate this node at117 Mya, and if the tree by Vinnersten & Bremer iscalibrated with our crown node age estimate for theorder, then age estimates within the order becomelargely similar.

Within Poales, our results are remarkably congru-ent with the age estimates of Bremer (2002). Mostdeviations are within the expected confidence inter-vals. Bromeliaceae are about 26 Myr older in ouranalysis, which might be a result of their differentplacement in the tree. This effect is more dramaticfor Centrolepidaceae, the stem node of which wasestimated to be at 97 Mya but around 45 Mya whennested within Restionaceae. We find Cyperaceae36 Myr older than did Bremer, a difference probablydue to differences in sampling. Bremer sampledeight taxa of Cyperaceae compared with 48 in ouranalysis.

It emerges from the above discussion thatincreased sampling pushes divergence times backin time. Although some theoretical considerations(Sanderson, 1990) might explain such behaviour to acertain extent, this tendency is worrying. The alreadyexisting gap between molecular age estimates and theages obtained from the fossil record will be enhancedunless we hit an upper limit for the ages at a certainsampling density. The observed phenomenon mightalso, however, be a systematic error and may point tothe necessity of methodological improvement inmolecular dating. If the observed tendency is not amethodological artefact, then we must assume thatcurrent age estimates are generally too youngbecause the present sampling is far from complete inmost families.

The crown nodes of Araceae, Petrosaviaceae,Orchidaceae, Arecaceae and Dasypogonaceae havebeen estimated to date back to the Early Creta-ceous. Among these, Petrosaviaceae and Dasy-pogonaceae are small families with a limitedrepresentation in our data, and they occupy posi-tions as sister to major clades in core monocots andcommelinids, respectively. In these cases the ageestimates are certainly enhanced by the phyloge-netic positions and should be interpreted withcaution. It is not surprising to find Araceae andArecaceae among the old families, with macrofossilsbeing known from the Albian and the Campanian,respectively (Herendeen & Crane, 1995). A crownnode age of 111 Myr for Orchidaceae is unexpected,however. Traditionally, this family has been lookedat as a very specialized and hence, probably, a younggroup. Considering the extensive sampling withinorchids (145 genera) and its firm phylogeneticposition at the base of the Asparagales, this ageestimate appears to be well supported. However, amethodological bias due to the extended samplingstill cannot be excluded. If our age estimate turnsout to be true, the evolutionary history of this fam-ily could be seen in a new light. Orchid diversity isnot necessarily due to a rapid and recent radiation,and similar patterns, for example in palms, might behypothesized.

DIVERGENCE TIMES AND THE FOSSIL RECORD

Molecular age estimates generally predate fossil ages.The fossil record of monocots is comparatively poor,with few fossils attributable to families reaching backbeyond the Maastrichtian. Reasons for this may bethe herbaceous habit and widespread zoophilouspollination (Herendeen & Crane, 1995). Monocotpollen is known from the Early Cretaceous (125 Mya),and fossils attributable to taxonomic groups date fromthe Turonian (Arecaceae pollen, 90 Mya) and theSantonian–Campanian boundary (Zingiberales fruits,83 Mya; Bremer, 2000). The ages estimated here aresomewhat older; for monocots 9 Myr, for Arecaceae10 Myr and for Zingiberales 5 Myr older.

From fossil evidence, a major radiation ofangiosperms is obvious in the mid-Cretaceous 130–90Mya (Lidgard & Crane, 1990; Herendeen & Crane,1995). With the diversification burst of angiospermssetting in no earlier than about 115 Mya, there stillremains a considerable gap between the fossil recordand our molecular dating, with the majority of lin-eages already present by that time. At this time wecannot decide whether this is an indication of anincomplete fossil record or an argument against cur-rently available molecular dating methodologies.

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DIVERGENCE TIMES – ERROR SOURCES

Molecular dating may be affected by a variety of errorsources (Sanderson & Doyle, 2001), the majority ofwhich are not sufficiently theoretically understood.Substitutional noise as a stochastic characteristic ofthe substitution process as such imposes a lowerboundary on error and finds its expression in minimalconfidence intervals. Branch length estimation will beaffected by rate variation across sites and across lin-eages, as well as by the optimization method used.Rate variation across sites might lead to incorrect esti-mates, especially at high rates (Sanderson & Doyle,2001). According to these authors, the use of a gammadistribution in the maximum likelihood model leads to10–30 Myr younger ages compared with the assump-tion of equal rates.

Rate variation across lineages is harder to dealwith. Clock-like behaviour over the whole topologywould be ideal for molecular dating, but with more lin-eages being included in the analysis this assumptionbecomes more and more unlikely. NPRS (Sanderson,1997) is a dating method that deals with rate hetero-geneities by minimizing rate changes between adja-cent branches. The assumption of rate autocorrelationappears to be reasonable, but there is also no a priorireason to exclude the possibility of abrupt ratechanges between adjacent lineages. Until more workhas been done on the evolution of evolutionary rates,dating will largely depend on the underlying hypoth-eses and the optimization method used.

Molecular dating is unable to provide absoluteages. The phylogenetic tree has to be placed in anappropriate stratigraphic context to calibrate theobtained relative ages. Calibration is a major errorsource because it depends on the dating of the fossilitself and on its attachment point to the phylogeny.Hence, several reference fossils are desirable in largetrees comprising a variety of taxa and considerablerate heterogeneity.

We found that, for a restricted taxon sampling, theages inferred from rbcL were only comparable withthose obtained with a three-gene matrix (rbcL + atpB+ 18S rDNA). It has been argued that evolutionaryrates do not differ significantly among different cellu-lar compartments (Sanderson & Doyle, 2001). If this istrue, the bias introduced using a single gene for datingmight be negligible.

Dating largely depends on the underlying topologyused to estimate branch lengths. Nodes marked withan asterisk in Figure 1 have not been constrainedprior to phylogenetic reconstruction. Alternativetopologies, especially those congruent with earlierstudies (see above), are less parsimonious. However,topological rearrangements at these nodes mightaffect age inference. Branches are short at a deeper

level in our tree, and rearrangements at this level onlyslightly affect branch length estimates. Rearrange-ments at a higher level should only slightly affectbranch lengths within the rearranged clade. Underthe assumption that no major rearrangements may beexpected, the topological bias should be reduced to anacceptable minimum. The absence of major conflict(see above) between our topology and earlier recon-structions renders this assumption reasonable.

Sampling biases, especially an overrepresentationof rapidly evolving herbaceous lineages, may bealleviated by including more and diverse taxa. Ourapproach, to include information from all availablegenera, is the only way to avoid a priori selection ofdata, but it may not necessarily lead to a representa-tive taxon sampling. Two families, Orchidaceae andArecaceae, may be over-represented in our data set.

Sanderson & Doyle (2001) pointed out that non-clocklike behaviour of evolutionary rates might leadto significant deviation among results obtained withdifferent dating methods. Different methods mayintroduce systematic biases, which are generally hardto detect. If our finding that increased sampling leadsto older age estimates is corroborated in the future,then current dating methods need revision.

ACKNOWLEDGEMENTS

We thank J. Nylander for computer assistance and T.Britton for running MPL analyses. The study wassupported by a Swedish Research Council grant to KB.

REFERENCES

APG. 1998. An ordinal classification for the families offlowering plants. Annals of the Missouri Botanical Garden85: 531–553.

APG II. 2003. An update of the Angiosperm Phylogeny Groupclassification for the orders and families of flowering plants:APG II. Botanical Journal of the Linnean Society 141: 399–436.

Baldwin BG, Sanderson MJ. 1998. Age and rate of diversi-fication of the Hawaiian silversword alliance (Compositae).Proceedings of the National Academy of Sciences, USA 95:9402–9406.

Bremer K. 2000. Early Cretaceous lineages of monocotflowering plants. Proceedings of the National Academy ofSciences, USA 97: 4707–4711.

Bremer K. 2002. Gondwanan evolution of the grass alliance offamilies (Poales). Evolution 56: 1374–1387.

Bremer K, Friis EM, Bremer B. 2004. Molecular phyloge-netic dating of asterid flowering plants shows Early Creta-ceous diversification. Systematic Biology 53: 496–505.

Britton T. 2002. PATH. Computer program and documentationavailable from http://www.math.uu.se/~tomb/PATH.html

Britton T, Oxelman B, Vinnersten A, Bremer K. 2002.Phylogenetic dating with confidence intervals using mean

394 T. JANSSEN and K. BREMER

© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 385–398

path lengths. Molecular Phylogenetics and Evolution 24: 58–65.

Caddick LR, Rudall PJ, Wilkin P, Hedderson TAJ, ChaseMW. 2002a. Phylogenetics of Dioscoreales based on com-bined analyses of morphological and molecular data. Botan-ical Journal of the Linnean Society 138: 123–144.

Caddick LR, Wilkin P, Rudall PJ, Hedderson TAJ, ChaseMW. 2002b. Yams reclassified: a recircumscription ofDioscoreaceae and Dioscoreales. Taxon 51: 103–114.

Chase MW, Soltis DE, Soltis PS, Rudall PJ, Fay MF,Hahn WH, Sullivan S, Joseph J, Molvray M, Kores PJ,Givnish TJ, Sytsma KJ, Pires JC. 2000. Higher-level sys-tematics of the monocotyledons: An assessment of currentknowledge and a new classification. In: Wilson KL, MorrisonDA, eds. Monocots: systematics and evolution. Melbourne:CSIRO. 3–16.

Fay MF, Rudall PJ, Sullivan S, Stobart KL, de Bruijn AY,Reeves G, Qamaruze-Zaman F, Hong W-P, Joseph J,Hahn WJ, Conran JG, Chase MW. 2000. Phylogeneticstudies of Asparagales based on four plastid DNA regions.In: Wilson KL, Morrison DA, eds. Monocots: systematics andevolution. Melbourne: CSIRO, 360–371.

Givnish TJ, Evans TM, Zjhra ML, Patterson TB, BerryPE, Sytsma KJ. 2000. Molecular evolution, adaptive radi-ation, and geographic diversification in the amphiatlanticfamily Rapateaceae: evidence from ndhF sequences and mor-phology. Evolution 54: 1915–1937.

Herendeen PS, Crane PR. 1995. The fossil history of themonocotyledons. In: Rudall PJ, Cribb PJ, Cutler DF,Humphries CJ, eds. Monocotyledons: systematics and evolu-tion. Kew: Royal Botanic Gardens, 1–21.

Källersjö M, Farris JS, Chase MW, Bremer B, Fay MF,Humphries CJ, Petersen G, Seberg O, Bremer K. 1998.Simultaneous parsimony jackknife analysis of 2538 rbcLDNA sequences reveals support for major clades of greenplants, land plants, seed plants and flowering plants. PlantSystematics and Evolution 213: 259–287.

Kishino H, Thorne JL, Bruno WJ. 2001. Performance of adivergence time estimation method under a probabilisticmodel of rate evolution. Molecular Biology and Evolution 18:352–361.

Kress WJ, Prince LM, Hahn WJ, Zimmer EA. 2001. Unrav-elling the evolutionary radiation of the families of theZingiberales using morphological and molecular evidence.Systematic Biology 50: 926–944.

Les DH, Cleland MA, Waycott M. 1997. Phylogeneticstudies in Alismatidae, II: Evolution of marine angiosperms(seagrasses) and hydrophily. Systematic Botany 22: 443–463.

Lidgard S, Crane PR. 1990. Angiosperm diversification andCretaceous floristic trends: a comparison of palynofloras andleaf macrofloras. Paleobiology 16: 77–93.

Meerow AW, Fay MF, Guy CL, Li Q-B, Zaman FQ, ChaseMW. 1999. Systematics of Amaryllidaceae based on cladisticanalysis of plastid rbcL and trnL-F sequence data. AmericanJournal of Botany 86: 1325–1345.

Rydin C, Källersjö M. 2002. Taxon sampling and seed plantphylogeny. Cladistics 18: 485–513.

Sanderson MJ. 1990. Estimating rates of speciation andevolution: a bias due to homoplasy. Cladistics 6: 387–391.

Sanderson MJ. 1997. A nonparametric approach to estimat-ing divergence times in the absence of rate constancy. Molec-ular Biology and Evolution 14: 1218–1231.

Sanderson MJ. 1998. Estimating rate and time in molecu-lar phylogenies: Beyond the molecular clock? In: Soltis DE,Soltis PS, Doyle JJ, eds. Molecular systematics of plants II:DNA sequencing. Boston: Kluwer Academic Press, 242–264.

Sanderson MJ. 1999. r8s 0.20. Computer program anddocumentation available from http://phylo.ucdavis.edu/r8s/r8s.html

Sanderson MJ. 2002. Estimating absolute rates of molecularevolution and divergence times: a penalized likelihoodapproach. Molecular Biology and Evolution 19: 101–109.

Sanderson MJ, Doyle JA. 2001. Sources of error and confi-dence intervals in estimating the age of angiosperms fromrbcL and 18S rDNA data. American Journal of Botany 88:1499–1516.

Soltis DE, Soltis PS, Chase MW, Mort ME, Albach DC,Zanis M, Savolainen V, Hahn WH, Hoot SB, Fay MF,Axtell M, Swenson SM, Prince LM, Kress WJ, NixonKC, Farris JS. 2000. Angiosperm phylogeny inferred from18S rDNA, rbcL, and atpB sequences. Botanical Journal ofthe Linnean Society 133: 381–461.

Stockey RA. 2003. The fossil record of basal monocots.Abstract in Monocots III. Los Angeles, California: RanchoSanta Ana Botanic Garden, 89.

Swofford DL. 2001. PAUP: phylogenetic analysis using par-simony (*and other methods). Version 4.0b6. Sunderland:Sinauer Associates.

Thorne JL, Kishino H. 2002. Divergence time and evolution-ary rate estimation with multilocus data. Systematic Biology51: 689–702.

Vinnersten A, Bremer K. 2001. Age and biogeography ofmajor clades in Liliales. American Journal of Botany 88:1695–1703.

Wikström N, Savolainen V, Chase MW. 2001. Evolution ofthe angiosperms: calibrating the family tree. Proceedings ofthe Royal Society of London B 268: 2211–2220.

Wilson MA, Gaut B, Clegg MT. 1990. Chloroplast DNAevolves slowly in the palm family (Arecaceae). MolecularBiology and Evolution 7: 303–314.

Zwickl DJ, Hillis DM. 2002. Increased taxon samplinggreatly reduces phylogenetic error. Systematic Biology 51:588–598.

APPENDIX

GenBank accession numbers to the rbcL sequences forall genera sampled.

INCERTAE SEDIS

Dasypogonaceae: Calectasia (AJ286557), Dasypogon(AF206758), Kingia (AJ404840), Petrosaviaceae: Japo-nolirion (AF206784), Petrosavia (AF206806).

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ACORALES

Acoraceae: Acorus (M91625).

ALISMATALES

Alismataceae: Alisma (L08759), Baldellia (U80677),Damasonium (U80678), Echinodorus (U80679), Luro-nium (U80680), Ranalisma (U80681), Sagittaria(L08767), Wiesneria (U80682), Aponogetonaceae:Aponogeton (U80684), Araceae: Amorphophallus(AF497060), Anchomanes (AF497108), Anthurium(AJ005627), Ariopsis (L10255), Arisaema (AF497109),Dieffenbachia (AJ005631), Filarum (AF497110),Gonatopus (AF497111), Gymnostachys (M91629),Hapaline (AF497112), Landoltia (AY034223), Lasia(L10250), Lemna (AY034234), Montrichardia (L10248),Orontium (AJ005632), Peltandra (AJ005628), Philoden-dron (AJ005623), Pistia (M96963), Pseudodracontium(AF497106), Scindapsus (AJ005625), Spathiphyllum(AJ235807), Spirodela (AY034222), Symplocarpus(L10247), Typhonium (AF497113), Wolffia (AY034254),Wolffiella (AY034244), Xanthosoma (AJ007543),Zamioculcas (AJ005624), Zantedeschia (AF065474),Butomaceae: Butomus (U80685), Cymodoceaceae:Amphibolis (U80686), Cymodocea (U80687), Halodule(U80689), Syringodium (U03727), Thalassodendron(U80692), Hydrocharitaceae: Apalanthe (U80693),Blyxa (U80694), Egeria (AB004887), Elodea(AB004888), Enhalus (AB004889), Halophila (U80698),Hydrilla (AB004891), Hydrocharis (U80701), Lagarosi-phon (AB004893), Limnobium (AB004894), Najas(U03731), Nechamandra (U80706), Ottelia (AB004895),Stratiotes (AB004896), Thalassia (AB004897), Vallisne-ria (U03726), Juncaginaceae: Cycnogeton (U80713),Lilaea (U80715), Triglochin (U80714), Limnocharita-ceae: Hydrocleys (AB004900), Limnocharis (U80717),Posidoniaceae: Posidonia (U80718), Potamogetonaceae:Coleogeton (U80727), Groenlandia (U80720), Lepilaena(U80729), Potamogeton (L08765), Zannichellia(U03725), Ruppiaceae: Ruppia (U03729), Scheuchzeri-aceae: Scheuchzeria (U03728), Tofieldiaceae: Pleea(AJ131774), Tofieldia (AJ286562), Zosteraceae: Hetero-zostera (U80730), Phyllospadix (U80731), Zostera(AY077964).

ARECALES

Arecaceae: Acrocomia (AY044625), Aiphanes(AJ404831), Allagoptera (AJ404828), Ammandra(AJ404838), Aphandra (AJ404837), Archontophoenix(AF449156), Areca (AJ404819), Arenga (AJ404788),Asterogyne (AJ404833), Astrocaryum (AY012510),Attalea (AJ404829), Bactris (AY044627), Balaka(AJ404814), Barcella (AY044630), Beccariophoenix(AJ404826), Bentinckia (AY012499), Borassodendron(AJ404768), Borassus (AY012469), Brassiophoenix(AJ404815), Burretio (AY012500), Butia (AY044632),Calamus (AJ404775), Calyptrocalyx (AY012501), Calyp-tronoma (AJ404832), Caryota (AJ404790), Ceroxylon(AJ404781), Chamaedorea (AF206748), Chamaerops(AJ404754), Chambeyronia (AY012489), Chelyocarpus(AY012457), Chuniophoenix (AJ404764), Clinostigma

(AF449157), Coccothrinax (AJ404751), Cocos(AY012507), Corypha (AJ404762), Cryosophila(AJ404747), Cyphophoenix (AJ404821), Cyrtostachys(AJ404810), Desmoncus (AY044628), Dictyocaryum(AY012479), Dictyosperma (AY012503), Drymophloeus(AY012494), Dypsis (AY012486), Elaeis (AJ404830), cf.Eremospatha (AJ404773), Eugeissona (AJ404774),Euterpe (AJ404802), Gastrococos (AY044629), Gaussia(AJ404784), Geonoma (AJ404834), Gronophyllum(AJ404816), Guihaia (AJ404755), Hedyscepe(AJ404807), Howea (AY012492), Hydriastele(AJ404817), Hyophorbe (AJ404785), Hyospathe(AJ404804), Hyphaene (AY012470), Iguanura(AJ404820), Iriartea (AF233088), Itaya (AJ404748),Johannesteijsmannia (AJ404758), Kentiopsis(AJ404809), Kerriodoxa (AJ404765), Laccospadix(AJ404812), Lemurophoenix (AJ404801), Leopoldinia(AJ404798), Licuala (AY012462), Linospadix(AF449158), Livistona (AJ404757), Lodoicea(AJ404769), Lytocaryum (AY044633), Manicaria(AJ404797), Marojejya (AJ404825), Masoala(AJ404824), Mauritia (AJ404777), Metroxylon(AF233089), Nannorrhops (AJ404763), Nenga(AJ404818), Neonicholsonia (AJ404803), Nypa(AJ404778), Oenocarpus (AY044624), Oncocalamus(AJ404776), Oncosperma (AY012505), Orania(AJ404796), Oraniopsis (AJ404782), Orbignya(AY012508), Phoenix (M81814), Phytelephas(AJ404835), Podococcus (AF233090), Prestoea(AY012487), Pritchardiopsis (AY012464), Pseudophoe-nix (AJ404779), Ptychosperma (AY012495), Ravenea(AY012475), Reinhardtia (AJ404799), Rhapidophyl-lum (AJ404753), Rhapis (AJ404756), Rhopalostylis(AJ404808), Roscheria (AJ404822), Roystonea(AY012488), Sabal (AJ404766), Salacca (AY012472),Satranala (AJ404771), Scheelea (AY044636), Schippia(AJ404749), Sclerosperma (AJ404823), Serenoa(AJ404760), Socratea (AY012480), Syagrus (AY044634),Synechanthus (AJ404786), Thrinax (AJ404750),Trachycarpus (AJ404752), Trithrinax (AJ404745), Veit-chia (AJ404813), Voanioa (AY044635), Wallichia(AJ404792), Washingtonia (AY012465), Wendlandiella(AY012477), Wettinia (AJ404794).

ASPARAGALES

Alliaceae: Acis (Z77256), Agapanthus (Z69221), Allium(Z69204), Amaryllis (Z69219), Boophane (AF116945), cf.Brunsvigia (AF116946), Caliphruria (AF116947),Calostemma (AF116948), Clivia (L05032), Crinum(AF116951), Cryptostephanus (AF116952), Cyrtanthus(AF116953), Eucharis (AF116954), Eucrosia(AF116955), Eustephia (AF116956), Galanthus(Z69218), Gethyllis (AF116957), Gilliesia (Z69208),Griffinia (AF116958), Habranthus (AF116959), Hae-manthus (AF116960), Hannonia (AF116961), Hessea(AF116962), Hieronymiella (AF116963), Hippeastrum(AF206776), Hymenocallis (AF116965), Ipheion(Z69201), Ismene (AF116966), Leptochiton (AF116970),Leucocoryne (Z69199), Lycoris (AB034753), Milula(AF116991), Narcissus (AF116972), Nerine (AF116973),Nothoscordum (Z69202), Pamianthe (AF116974), Pan-cratium (AF116975), Paramongaia (AF116976), Phae-

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dranassa (AF116977), Proiphys (AF116978), Rauhia(AF116979), Rhodophiala (AF116980), Scadoxus(AF116981), Solaria (Z69207), Sprekelia (AF116982),Stenomesson (Z69217), Sternbergia (AF116984), Stru-maria (AF116985), Traubia (AF116986), Tristagma(Z69206), Tulbaghia (Z69203), Ungernia (AF116987),Vagaria (AF116988), Worsleya (AF116989), Zephyran-thes (AF116990), Asparagaceae: Acanthocarpus(Z77296), Agave (AF206729), Albuca (Z69223),Androstephium (AJ311060), Anemarrhena (Z77251),Anthericum (Z69225), Aphyllanthes (Z77259), Arthropo-dium (Z69233), Asparagus (L05028), Aspidistra(Z77269), Behnia (Z69226), Bessera (Z69215), Bowiea(Z69237), Brodiaea (Z69210), Calibanus (Z77276),Camassia (Z69238), Campylandra (AB029835),Chamaexeros (Z77298), Chlorogalum (Z69228), Chloro-phytum (L05031), Comospermum (Z73679), Convallaria(D28334), Danae (L05034), Dasylirion (AB029847),Dichelostemma (AJ311063), Disporopsis (D17373), Dra-caena (AB029848), Echeandia (Z69212), Eriospermum(Z77277), Hemiphylacus (Z73688), Herreria (Z69230),Heteropolygonatum (AB029831), Hosta (L10253), Hya-cinthus (AF116995), Ledebouria (L05038), Leucocrinum(Z77252), Liriope (Z77271), Lomandra (L05039), Maian-themum (D17378), Muilla (Z69213), Muscari (Z77278),Myrsiphyllum (Z77260), Nolina (L05030), Ophiopogon(AB029840), Ornithogalum (Z69224), Paradisea(Z69229), Peliosanthes (AB029843), Polianthes(Z69227), Polygonatum (AB029827), Reineckea(AB029834), Rohdea (AB029836), Ruscus (Z77274),Sansevieria (Z73698), Scilla (D28161), Semele (Z77275),Smilacina (D17380), Sowerbaea (Z69234), Thysanotus(Z69236), Tricalistra (AB029839), Triteleia (Z69198),Tupistra (AB029838), Whiteheadia (Z77279), Xerolirion(Z77299), Asteliaceae: Astelia (Z77261), Collospermum(Y14986), Milligania (Z73693), Blandfordiaceae: Bland-fordia (Z73694), Boryaceae: Alania (Y14982), Borya(Z77262), Doryanthaceae: Doryanthes (Z73697), Hypoxi-daceae: Curculigo (Z73701), Empodium (Y14987),Hypoxis (Z73702), Pauridia (Y14991), Rhodohypoxis(Z77280), Spiloxene (Z77281), Iridaceae: Alophia(AJ309678), Anomatheca (Z73703), Aristea (Z77282),Babiana (AJ309673), Belamcanda (AJ307078), Bobar-tia (AJ307079), Calydorea (AJ309682), Chasmanthe(AJ309660), Cipura (AJ309681), Crocus (AJ309668),Cypella (AJ309683), Dietes (AJ307080), Diplarrhena(AJ309686), Eleutherine (Z77283), Ennealophus(AJ309684), Ferraria (AJ307081), Freesia (Z77284), Gal-axia (AJ309685), Geissorhiza (AJ309676), Gelasine(AJ309674), Geosiris (Z77285), Gladiolus (Z77286),Gynandriris (AJ309698), Herbertia (AJ309692), Hesper-antha (AJ309656), Hesperoxiphion (AJ309677), Home-ria (AJ309691), Iris (L05037), Isophysis (Z77287), Ixia(Z77288), Klattia (AJ309667), Lapeirousia (AJ309665),Libertia (AJ309687), Micranthus (AJ309662), Moraea(AJ307084), Neomarica (AJ309679), Nivenia (Z77289),Olsynium (AJ309688), Orthrosanthus (L10249), Par-danthopsis (AJ309696), Patersonia (AJ277879), Pillan-sia (AJ309671), Radinosiphon (AJ309661), Romulea(AJ309659), Savannosiphon (AJ309664), Schizostylis(AJ309657), Sisyrinchium (Z77290), Solenomelus(AJ309689), Sparaxis (AJ309669), Syringodea(AJ309670), Thereianthus (AJ309663), Tigridia(AJ309680), Trimezia (AJ309672), Tritonia (AJ309675),

Tritoniopsis (AJ309658), Watsonia (AJ309666), Witse-nia (AJ277880), Ixioliriaceae: Ixiolirion (Z73704),Lanariaceae: Lanaria (Z77313), Orchidaceae: Acanthep-hippium (AF074100), Acianthus (AF074101), Acineta(AF074102), Aeranthes (AF074104), Altensteinia(AF074105), Ancistrochilus (AF264152), Angraecum(AF074106), Ansellia (AF074107), Anthogonium(AF264153), Aplectrum (AF074108), Apostasia(Z73705), Arethusa (AF074109), Arpophyllum(AF074110), Arundina (AF074111), Basiphyllaea(AF264155), Bifrenaria (AF074112), Bletia (AF264156),Bletilla (AF074114), Bothriochilus (AF264158), Bulbo-phyllum (AF074115), Cadetia (D58406), Caladenia(AF074116), Calanthe (AF264159), Calochilus(AF074118), Calopogon (AF074119), Calypso(AF074120), Catasetum (AF074121), Cattleya(AF074122), Cephalanthera (AF074123), Chiloglottis(AF074124), Chloraea (AF074125), Chysis (AF074126),Cleisostoma (AF074130), Cleistes (AF074127), Clem-atepistephium (AF074131), Coelia (AF074132), Coelog-yne (AF074133), Collabium (AF264163), Coryanthes(AF074134), Corybas (AF074135), Corymborkis(AF074136), Cranichis (AF074137), Cryptarrhena(AF074138), Cryptocentrum (AF074139), Cryptostylis(AF074140), Cymbidium (AF074141), Cypripedium(AF074142), Cyrtopodium (AF074143), Dendrobium(AF074145), Dendrochilum (AF264164), Diaphananthe(AF074147), Diceratostele (AF074148), Dichaea(AF074149), Dilochia (AF264165), Dilomilis(AF074150), Diplocaulobium (D58409), Disa(AF274006), Diuris (AF074152), Dressleria (AF074153),Duckeella (AF074154), Earina (AF074155), Eleorchis(AF264166), Elleanthus (AF074156), Encyclia(AF074157), Epigeneium (D58410), Epipactis (Z73707),Epistephium (AF074162), Eria (AF074164), Eriaxis(AF074165), Eriochilus (AF074166), Eriopsis(AF074167), Erythrorchis (AF074168), Eulophia(AF074170), Flickingeria (D58411), Galeandra(AF074171), Glomera (AF074172), Glossodia(AF074173), Goodyera (AF074174), Govenia(AF074175), Grammatophyllum (AF074176), Habe-naria (AF074177), Hexalectris (AF264168), Houlletia(AF074178), Huntleya (AF074179), Isotria (AF074180),Kegeliella (AF074181), Koellensteinia (AF074182),Liparis (AF074183), Listera (AF074184), Lycaste(AF074185), Lycomormium (AF074186), Lyperanthus(AF074187), Malaxis (AF074188), Masdevallia(AF074189), Maxillaria (AF074190), Megastylis(AF074191), Meiracyllium (AF074192), Mexipedium(AF074193), Microtis (AF074194), Mischobulbum(AF264169), Monophyllorchis (AF074195), Mormodes(AF074196), Neofinetia (AF074197), Neomoorea(AF074198), Nephelaphyllum (AF264170), Nervilia(AF074199), Neuwiedia (AF074200), Oncidium(AF074201), Ophrys (AF074202), Orthoceras(AF074204), Pachyplectron (AF074205), Palmorchis(AF074206), Paphiopedilum (AF074207), Phaius(AF074210), Phalaenopsis (AF074211), Phragmipe-dium (AF074212), Phreatia (AF074214), Platanthera(AF074215), Platythelys (AF074216), Pleione(AF264173), Pleurothallis (AF074217), Podochilus(AF074218), Pogonia (AF074219), Polystachya(AF074222), Ponthieva (AF074223), Pseuderia(D58412), Pterostylis (AF074224), Satyrium

AGE OF MONOCOT GROUPS 397

© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 385–398

(AF074226), Selenipedium (AF074227), Sobralia(AF074228), Spathoglottis (AF264175), Spiranthes(AF074229), Stanhopea (AF074230), Stellilabium(AF074231), Tainia (AF264176), Thelymitra(AF074232), Thunia (AF074233), Tipularia (AF074234),Trichotosia (AF074235), Triphora (AF074236), Tropidia(AF074237), Vanilla (AF074239), Xerorchis (AF074244),Xylobium (AF074245), Zygopetalum (AF074246), Teco-philaeaceae: Conanthera (Z77311), Cyanastrum(U41572), Cyanella (Z77312), Kabuyea (Y17336), Odon-tostomum (Z77314), Tecophilaea (Z73709), Walleria(Y17338), Zephyra (Y17340), Xanthorrhoeaceae: Aloe(L05029), Asphodeline (Z73681), Asphodelus (Z73682),Astroloba (Z73683), Bulbine (Z73684), Bulbinella(Z73685), Caesia (Z77297), Eremurus (Z73686), Eustre-phus (AF116996), Gasteria (Z73687), Geitonoplesium(AF116997), Haworthia (L05035), Hemerocallis(L05036), Herpolirion (Z77303), Jodrellia (Y17335),Johnsonia (Z77304), Kniphofia (Z73689), Lomatophyl-lum (Z73690), Pasithea (Z77305), Phormium (Z69232),Poellnitzia (Z73691), Simethis (Z69231), Stawellia(Z77306), Stypandra (Z77307), Trachyandra (Z73692),Tricoryne (Z77308), Xanthorrhoea (Z73710), Xerone-mataceae: Xeronema (Z69235).

COMMELINALES

Commelinaceae: Amischotolype (AF312239), Aneilema(AF312252), Anthericopsis (AF312259), Belosynapsis(AF312257), Buforrestia (AF036886), Callisia(AF312248), Cochliostema (AF312244), Coleotrype(AF312243), Commelina (L05033), Cyanotis(AF312241), Dichorisandra (AF312242), Elasis(AF312251), Floscopa (AF312255), Geogenanthus(AF312261), Gibasis (AF312250), Murdannia(AF312256), Palisota (AF312240), Pollia (AF312262),Polyspatha (AF312263), Rhopalephora (AF312264),Siderasis (AF312254), Stanfieldiella (AF312265),Thyrsanthemum (AF312246), Tinantia (AF312260),Tradescantia (L05041), Tripogandra (AF312249), Wel-denia (AF312245), Haemodoraceae: Anigozanthos(AJ286556), Wachendorfia (AF312266), Hanguanaceae:Hanguana (AJ404842), Philydraceae: cf. Helmholtzia(AF206774), Philydrella (AF206808), Pontederiaceae:Eichhornia (U41574), Heteranthera (U41581), Hydro-thrix (U41582), Monochoria (U41588), Pontederia(U41593).

DIOSCOREALES

Burmanniaceae: Burmannia (AF206742), Geomitra(AF307488), Gymnosiphon (AF307489), Dioscoreaceae:Avetra (AF307476), Dioscorea (D28327), Nanarepenta(AF307473), Rajania (AF307472), Stenomeris(AF307475), Tacca (AJ286561), Tamus (AF307474),Trichopus (AF307477), Nartheciaceae: Aletris (M. W.Chase, pers. comm.), Lophiola (AJ417897), Narthecium(AJ286560).

LILIALES

Alstroemeriaceae: Alstroemeria (Z77254), Bomarea(Z77255), Leontochir (AY120369), Campynemataceae:

Campynema (Z77264), Campynemanthe (AJ276349),Colchicaceae: Androcymbium (Z77265), Burchardia(Z77266), Colchicum (L12673), Disporum (D17376), Glo-riosa (D28867), Iphigenia (AJ417893), Petermannia(Z77267), Tripladenia (Z77268), Uvularia (AB009952),Liliaceae: Amana (AB024385), Calochortus (Z77263),Cardiocrinum (AB034918), Clintonia (AB056856),Erythronium (D28156), Fritillaria (Z77293), Gagea(AB034752), Lilium (AB034926), Lloydia (Z77294),Medeola (D28158), Nomocharis (Z77295), Notholirion(AB034919), Prosartes (D17374), Scoliopus (D28163),Streptopus (D17381), Tricyrtis (D17382), Tulipa(Z77292), Luzuriagaceae: Drymophila (AJ276346),Luzuriaga (Z77300), Melanthiaceae: Amianthium(AJ417895), Chamaelirium (AJ276347), Daiswa(D28155), Heloniopsis (AJ417894), Kinugasa (D28157),Melanthium (AJ276348), Paris (D28159), Trillium(AB018845), Veratrum (D28168), Xerophyllum(AJ276350), Philesiaceae: Lapageria (Z77301), Philesia(Z77302), Rhipogonaceae: Rhipogonum (Z77309), Smila-caceae: Smilax (Z77310).

PANDANALES

Cyclanthaceae: Carludovica (AF197596), Cyclanthus(AY007660), Ludovia (L10251), Sphaeradenia(AJ235808), Pandanaceae: Freycinetia (AF206770),Pandanus (M91632), Stemonaceae: Croomia (D28154),Pentastemona (AF307490), Stemona (AJ131948), Sti-choneurone (AF307492), Velloziaceae: Barbacenia(AJ131946), Vellozia (L19970).

POALES

Anarthriaceae: Anarthria (AF148760), Hopkinsia(AF148777), Lyginia (AF148787), Bromeliaceae: Aech-mea (L19978), Ananas (L19977), Catopsis (L19976),Glomeropitcairnia (L19975), Hechtia (L19974), Puya(L19973), Tillandsia (L19971), Centrolepidaceae:Centrolepis (AF148766), Cyperaceae: Abildgaardia(Y12985), Actinoscirpus (Y12953), Alinula (AJ278290),Ascolepis (Y13003), Becquerelia (Y12948), Blysmus(AJ404700), Bolboschoenus (Y12996), Bulbostylis(Y12992), Carex (Y12999), Carpha (AF307909), Caustis(Y12976), Chorizandra (AJ419939), Chrysithrix(AJ419938), Cladium (Y12988), Coleochloa (Y12975),Courtoisina (AY040590), Cyperus (Y13016), Desmo-schoenus (AJ404701), Eleocharis (Y13012), Eriophorum(Y12951), Ficinia (Y12963), Fimbristylis (Y13009),Fuirena (Y12971), Gahnia (Y12973), Hellmuthia(Y13000), Hypolytrum (Y12956), Isolepis (Y12962),Kobresia (U49232), Kyllinga (Y12979), Kyllingiella(AY040592), Lepironia (Y12957), Lipocarpha (Y12991),Mapania (Y12955), Mesomelaena (Y12959), Nemum(Y12945), Oreobolus (Y12972), Oxycaryum (Y13006),Pleurostachys (Y12989), Pycreus (Y13005), Remirea(AY040593), Rhynchospora (AF206818), Schoenoplectus(Y12943), Schoenus (Y12983), Scirpoides (Y13001),Scleria (Y12968), Scripus (AJ297509), Sphaerocyperus(AJ404699), Trichophorum (Y12969), Ecdeiocoleaceae:Ecdeiocolea (AJ286559), Georgeantha (AF148772),Eriocaulaceae: Eriocaulon (L10252), Lachnocaulon(Y13019), Mesanthemum (AJ419941), Paepalanthus

398 T. JANSSEN and K. BREMER

© 2004 The Linnean Society of London, Botanical Journal of the Linnean Society, 2004, 146, 385–398

(AJ419942), Syngonanthus (AJ419943), Tonina(AF036878), Flagellariaceae: Flagellaria (L12678),Hydatellaceae: Trithuria (AF458076), Joinvilleaceae:Joinvillea (L01471), Juncaceae: Distichia (AJ419944),Juncus (L12681), Luzula (AJ419945), Marsipposper-mum (AJ419946), Oxychloe (Y12978), Rostkovia(AJ419947), Mayacaceae: Mayaca (AJ419948), Poaceae:Amphipogon (U88403), Anomochloa (AF021875), Aris-tida (U31359), Arundo (U13226), Avena (L15300), Bam-busa (M91626), Bromus (Z49836), Cenchrus (L14622),Centropodia (U31100), Chasmanthium (U31101),Chusquea (U13227), Cyperochloa (U88404), Danthonia(U31102), Elymus (Z49837), Elytrophorus (U88405),Enneapogon (U31103), Eragrostis (U31104), Eremium(Z49840), Eriachne (AF352580), Guaduella (AF164778),Gynerium (U31105), Hordeum (Z49842), Hyparrhenia(U31436), Karroochloa (U31437), Leersia (U13228),Leymus (Z49843), Lithachne (U13231), Merxmuellera(U31438), Moliniopsis (U31439), Monachather(U31379), Neurachne (X55827), Oryza (D00207),Pennisetum (L14623), Peridictyon (Z49845), Pharus(AJ419950), Phragmites (U29900), Phyllostachys(U13230), Plinthanthesis (U31440), Pseudoroegneria(Z49846), Puccinellia (L14621), Puelia (AF164780),Rytidosperma (U31441), Setaria (X79900), Stipa(U31442), Stipagrostis (U31378), Streptochaeta(AJ419949), Styppeiochloa (U88406), Thysanolaena(U31380), Tristachya (U31381), Triticum (D00206),Zea (11990232), Zizania (L05043), Rapateaceae:Cephalostemon (AF036884), Kunhardtia (AF036883),Rapatea (AF460969), Schoenocephalium (AF460970),Spathanthus (AF460971), Stegolepis (L19972), Restion-aceae: Acion (AF148762), Alexgeorgea (AF148759),

Baloskion (AF148764), Calorophus (AF148765), Chaet-anthus (AF148782), Chordifex (AF148789), Coleocarya(AF148769), Dapsilanthus (AF148780), Desmocladus(AF148770), Dielsia (AF148771), Elegia (L12675),Empodisma (AF148775), Eurychorda (AF148790),Guringalia (AF148763), Harperia (AF148776), Kulinia(AF148778), Lepidobolus (AF148779), Leptocarpus(AF307924), Lepyrodia (AF148785), Loxocarya(AF148786), Meeboldina (AF148783), Melanostachya(AF148788), Restio (AJ419951), Saropsis (AF148791),Sporadanthus (AF148793), Taraxis (AF148794), Trem-ulina (AF148792), Tyrbastes (AF148795), Winifredia(AF148796), Sparganiaceae: Sparganium (M91633),Thurniaceae: Prionium (U49223), Thurnia (AF036881),Typhaceae: Typha (M91634), Xyridaceae: Abolboda(AJ419952), Aratitiyopea (AF461418), Orectanthe(AF036880), Xyris (AJ286563).

ZINGIBERALES

Cannaceae: Canna (AF378774), Costaceae: Costus(AF243510), Dimerocostus (AF243838), Monocostus(AF243839), Tapeinochilos (AF243840), Heliconiaceae:Heliconia (L05451), Lowiaceae: Orchidantha(AF243841), Marantaceae: Calathea (AF243842),Maranta (AF378768), Marantochloa (AF378769),Pleiostachya (AF378781), Musaceae: Ensete(AF243843), Musa (AF378770), Musella (AF243844),Strelitziaceae: Phenakospermum (AF243845), Ravenala(L05459), Strelitzia (AF243846), Zingiberaceae: Globba(AF243847), Hedychium (AF243848), Riedelia(AF243849), Zingiber (AF243850).