TREE-1847; No.
of Pages 8
Opinion
The changing face of the molecular
evolutionary clock
Simon Y.W. Ho
School of Biological Sciences, University of Sydney, Sydney, NSW, Australia
The molecular clock has played an important role in
biological research, both as a description of the evolu-
tionary process and as a tool for inferring evolutionary
timescales. Genomic data have provided valuable
insights into the molecular clock, allowing the patterns
and causes of evolutionary rate variation to be charac-
terized in increasing detail. I explain how genome
sequences offer exciting opportunities for estimating
the timescale of the Tree of Life. I describe the different
approaches that have been used to deal with the compu-
tational and statistical challenges encountered in molec-
ular clock analyses of genomic data. Finally, I offer a
perspective on the future of molecular clocks, highlight-
ing some of the key limitations and the most promising
research directions.
The molecular clock
One of the fundamental goals of biological research is to
understand the evolutionary process. By allowing the raw
materials of evolution to be analyzed, genetic data have had
an immense impact on this endeavor. In this context, the
molecular clock has been extremely valuable owing to its dual
role as a description of the pattern of molecular evolution and
as a tool for estimating evolutionary rates and timescales.
The importance of the molecular clock has not diminished
over the years, with its role shifting to the analysis of evolu-
tionary patterns and processes on a genomic scale.
The molecular clock hypothesis, which postulates a con-
stancy of evolutionary rates among lineages, was introduced
in the early 1960s [1] and played a part in the development of
molecular evolutionary theory. The apparent homogeneity
of rates among lineages was one of the inspirations for the
neutral theory, which proposed that a large proportion of
mutations do not alter the fitness of an organism [2]. The
neutral theory emphasized the importance of genetic drift
and predicted that evolutionary rates depended on rates of
spontaneous mutation, independently of population size.
According to the neutral theory, however, absolute rates
of evolution (per unit time) are expected to have a negative
relationship with generation length. This is because most
inherited mutations are thought to occur during replication
of germline DNA, and species with long generations there-
fore tend to accumulate fewer average mutations per year.
Corresponding author: Ho, S.Y.W. (simon.ho@sydney.edu.au).
Keywords: molecular clock; genomic data; rate heterogeneity; pacemaker models;
phylogenetic analysis.
0169-5347/
ß 2014 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.tree.2014.07.004
The observation that evolutionary rates in coding sequences
were actually independent of generation length was one of
the motivations for the development of the nearly-neutral
theory shortly afterwards [3]. The nearly-neutral theory
gave population size a key role in governing the relative
impacts of drift and selection. To this day, the neutral theory
and the molecular clock remain important null models in
evolutionary analysis.
The molecular clock is familiar to many researchers as a
tool for estimating evolutionary rates and timescales. Ear-
ly molecular clock studies focused on the timescale of
hominid evolution [4] and were followed by ambitious
efforts to date some of the deepest nodes in the Tree of
Life [5,6]. Alongside these studies was a growing recogni-
tion of rate variation among lineages, in contradiction to
the molecular clock. This motivated the development of
more powerful methods of estimating evolutionary time-
scales, including models that incorporated rate heteroge-
neity across lineages [7]. As a result, the role of the
molecular clock grew dramatically and molecular dating
analyses now form an important component of many evo-
lutionary studies [8,9]. Our understanding of molecular
evolutionary rates has been aided by the growth in genomic
sequence data. These data have brought significant compu-
tational challenges but offer a rich source of information for
resolving the timescale of the Tree of Life.
Heterogeneity in molecular evolutionary rates
Molecular evolution involves dynamic interactions among
the forces of mutation, selection, and drift. As a conse-
quence, rate variation across lineages and across the ge-
nome are ubiquitous features of the evolutionary process.
Large datasets increase the power of statistical methods to
test hypotheses about molecular evolution, including the
impacts of different biological and environmental factors
that affect evolutionary rates.
The causes of rate variation can be broadly divided into
gene effects, lineage effects, and residual effects [10,11].
Gene effects lead to different rates between loci, and have
long been recognized as an intrinsic feature of the molecu-
lar evolutionary process [12]. They can be caused by differ-
ences in the proportion of functionally constrained sites
and by regional heterogeneities in mutation rates across
the genome [11]. Gene effects represent the only form of
rate variation recognized in the simplest clock model,
known as the strict or global molecular clock.
Lineage effects refer to factors that act across the whole
genome, such as differences in particular physiological and
life-history traits. These might include generation time,
Trends in Ecology & Evolution xx (2014) 1–8 1
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Opinion Trends in Ecology & Evolution xxx xxxx, Vol. xxx, No. x

r Deg
ake en
m pa

er
ac

ate
Universal p

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ma

mulple
ker
Mu r
l p l ke
e pace ma

TRENDS in Ecology & Evolution

Figure 1. Comparison of the three pacemaker models of genome evolution [16], each illustrated using rooted four-taxon trees from five loci. In the Universal Pacemaker
model, loci evolve at different rates but share the same pattern of rate variation across branches. In the Multiple Pacemaker model, loci evolve at different rates but
groups of loci share the same patterns of rate variation across branches. In the Degenerate Multiple Pacemaker model, each locus has its own, distinct pattern of rate
variation across branches. These three models involve different interactions between gene effects and lineage effects, leading to contrasting patterns of rate variation
between loci.

metabolic rate, body size, and the performance of DNA
repair mechanisms [13]. Interactions between lineage and
gene effects are known as residual effects and act hetero-
geneously across the genome. Residual effects can be
caused by selection, variation in population size, and other
factors [14,15]. By causing the pattern and extent of
among-lineage rate heterogeneity to vary across loci, re-
sidual effects are particularly important at the genome
scale.
The interplay between gene and lineage effects is en-
capsulated in the pacemaker models of genome evolution
(Figure 1) [16]. The Universal Pacemaker model ascribes
variation in molecular rates to the presence of both gene
and lineage effects. In this model, loci can evolve at distinct
rates from one another, but are all governed equally by
lineage effects. Notably, this model assumes the absence of
residual effects. Studies of archaeal and bacterial genomes
[16,17] and of genome-wide collections of trees from Dro-
sophila and yeast [18] found that the Universal Pacemaker
model explained much of the observed variation in rates.
This stands in contrast with the Degenerate Multiple
Pacemaker model, which assumes that each locus has a
distinct pattern of rate variation across lineages, and
genomic evolution is thereby dominated by residual effects.
2
Between these two models sits the Multiple Pacemaker
model, which involves a moderate level of residual effects.
In this model there are clusters of genes that share the
same lineage effects. This is the most plausible model of
genomic evolution and appears to have some statistical
support [16]. The growing availability of genomic datasets
will enable further comparisons of these models of molec-
ular evolution.
Dating using genome sequences
The molecular clock continues to be an important tool for
estimating evolutionary rates and timescales in the geno-
mic era. There has been a steady increase in the size of
datasets used for such phylogenetic dating analyses, both
inspiring and enabling the development of sophisticated,
parameter-rich models of molecular evolution. Much of the
progress in molecular clock methods over the past few
decades can be described as an effort to account for lineage
effects – that is, dealing with rate variation among
branches in the phylogeny [7]. With the growing use of
large datasets comprising multiple markers, the impacts of
residual effects are becoming increasingly important. To
appreciate how methodological development has benefited
from the increase in genetic data it is worthwhile to
TREE-1847; No. of Pages 8
Opinion
consider how different methods have been designed to
account for the three major forms of rate variation. This
review focuses on sequence data, which are by far the most
widely used type of genome-scale data in molecular clock
analyses.
Accounting for rate variation across lineages
For the first few decades of its history, the term ‘molecular
clock’ referred to the strict-clock model, which does not
account for lineage effects. This still provides a useful null
model for testing rate variation among lineages. In addi-
tion, use of the strict clock remains commonplace in anal-
yses of datasets with low genetic variation, such as those
comprising samples from a single population or from close-
ly related species [19].
Since the late 1990s there has been a proliferation of
methods that relax the assumption of rate constancy [7].
These relaxed-clock models allow the rate to vary across
lineages such that each branch of the phylogeny can have a
distinct evolutionary rate. The various relaxed-clock mod-
els make different assumptions about how rates vary
throughout the tree, such as the degree of correlation
between rates in neighboring branches. These models have
been reviewed and compared in detail [7,20], although
some uncertainty remains about their relative merits
[21–23].
The wide range of molecular clock models has made the
estimation of evolutionary rates and timescales a substan-
tially statistical exercise. However, using an inappropriate
clock model can produce highly misleading estimates of
evolutionary rates and timescales [24–27]. Accordingly,
choosing an appropriate model is an important step in
any phylogenetic dating analysis, and there are several
methods that can be used for model selection. Although the
various clock models provide different descriptions of rate
variation across lineages, they do not make any statements
about absolute rates or node times. In this respect, all
molecular clock methods share a reliance on calibrations,
which are usually informed by paleontological or geological
data (Box 1).
Accounting for rate variation along the sequence
Evolutionary rates can vary across the genome, among
regions, among loci, and even between nucleotide sites. In
phylogenetic analyses of DNA sequence data, rate hetero-
geneity among sites has typically been taken into account
by assigning sites to a small number of discrete rate
categories, based on the gamma distribution [28]. This
method can also be used when analyzing genomic datasets,
but an alternative approach is to allow each locus or group
of loci to have a distinct evolutionary rate [29–31]. For
example, a relative rate parameter can be assigned to each
locus, with these rates following a gamma [30] or Dirichlet
distribution [31]. These methods, however, are only appro-
priate when gene and lineage effects are present, but not
residual effects (conforming to the Universal Pacemaker
model of genome evolution; Figure 1).
Accounting for residual effects
A more difficult problem emerges when there are signifi-
cant residual effects such that different sites or loci show
Trends in Ecology & Evolution xxx xxxx, Vol. xxx, No. x
Box 1. Calibrating the molecular clock
Molecular clock models describe the patterns of rate variation
across lineages, allowing estimation of the relative ages of nodes in
the phylogeny. To place an absolute timescale on the phylogenetic
tree, the molecular clock needs to be calibrated. This can be done in
one of two ways: by setting the rate to a known value, or by
constraining the age of at least one node in the phylogeny. The
scarcity of reliable rate estimates means that the latter approach is
usually preferred, especially when there is substantial rate variation
among lineages. Generally, increasing the number of calibrations
leads to an improvement in molecular clock estimation [25,66].
Ages of nodes can be constrained on the basis of fossil or
geological information. The earliest fossil representative of a
lineage can be used to infer the divergence time of that lineage
from its sister lineage [67]. The age constraint can be implemented
in several ways, of which the simplest is to fix the age of the node to
a point value. However, this ignores any uncertainty in the
calibration age, such as that associated with radiometric dating or
taxonomic assignment. Instead, a preferable approach is to account
for uncertainty by allowing the node age to vary within chosen
constraints [68]. In Bayesian phylogenetic analysis this can be done
by specifying an informative prior distribution for the node age,
typically in the form of a lognormal or exponential distribution [69].
Choosing the parameters of these distributions can be a difficult
exercise [70], but there are several formalized methods that allow
this process to be informed by the fossil data [71,72]. Alternatively,
calibrating fossils can be included in combined analyses of
morphological and molecular data such that their associated
temporal information is incorporated implicitly [63,73].
If ancient genomes are available, the ages of the sequences can be
used for calibration. This is possible in studies based on well-
preserved ancient specimens or on rapidly evolving viruses
sampled through time [74]. Calibrations based on ancient se-
quences can be very effective because there is usually no
uncertainty in the attachment of dates to nodes in the phylogeny.
Moreover, these dates are often known with considerable precision.
Any nontrivial uncertainty in the sequence ages can be incorporated
into the molecular clock analysis [75,76].
contrasting patterns of rate variation in different branches
of the tree. A familiar example of this is associated with the
codon structure of protein-coding genes: patterns of rate
variation at second codon positions are influenced by se-
lection, whereas rates at third codon sites are more likely
to be subject to lineage effects [32]. In this particular
example, a simple solution is to assign a separate model
of among-lineage rate variation to each codon position [33].
More generally, one can account for residual effects by
partitioning the data into subsets based on their pattern of
among-lineage rate heterogeneity. An appropriate strate-
gy might be to assign separate molecular clock models to
different genomic regions, different loci, or different codon
positions in protein-coding genes [33]. Alternatively, a
statistical approach can be taken to identify the optimal
partitioning scheme for the data. This can be done, for
example, by comparing the estimates of branch lengths
from the different loci in the dataset and using a clustering
method to group loci with similar patterns of rate variation
among branches [29,34]. These patterns can be summa-
rized using tree-distance metrics [34] or principal-compo-
nents analysis [29]. In the molecular clock analysis, a
separate model of among-lineage rate variation can then
be assigned to each subset of the data. This is analogous to
the common practice of partitioning the dataset and
assigning a separate substitution model to each data sub-
set [35].
3
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Opinion
Analyzing large datasets
With the drive to base evolutionary inferences on datasets
of increasing size, an ongoing feature of sampling has been
the trade-off between numbers of loci and taxa. Some
studies have focused on estimating evolutionary time-
scales for very large numbers of taxa, but these have
typically been restricted to small numbers of loci or have
used supertree methods to merge smaller trees inferred
from different markers [36,37]. The emergence of high-
throughput sequencing technology has made it possible to
assemble large datasets that can be characterized as hav-
ing (i) a small number of markers for a large number of
taxa; (ii) a large number of markers for a small number of
taxa; or (iii) a large number of markers for a large number
of taxa. As I explain below, these three types of dataset
present different challenges for molecular clock analyses.
Datasets comprising small numbers of markers for a
very large sample of taxa can be used to answer a variety of
evolutionary questions, particularly those associated with
macroevolutionary processes such as diversification
[36,37]. Several methods can be used to estimate evolu-
tionary timescales for very large phylogenetic trees [38–41]
(Box 2). These methods share several features in common.
All treat inference of the phylogeny as a separate problem,
and thus the topology and branch lengths (measuring the
amount of genetic change) are assumed to be known for the
dating analysis. Although this leads to a considerable
reduction in the computational burden, it also places a
limit on model complexity because the sequence data are
not always analyzed directly. In addition, most of the
methods that can analyze large numbers of taxa are unable
to accommodate complex calibrating information. Thus,
these methods typically do not handle uncertainty in cali-
brations and the phylogeny in an ideal manner.
In molecular dating analyses there has been growing
use of datasets that comprise large amounts of sequence
data from small numbers of taxa. As with many advances
Box 2. Methods for estimating evolutionary timescales from large datasets
A wide range of molecular clock methods for estimating evolu-
tionary timescales are available in various software packages. The
methods differ in terms of their statistical motivations, assumptions
about rate variation, and ability to handle different forms of
calibration. These characteristics typically need to be weighed
against the computational burden and running time of the different
methods.
Perhaps the most popular approaches, at least in recent years, are
those implemented in a Bayesian phylogenetic framework. Bayesian
methods are popular because they can readily incorporate complex
models of molecular evolution. Bayesian molecular clocks are
available in programs such as BEAST [77], MrBayes [78], MCMCtree
[79], PhyloBayes [80], and DPPdiv [20]. Some of these require a fixed
tree topology, whereas others are able to co-estimate the phylogeny
and node times. These programs are usually able to implement a
range of relaxed-clock models. Clock calibrations are typically
incorporated as priors on node ages. Estimates of node times can
be sensitive to the choice of priors, making it important to examine
the priors carefully [31,81].
Unlike most other Bayesian methods, MCMCtree has the option
of using approximate likelihood calculation [52]. This makes it
feasible for MCMCtree to analyze genome-scale datasets [29,82].
Despite such computational improvements, Bayesian molecular
4
Trends in Ecology & Evolution xxx xxxx, Vol. xxx, No. x
in molecular phylogenetics, the first steps in this direction
were taken using organellar genomes. Molecular dating
using complete sequences of organellar genomes is now
relatively common (e.g., [42,43]), whereas few dating stud-
ies based on nuclear DNA have taken advantage of the
data produced by early genome projects (e.g., [44]). When
there are few taxa in the dataset, the number of distinct
site patterns in the alignment is small. Sites sharing the
same pattern of variation across taxa can be grouped for
the purposes of likelihood calculation such that molecular
clock analyses of these datasets are computationally trac-
table even with intensive Bayesian and likelihood methods
(Box 2). However, datasets with many loci but few taxa are
subject to several disadvantages associated with sparse
taxon sampling, with impacts on tree balance, performance
of phylogenetic inference, and estimation of macroevolu-
tionary parameters [45]. The addition of taxa often comes
at the cost of an increased proportion of missing data, with
uncertain impacts on molecular clock analysis [46,47].
Analyses of multilocus data must also deal with incongru-
ence between trees estimated from different loci, the extent
of which will depend on the taxonomic scale being investi-
gated (Box 3).
Genome-scale datasets from moderate to large numbers
of taxa are still relatively rare. However, with various
genome-sequencing initiatives underway, such as the Ge-
nome 10K Project [48] and the i5K Project [49], very large
datasets will soon become much more common. Analyses of
these data, which have the potential to comprise millions of
characters from tens to hundreds of taxa, can be handled
using rapid likelihood-based methods but remain problem-
atic for most Bayesian phylogenetic methods. In particu-
lar, enormous computational demands are made by the
calculation of the full likelihood and by the estimation of
the posterior using Markov chain Monte Carlo sampling
[50]. These analyses will benefit from efforts to improve the
computational efficiency of molecular dating analyses. In
clock methods can usually only handle datasets comprising
moderate numbers of taxa. Further developments will allow
Bayesian molecular clock methods to be applied to much larger
datasets [50].
Several non-Bayesian methods have been developed for analyzing
datasets containing large numbers of taxa and are available in such
software as PATHd8 [41], treePL [40], DAMBE [39], chronos [66], and
RelTime [38]. These typically rely on other methods to produce
estimates of the tree topology and branch lengths. Genome-scale
datasets can be used for this purpose if rapid phylogenetic analysis is
performed by methods such as RAxML [83]. Most non-Bayesian
methods for molecular clock analysis employ a rate-smoothing
approach in which large rate changes between neighboring branches
in the phylogeny are disfavored [68]. Calibrations are implemented as
constraints on node times or by fixing the ages of nodes to point
values. Choosing a molecular clock method depends on a balance
between the features of the method and its computational demands.
With a sufficiently large number of calibrations, however, we would
expect most molecular clock methods to converge on similar
estimates of evolutionary timescales [25,38,66]. Nevertheless, the
various methods report uncertainty in different ways, depending on
how they are able to handle error in the calibrations and the
phylogeny.
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Opinion Trends in Ecology & Evolution xxx xxxx, Vol. xxx, No. x

Box 3. Taxonomic scales of analysis

Molecular clock studies usually deal with evolutionary patterns and
processes occurring across geological timeframes. However, many
studies are conducted at much lower taxonomic scales, with multiple
samples drawn from a single species or from each of several closely
related species. These datasets pose very different challenges for
molecular clock analysis, and for phylogenetic analysis more
generally, because recombination removes the expectation of con-
gruence among the trees estimated from different loci. Accordingly,
the evolutionary process is better described using the coalescent
framework than by birth–death models of speciation and extinction.
Genome-scale intraspecific datasets are becoming more common
(e.g., [84,85]), raising the question of how such data can be analyzed
using molecular clocks. Methods based on concatenation, which
assume that all loci share the same tree, would clearly be
inappropriate [86]. One approach is to obtain an estimate of the
timescale from each locus: this can easily be achieved even using
intensive Bayesian or likelihood-based dating methods. The age
estimates for nodes of interest, such as those that are present in a
majority of gene trees, can then be summarized across loci. However,
any approaches based on consensus of gene trees can be highly
misleading, even at higher taxonomic levels [87].
When the species in the dataset are represented by multiple
samples, species-tree methods can be used. These allow variation
across individual gene trees but assume that they are all embedded
in a single, underlying species tree (e.g., [88,89]). These methods
tend to be computationally intensive and cannot be readily applied
to genome-scale datasets. Moreover, species-tree methods are
generally not amenable to the incorporation of calibrating informa-
tion.
Among studies of recent evolutionary events, a common problem
is a lack of reliable and appropriate calibrations [90]. Fossil
calibrations are generally more useful for analyses of deeper
timescales, whereas biogeography-based calibrations usually in-
volve strong assumptions about gene flow – and are not always
available for the taxa being studied. The paucity of reliable
calibrations also exacerbates the problem of time-dependent biases
in rate estimation [91]. Purifying selection and other factors cause
evolutionary rates to appear higher over short timeframes, an
effect that is particularly pronounced over the time-depths involved
in intraspecific analyses. Failing to account for this problem can
result in highly misleading estimates of evolutionary timescales
[92,93].

150 100 50

Jurassic Cretaceous Paleogene Neogene

Monotremata

Marsupiala

Xenarthra

Afrotheria
Theria

Lagomorpha

Placentalia Rodena

Scandena

Primates

Eulipotyphla

Cetarodactyla

Chiroptera

Perissodactyla

Carnivora

TRENDS in Ecology & Evolution

Figure 2. Evolutionary timescale of ordinal diversification in mammals. Chronogram (dates in Ma) estimated in a Bayesian relaxed-clock analysis of 14 632 nuclear genes,
calibrated using 38 fossil-based constraints on the ages of nodes [29]. Light-blue bars at nodes represent 95% credibility intervals of divergence-time estimates. Triangles
denote clades represented by more than one species in the analysis. Even with genome-scale data, the age estimates for some nodes have considerable uncertainty. The
orange vertical line indicates the timing of the Cretaceous–Paleogene boundary. Most orders of placental mammals diversified in the Paleogene, but the basal divergences
in Placentalia occurred in the Late Cretaceous. The date estimates were robust to a range of factors including data partitioning and various model priors [29,31].

5
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Opinion
Box 4. Case study – phylogenomic estimate of the
mammalian evolutionary timescale
The early history of mammals, particularly the timing of their
diversification in relation to the extinction of the dinosaurs at the
Cretaceous–Paleogene boundary (65.5 million years ago), has been
the subject of considerable research [94]. Most molecular clock
analyses have placed the diversification of mammalian orders in the
Cretaceous (e.g., [36,95]). In a recent study based on genomic data,
dos Reis and colleagues [29] estimated the timescale of mammalian
diversification. They analyzed the DNA sequences of 14 632 genes
that were partitioned into 20 subsets on the basis of their relative
rate of evolution. This partitioning was carried out using the
distance between human and mouse for each gene.
The authors performed further analyses using a smaller set of 857
genes that were present in all 36 genomes. As with the larger
dataset, the authors partitioned the genes according to their relative
evolutionary rate. For the 857 gene dataset, however, dos Reis et al.
[29] also compared a partitioning scheme in which genes were
grouped according to their patterns of rate variation across
branches. This partitioning was performed using a clustering
approach based on the branch lengths estimated from each gene.
In both cases, the evolutionary timescale was estimated using a
Bayesian molecular clock analysis in the software MCMCTREE [79],
calibrated by 38 fossil-based age constraints. To make the analysis
computationally tractable, the authors used approximate likelihood
calculation. Even with this approach, the analysis of the full dataset
took about 20 days.
The results of the analysis showed that the ordinal crown groups of
mammals originated after the Cretaceous–Paleogene boundary (see
Figure 2 in main text). The basal divergences among these orders,
however, largely occurred in the Late Cretaceous. This supports a
scenario in which placental mammals had a long evolutionary fuse in
the Cretaceous before radiating explosively in the Paleogene [96]. The
estimate of the evolutionary timescale was robust to a range of
factors, including the data-partitioning strategy. This attested to the
benefits of using genome-scale data in this study. Nevertheless, the
age-estimates for some of the divergence events in the tree had
considerable uncertainty, showing that even genomic data do not
necessarily yield errorless estimates of evolutionary parameters.
this regard, an effective technique is the use of approxi-
mate likelihood calculations in Bayesian relaxed-clock
methods, which can lead to a 1000-fold reduction in com-
puting time [50–52]. A recent example of the application of
these methods involved the timing of the early diversifica-
tion of placental mammals (Figure 2; Box 4). Substantial
reductions in the time needed for molecular clock analyses
can also be achieved by distributing computational load,
such as by multithreading or parallelization [53,54].
Looking towards the future
There has been a rapid growth in genomic data over the
past few years, but are there diminishing returns when
using larger datasets for molecular clock analysis? Even
with infinite data, molecular dating analyses cannot pro-
duce errorless estimates of evolutionary timescales [55,56].
Assuming that the phylogenetic tree has been estimated
accurately and that an appropriate substitution model has
been chosen, uncertainty in estimates of node ages arises
from three sources [39,57]. First, there is estimation error
in the branch lengths. Second, there is uncertainty in the
pattern of rate variation among branches in the phylogeny.
Third, uncertainty in the fossil or geological information
used to construct calibrations is rarely trivial.
In analyses of genomic data, the branch lengths are
effectively estimated without error, and uncertainties in
6
Trends in Ecology & Evolution xxx xxxx, Vol. xxx, No. x
estimates of node times therefore depend on the clock
model and on the calibrations [39]. Assuming that the
chosen clock model accurately describes the rate variation
across the tree, uncertainty in the estimates of node times
converges to the uncertainty in the calibrations; this can
occur even with relatively small amounts of sequence data
[57,58]. In particular, estimation error in node ages is most
strongly influenced by the most precisely defined calibra-
tions [57]. Therefore, without reliable calibrations, the
advantages of using genomic data are largely blunted.
As a consequence, identification of reliable calibrations
remains the most crucial component of molecular dating
analyses [59].
The growing wealth of genomic data presents an excit-
ing opportunity for improving models of the molecular
clock. Comprehensive comparisons of different relaxed-
clock models will help to identify the models that provide
the best description of biological reality. Furthermore,
improvements in our understanding of the causes of rate
variation among lineages and among loci bring the pros-
pect of building mechanistic models of molecular rates.
This will be aided by studies of biological correlates of rates
[60]. In this regard, models of covariation between traits
and rates present a promising line of inquiry [61]. Although
this review has focused on sequence data, future model
development might be able to accommodate other forms of
genome-scale data, such as ultra-conserved elements and
panels of single-nucleotide polymorphisms. For example, a
clock-based analysis of protein folds has been used to
investigate the early evolutionary history of life on Earth
[62].
Refinement of calibration methods, discoveries of infor-
mative fossils, and improved understanding of biogeo-
graphic calibrations will all play important roles in
strengthening the reliability of molecular clock estimates.
An exciting area of work is the development of methods for
incorporating fossil data into molecular clock analyses.
These can either involve combined analyses of morpholog-
ical and molecular data [63] or other methods of using
fossil-occurrence data for calibration [64,65].
Concluding remarks and future directions
The molecular clock has made important contributions to
our understanding of biological processes and evolutionary
timescales. Now in the genomic era, we have experienced a
deluge of data that can be used to gain fascinating insights
into evolutionary questions. To use the molecular clock to
its full potential, methodological and computational devel-
opment must keep pace.
Addressing computational challenges should not merely
depend on technological progress but should involve the
improvement of methods of phylogenetic and molecular
clock estimation. Further development of models of rate
variation and methods of clock calibration will help to
improve the reliability of molecular clock inferences. With
data availability no longer being a major limiting factor,
efforts should also be directed towards identifying the
genomic data subsets that are the most suitable for esti-
mating evolutionary timescales. With these improvements
in hand, the Time-Tree of Life will have its roots in firm
ground.
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Opinion
Acknowledgments
The author thanks Sebastián Duchêne for useful discussions and Paul
Craze, Tracy Heath, and anonymous reviewers for their constructive
comments on the manuscript. S.Y.W.H. was supported by the Australian
Research Council.
Appendix A. Supplementary data
Supplementary data associated with this article can be found, in the online
version, at http://dx.doi.org/10.1016/j.tree.2014.07.004.
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