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Downloads for data in this track are available:
Description
This track shows multiple alignments of 13 vertebrate
species and measurements of evolutionary conservation using
two methods (phastCons and phyloP) from the
PHAST package, for all species.
The multiple alignments were generated using multiz and
other tools in the UCSC/Penn State Bioinformatics
comparative genomics alignment pipeline.
Conserved elements identified by phastCons are also displayed in
this track.
PhastCons is a hidden Markov model-based method that estimates the probability that each
nucleotide belongs to a conserved element, based on the multiple alignment.
It considers not just each individual alignment column, but also its
flanking columns. By contrast, phyloP separately measures conservation at
individual columns, ignoring the effects of their neighbors. As a
consequence, the phyloP plots have a less smooth appearance than the
phastCons plots, with more "texture" at individual sites. The two methods
have different strengths and weaknesses. PhastCons is sensitive to "runs"
of conserved sites, and is therefore effective for picking out conserved
elements. PhyloP, on the other hand, is more appropriate for evaluating
signatures of selection at particular nucleotides or classes of nucleotides
(e.g., third codon positions, or first positions of miRNA target sites).
Another important difference is that phyloP can measure acceleration
(faster evolution than expected under neutral drift) as well as
conservation (slower than expected evolution). In the phyloP plots, sites
predicted to be conserved are assigned positive scores (and shown in blue),
while sites predicted to be fast-evolving are assigned negative scores (and
shown in red). The absolute values of the scores represent -log p-values
under a null hypothesis of neutral evolution. The phastCons scores, by
contrast, represent probabilities of negative selection and range between 0
and 1.
Both phastCons and phyloP treat alignment gaps and unaligned nucleotides as
missing data, and both were run with the same parameters.
UCSC has repeatmasked and aligned all genome assemblies, and
provides all the sequences for download. For genome assemblies
not available in the genome browser, there are alternative assembly hub
genome browsers. Missing sequence in any assembly
is highlighted in the track display by regions of yellow when
zoomed out and by Ns when displayed at base level (see Gap Annotation, below).
Table 1. Genome assemblies included in the 13-way Conservation track.
Display Conventions and Configuration
In full and pack display modes, conservation scores are displayed as a
wiggle track (histogram) in which the height reflects the
size of the score.
The conservation wiggles can be configured in a variety of ways to
highlight different aspects of the displayed information.
Click the Graph configuration help link for an explanation
of the configuration options.
Pairwise alignments of each species to the rat genome are
displayed below the conservation histogram as a grayscale density plot (in
pack mode) or as a wiggle (in full mode) that indicates alignment quality.
In dense display mode, conservation is shown in grayscale using
darker values to indicate higher levels of overall conservation
as scored by phastCons.
Checkboxes on the track configuration page allow selection of the
species to include in the pairwise display.
Note that excluding species from the pairwise display does not alter the
the conservation score display.
To view detailed information about the alignments at a specific
position, zoom the display in to 30,000 or fewer bases, then click on
the alignment.
Gap Annotation
The Display chains between alignments configuration option
enables display of gaps between alignment blocks in the pairwise alignments in
a manner similar to the Chain track display. The following
conventions are used:
- Single line: No bases in the aligned species. Possibly due to a
lineage-specific insertion between the aligned blocks in the rat genome
or a lineage-specific deletion between the aligned blocks in the aligning
species.
- Double line: Aligning species has one or more unalignable bases in
the gap region. Possibly due to excessive evolutionary distance between
species or independent indels in the region between the aligned blocks in both
species.
- Pale yellow coloring: Aligning species has Ns in the gap region.
Reflects uncertainty in the relationship between the DNA of both species, due
to lack of sequence in relevant portions of the aligning species.
Genomic Breaks
Discontinuities in the genomic context (chromosome, scaffold or region) of the
aligned DNA in the aligning species are shown as follows:
-
Vertical blue bar: Represents a discontinuity that persists indefinitely
on either side, e.g. a large region of DNA on either side of the bar
comes from a different chromosome in the aligned species due to a large scale
rearrangement.
-
Green square brackets: Enclose shorter alignments consisting of DNA from
one genomic context in the aligned species nested inside a larger chain of
alignments from a different genomic context. The alignment within the
brackets may represent a short misalignment, a lineage-specific insertion of a
transposon in the rat genome that aligns to a paralogous copy somewhere
else in the aligned species, or other similar occurrence.
Base Level
When zoomed-in to the base-level display, the track shows the base
composition of each alignment. The numbers and symbols on the Gaps
line indicate the lengths of gaps in the rat sequence at those
alignment positions relative to the longest non-rat sequence.
If there is sufficient space in the display, the size of the gap is shown.
If the space is insufficient and the gap size is a multiple of 3, a
"*" is displayed; other gap sizes are indicated by "+".
Codon translation is available in base-level display mode if the
displayed region is identified as a coding segment. To display this annotation,
select the species for translation from the pull-down menu in the Codon
Translation configuration section at the top of the page. Then, select one of
the following modes:
-
No codon translation: The gene annotation is not used; the bases are
displayed without translation.
-
Use default species reading frames for translation: The annotations from
the genome displayed in the Default species to establish reading frame
pull-down menu are used to translate all the aligned species present in the
alignment.
-
Use reading frames for species if available, otherwise no translation:
Codon translation is performed only for those species where the region is
annotated as protein coding.
- Use reading frames for species if available, otherwise use default species:
Codon translation is done on those species that are annotated as being protein
coding over the aligned region using species-specific annotation; the remaining
species are translated using the default species annotation.
Codon translation uses the following gene tracks as the basis for translation:
| Gene Track | Species |
| UCSC Genes | Human, Mouse |
| RefSeq Genes | Cow, Rat, Rhesus, Zebrafish |
| Ensembl Genes v75 | Chicken, Chimp, Dog, Guinea Pig, Opossum, Panda, Turkey |
Table 2. Gene tracks used for codon translation.
Methods
Pairwise alignments with the rat genome were generated for
each species using lastz from repeat-masked genomic sequence.
Pairwise alignments were then linked into chains using a dynamic programming
algorithm that finds maximally scoring chains of gapless subsections
of the alignments organized in a kd-tree.
The scoring matrix and parameters for pairwise alignment and chaining
were tuned for each species based on phylogenetic distance from the reference.
High-scoring chains were then placed along the genome, with
gaps filled by lower-scoring chains, to produce an alignment net.
For more information about the chaining and netting process and
parameters for each species, see the description pages for the Chain and Net
tracks.
An additional filtering step was introduced in the generation of the 13-way
conservation track to reduce the number of paralogs and pseudogenes from the
high-quality assemblies and the suspect alignments from the low-quality
assemblies:
the pairwise alignments of high-quality
sequences were filtered based on synteny;
those for 2X mammalian genomes were filtered to retain only
alignments of best quality in both the target and query ("reciprocal
best").
The resulting best-in-genome pairwise alignments
were progressively aligned using multiz/autoMZ,
following the tree topology diagrammed above, to produce multiple alignments.
The multiple alignments were post-processed to
add annotations indicating alignment gaps, genomic breaks,
and base quality of the component sequences.
The annotated multiple alignments, in MAF format, are available for
bulk download.
An alignment summary table containing an entry for each
alignment block in each species was generated to improve
track display performance at large scales.
Framing tables were constructed to enable
visualization of codons in the multiple alignment display.
Phylogenetic Tree Model
Both phastCons and phyloP are phylogenetic methods that rely
on a tree model containing the tree topology, branch lengths representing
evolutionary distance at neutrally evolving sites, the background distribution
of nucleotides, and a substitution rate matrix.
The
all-species tree model for this track was
generated using the phyloFit program from the PHAST package
(REV model, EM algorithm, medium precision) using multiple alignments of
4-fold degenerate sites extracted from the 100-way alignment
(msa_view). The 4d sites were derived from the RefSeq (Reviewed+Coding) gene
set, filtered to select single-coverage long transcripts.
This same tree model was used in the phyloP calculations; however, the
background frequencies were modified to maintain reversibility.
The resulting tree model:
all species.
PhastCons Conservation
The phastCons program computes conservation scores based on a phylo-HMM, a
type of probabilistic model that describes both the process of DNA
substitution at each site in a genome and the way this process changes from
one site to the next (Felsenstein and Churchill 1996, Yang 1995, Siepel and
Haussler 2005). PhastCons uses a two-state phylo-HMM, with a state for
conserved regions and a state for non-conserved regions. The value plotted
at each site is the posterior probability that the corresponding alignment
column was "generated" by the conserved state of the phylo-HMM. These
scores reflect the phylogeny (including branch lengths) of the species in
question, a continuous-time Markov model of the nucleotide substitution
process, and a tendency for conservation levels to be autocorrelated along
the genome (i.e., to be similar at adjacent sites). The general reversible
(REV) substitution model was used. Unlike many conservation-scoring programs,
phastCons does not rely on a sliding window
of fixed size; therefore, short highly-conserved regions and long moderately
conserved regions can both obtain high scores.
More information about
phastCons can be found in Siepel et al. 2005.
The phastCons parameters used were: expected-length=45,
target-coverage=0.3, rho=0.3.
PhyloP Conservation
The phyloP program supports several different methods for computing
p-values of conservation or acceleration, for individual nucleotides or
larger elements (
http://compgen.cshl.edu/phast/). Here it was used
to produce separate scores at each base (--wig-scores option), considering
all branches of the phylogeny rather than a particular subtree or lineage
(i.e., the --subtree option was not used). The scores were computed by
performing a likelihood ratio test at each alignment column (--method LRT),
and scores for both conservation and acceleration were produced (--mode
CONACC).
Conserved Elements
The conserved elements were predicted by running phastCons with the
--most-conserved option. The predicted elements are segments of the alignment
that are likely to have been "generated" by the conserved state of the
phylo-HMM. Each element is assigned a log-odds score equal to its log
probability under the conserved model minus its log probability under the
non-conserved model. The "score" field associated with this track contains
transformed log-odds scores, taking values between 0 and 1000. (The scores
are transformed using a monotonic function of the form a * log(x) + b.) The
raw log odds scores are retained in the "name" field and can be seen on the
details page or in the browser when the track's display mode is set to
"pack" or "full".
Credits
This track was created using the following programs:
- Alignment tools: lastz (formerly blastz) and multiz by Minmei Hou, Scott Schwartz and Webb
Miller of the Penn State Bioinformatics Group
- Chaining and Netting: axtChain, chainNet by Jim Kent at UCSC
- Conservation scoring: phastCons, phyloP, phyloFit, tree_doctor, msa_view and
other programs in PHAST by
Adam Siepel at Cold Spring Harbor Laboratory (original development
done at the Haussler lab at UCSC).
- MAF Annotation tools: mafAddIRows by Brian Raney, UCSC; mafAddQRows
by Richard Burhans, Penn State; genePredToMafFrames by Mark Diekhans, UCSC
- Tree image generator: phyloPng by Galt Barber, UCSC
- Conservation track display: Kate Rosenbloom, Hiram Clawson (wiggle
display), and Brian Raney (gap annotation and codon framing) at UCSC
The phylogenetic tree is based on Murphy et al. (2001) and general
consensus in the vertebrate phylogeny community. Thanks to Giacomo Bernardi for
help with the fish relationships.
References
Phylo-HMMs, phastCons, and phyloP:
Felsenstein J, Churchill GA.
A Hidden Markov Model approach to
variation among sites in rate of evolution.
Mol Biol Evol. 1996 Jan;13(1):93-104.
PMID: 8583911
Pollard KS, Hubisz MJ, Rosenbloom KR, Siepel A.
Detection of nonneutral substitution rates on mammalian phylogenies.
Genome Res. 2010 Jan;20(1):110-21.
PMID: 19858363; PMC: PMC2798823
Siepel A, Bejerano G, Pedersen JS, Hinrichs AS, Hou M, Rosenbloom K,
Clawson H, Spieth J, Hillier LW, Richards S, et al.
Evolutionarily conserved elements in vertebrate, insect, worm,
and yeast genomes.
Genome Res. 2005 Aug;15(8):1034-50.
PMID: 16024819; PMC: PMC1182216
Siepel A, Haussler D.
Phylogenetic Hidden Markov Models.
In: Nielsen R, editor. Statistical Methods in Molecular Evolution.
New York: Springer; 2005. pp. 325-351.
Yang Z.
A space-time process model for the evolution of DNA
sequences.
Genetics. 1995 Feb;139(2):993-1005.
PMID: 7713447; PMC: PMC1206396
Chain/Net:
Kent WJ, Baertsch R, Hinrichs A, Miller W, Haussler D.
Evolution's cauldron:
duplication, deletion, and rearrangement in the mouse and human genomes.
Proc Natl Acad Sci U S A. 2003 Sep 30;100(20):11484-9.
PMID: 14500911; PMC: PMC208784
Multiz:
Blanchette M, Kent WJ, Riemer C, Elnitski L, Smit AF, Roskin KM,
Baertsch R, Rosenbloom K, Clawson H, Green ED, et al.
Aligning multiple genomic sequences with the threaded blockset aligner.
Genome Res. 2004 Apr;14(4):708-15.
PMID: 15060014; PMC: PMC383317
Lastz (formerly Blastz):
Chiaromonte F, Yap VB, Miller W.
Scoring pairwise genomic sequence alignments.
Pac Symp Biocomput. 2002:115-26.
PMID: 11928468
Harris RS.
Improved pairwise alignment of genomic DNA.
Ph.D. Thesis. Pennsylvania State University, USA. 2007.
Schwartz S, Kent WJ, Smit A, Zhang Z, Baertsch R, Hardison RC,
Haussler D, Miller W.
Human-mouse alignments with BLASTZ.
Genome Res. 2003 Jan;13(1):103-7.
PMID: 12529312; PMC: PMC430961
Phylogenetic Tree:
Murphy WJ, Eizirik E, O'Brien SJ, Madsen O, Scally M, Douady CJ, Teeling E,
Ryder OA, Stanhope MJ, de Jong WW, Springer MS.
Resolution of the early placental mammal radiation using Bayesian phylogenetics.
Science. 2001 Dec 14;294(5550):2348-51.
PMID: 11743200
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