GMD Vignette:

Generic histogram construction, generic distance measure,

cluster analysis with evaluation and visualization

Xiaobei Zhao∗ Albin Sandelin∗

February 6, 2012

(Version: 0.3.1.1)
Abstract

The purpose of this GMD Vignette is to show how to get started with the R package GMD. GMD denotes Generalized
Minimum Distance between distributions, which is a true metric that measures the distance between the
shapes of any two discrete numerical distributions (e.g. histograms).
The vignette includes a brief introduction, an example to illustrate the concepts and the implementation of GMD
and case studies that were carried out using classical data sets (e.g. iris) and high-throughput sequencing data
(e.g. ChIP-seq) from biology experiments. The appendix on page 15 contains an overview of package functionality,
and examples using primary functions in histogram construction (the ghist function), how to measure distance
between distributions (the gdist function), cluster analysis with evaluation (the ‘‘elbow’’ method in the css
function) and visualization (the heatmap.3 function).

Keywords: histogram, distance, metric, non-parametric, cluster analysis, hierarchical clustering, sum-of-squares,
heatmap.3

Contents

1 Introduction 2

2 Minimal Example: “Hello, GMD!” 3

3 An example to understand GMD 4

3.1 Histogram: construction and visualization . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

3.2 Histogram: distance measure and alignment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

∗ Bioinformatics Centre, University of Copenhagen, Department of Biology and Biotech Research and Innovation Centre, Ole

Maaløes Vej 5, DK-2200, Copenhagen, Denmark. Email: xiaobei@binf.ku.dk ; albin@binf.ku.dk

1
4 Case study 9

4.1 CAGE: measuring the dissimilarities among TSSDs . . . . . . . . . . . . . . . . . . . . . . . . . 9

4.2 ChIP-seq: measuring the similarities among histone modification patterns . . . . . . . . . . . . 12

A Functionality 15

A.1 An overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

A.2 ghist: Generic construction and visualization of histograms . . . . . . . . . . . . . . . . . . . . 16

A.2.1 Examples using simulated data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 16

A.2.2 Examples using iris data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

A.2.3 Examples using nottem data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20

A.3 gdist: Generic construction and visualization of distances . . . . . . . . . . . . . . . . . . . . . 22

A.4 css: Clustering Sum-of-Squares and the ‘‘elbow’’ plot . . . . . . . . . . . . . . . . . . . . . . 24

A.5 heatmap.3: Visualization in cluster analysis, with evaluation . . . . . . . . . . . . . . . . . . . 27

A.5.1 Examples using mtcars data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 27

A.5.2 Examples using ruspini data . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 31

B Data 33

B.1 GMD dataset overview . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33

B.2 CAGE data: cage and cagel . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 33

B.3 ChIP-seq data: chipseq mES and chipseq hCD4T . . . . . . . . . . . . . . . . . . . . . . . . . . 34

1 Introduction

Similar to the Earth Mover’s distance, Generalized Minimum distance of Distributions (GMD) (based on
MDPA - Minimum Difference of Pair Assignment [3]) is a true metric of the similarity between the shapes of
two histograms1 . Considering two normalized histograms A and B, GMD measures their similarity by counting
the necessary “shifts” of elements between the bins that have to be performed to transform distribution A into
distribution B.

The GMD package provides classes and methods for computing GMD in R [5]. The algorithm has been implemented
in C to interface with R for efficient computation. The package also includes downstream cluster analysis in
function css (A.4 on page 24) that use a pairwise distance matrix to make partitions given variant criteria,
including the “elbow” rule as discussed in [7] or desired number of clusters. In addition, the function heatmap.3
(A.5 on page 27) integrates the visualization of the hierarchical clustering in dendrogram, the distance measure
1 In statistics (and many other fields), histogram refers to a graphical representation of category frequencies in the data. Here,

we use this term in a more mathematical sense, defined as a function that counts categorical data or a result returned by such a
function.

2
 in heatmap and graphical representations of summary statistics of the resulting clusters or the overall partition.
For more flexibility, the function heatmap.3 can also accept plug-in functions defined by end-users for custom
summary statistics.

The motivation to write this package was born with the project [7] on characterizing Transcription Start
Site (TSS) landscapes using high-throughput sequencing data, where a non-parametric distance measure was
developed to assess the similarity among distributions of high-throughput sequencing reads from biological
experiments. However, it is possible to use the method for any empirical distributions of categorical data.

The package is available on CRAN. The source code is available at http://cran.r-project.org/web/packages/

GMD/ under GPL license.

2 Minimal Example: “Hello, GMD!”

hello-GMD.R
1 ##' Check GMD's sanity and start up
2 ##' @name hello-GMD
3

4 ## GMD at CRAN, for source code download and installation

5 ## http://cran.r-project.org/web/packages/GMD/index.html
6

7 ## load GMD
8 library(GMD)
9

10 ## version of GMD and description

11 packageVersion("GMD")
12 packageDescription("GMD")
13

14 ## view GMD vignette

15 vignette("GMD-vignette",package="GMD")
16

17 ## list the available data sets in GMD

18 data(package="GMD")
19

20 ## list all the objects in the GMD

21 ls("package:GMD")
22

23 ## help info on GMD

24 help(package="GMD")
25

26 ## run a demo
27 demo("GMD-demo")
28

29 ## cite GMD in publications

30 citation(package="GMD")
31

Figure 1: Source code of “hello-GMD.R”.

hello-GMD.R (fig. 1) is a minimal example to load and check of that your GMD installation works. It also

3
includes code for viewing the package information and this “Vignette”, checking data sets provided by GMD,
starting a demo and listing the citation of GMD.

3 An example to understand GMD

This example, based on simulated data, is designed to illustrate the concepts and the implementation of GMD
by stepping through the computations in detail.

3.1 Histogram: construction and visualization

> ## ------------------------------------------------------------------------
> ## chunk1: Load library and simulate data
> ## ------------------------------------------------------------------------
> require("GMD") # load library
> ## create two normally-distributed samples
> ## with unequal means and unequal variances
> set.seed(2012)
> x1 <- rnorm(1000,mean=-5, sd=10)
> x2 <- rnorm(1000,mean=10, sd=5)

> ## ------------------------------------------------------------------------
> ## chunk2: Construct histograms
> ## ------------------------------------------------------------------------
> ## create common bins
> n <- 20 # desired number of bins
> breaks <- gbreaks(c(x1,x2),n) # bin boundaries
> ## make two histograms
> v1 <- ghist(x1,breaks=breaks,digits=0)
> v2 <- ghist(x2,breaks=breaks,digits=0)

> ## ------------------------------------------------------------------------
> ## chunk3: Save histograms as multiple-histogram (`mhist') object
> ## ------------------------------------------------------------------------
> x <- list(v1,v2)
> mhist.obj <- as.mhist(x)

4
> ## ------------------------------------------------------------------------
> ## chunk4: Visualize an `mhist' object
> ## ------------------------------------------------------------------------
> ## plot histograms side-by-side
> plot(mhist.obj,mar=c(1.5,1,1,0),main="Histograms of simulated normal distributions")

Histograms of simulated normal distributions

1
250 2

200

150
Count

100

50

−21 −5 11 27

Bin

5
> ## plot histograms as subplots, with corresponding bins aligned
> plot(mhist.obj,beside=FALSE,mar=c(1.5,1,1,0),
+ main="Histograms of simulated normal distributions")

Histograms of simulated normal distributions

250 1
200

150

100

50

0
Count

−21 −5 11 27

250 2
200

150

100

50

−21 −5 11 27

Bin

6
3.2 Histogram: distance measure and alignment

Here we measure the GMD distance between shapes of two histograms with option sliding on.
> ## ------------------------------------------------------------------------
> ## chunk5: Measure the pairwise distance between two histograms by GMD
> ## ------------------------------------------------------------------------
> gmdp.obj <- gmdp(v1,v2,sliding=TRUE)
> print(gmdp.obj) # print a brief version by default

[1] 1.334

> print(gmdp.obj,mode="detailed") # print a detailed version

GM-Distance: 1.334
Sliding: TRUE
Number of hits: 1
Gap:
v1 v2
Hit1 5 0

Resolution: 1

> print(gmdp.obj,mode="full") # print a full version

Distribution of v1:
1 2 10 19 28 46 59 101 109 119 133 108 90 77 42 29 17 6 3 1
(After normalization)
0.001 0.002 0.01 0.019 0.028 0.046 0.059 0.101 0.109 0.119 0.133 0.108 0.09 0.077 0.042 0.029 0.017 0.006 0.003 0.001
Distribution of v2:
0 0 0 0 0 0 0 0 0 1 5 30 94 206 258 199 139 58 8 2
(After normalization)
0 0 0 0 0 0 0 0 0 0.001 0.005 0.03 0.094 0.206 0.258 0.199 0.139 0.058 0.008 0.002
GM-Distance: 1.334
Sliding: TRUE
Number of hits: 1
Gap:
v1 v2
Hit1 5 0

Resolution: 1

Now, let’s have a look at the alignment by GMD, with a distance 1.334 and a “shift” of 5 in the 1st distribution.
It is important to note that the specific features (the values in this case) of the original bins in the histograms
are ignored with sliding on. To keep original bin-to-bin correspondence, please set sliding to FALSE (see
examples in section 4.2 on page 12).
> ## ------------------------------------------------------------------------
> ## chunk6: Show alignment
> ## ------------------------------------------------------------------------
> plot(gmdp.obj,beside=FALSE)

7
 Optimal alignment between distributions (with sliding)
GMD=1.334, gap=c(5,0)
0.25
v1
0.20

0.15

0.10

0.05

0.00

5 10 15 20 25
Fraction

0.25
v2
0.20

0.15

0.10

0.05

0.00

5 10 15 20 25

Position

8
 4 Case study

4.1 CAGE: measuring the dissimilarities among TSSDs

Studies have demonstrated that the spatial distributions of read-based sequencing data from different platforms
often indicate functional properties and expression profiles (reviewed in [6] and [8]). Analyzing the distributions
of DNA reads is therefore often meaningful. To do this systematically, a measure of similarity between
distributions is necessary. Such measures should ideally be true metrics, have few parameters as possible, be
computationally efficient and also make biological sense to end-users. Case studies were made in section 4.1
and 4.2 to demonstrate the applications of GMD using distributions of CAGE and ChIP-seq reads.

In this section we demonstrate how GMD is applied to measure the dissimilarities among TSSDs, histograms of
transcription start site (TSS) that are made of CAGE tags, with option sliding on. The spatial properties of
TSSDs vary widely between promoters and have biological implications in both regulation and function. The
raw data were produced by CAGE and downloaded from FANTOM3 ([2]) and CAGE sequence reads were
preprocessed as did in [7]. The following codes case-cage.R (fig. 2) are sufficient to perform both pairwise
GMD calculation by function gmdp and to construct a GMD distance matrix by function gmdm. A handful of
options are available for control and flexibility, particularly, the option sliding is enabled by default to allow
partial alignment.
case-cage.R
1 require("GMD") # load library
2 data(cage) # load data
3

4 ## measure pairwise distance

5 x <- gmdp(cage[["Pfkfb3 (T02R00AEC2D8)"]],cage[["Csf1 (T03R0672174D)"]])
6 print(x) # print a brief version by default
7 print(x, mode="full") # print a full version by default
8

9 ## show alignment
10 plot(x,labels=c("Pfkfb3","Csf1"),beside=FALSE)
11

12 ## show another alignment

13 plot(gmdp(cage[["Hig1 (T09R0743763C)"]],cage[["Cd72 (T04R028B8BC9)"]]),
14 labels=c("Hig1 (T09R0743763C)","Cd72 (T04R028B8BC9)"),
15 beside=FALSE)
16

17 ## construct a distance matrix and visualize it

18 short.labels <- gsub("(.+) \\(.+","\\1",names(cage)) # get short labels
19 x <- gmdm(cage[1:6],labels=short.labels[1:6])
20 plot(x)

Figure 2: Source code of “case-cage.R”.

9
 Optimal alignment between distributions (with sliding)
GMD=7.881, gap=c(0,100)
0.8

0.6 Pfkfb3 (T02R00AEC2D8)

0.4

0.2

0.0

50 100 150 200 250

Fraction

0.8

0.6 Csf1 (T03R0672174D)

0.4

0.2

0.0

50 100 150 200 250

Position

Figure 3: Graphical output 1 of source code “case-cage.R”.

Optimal alignment between distributions (with sliding)

GMD=3.992, gap=c(0,24)
0.3 Hig1 (T09R0743763C)
0.2

0.1

0.0

20 40 60 80 100 120
Fraction

0.3 Cd72 (T04R028B8BC9)

0.2

0.1

0.0

20 40 60 80 100 120

Position

Figure 4: Graphical output 2 of source code “case-cage.R”.

10
 Optimal alignments among distributions (with sliding)
0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
NA 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
5 5 5 5 2 50 200 50 50 50 50 20 50 50 200 50 10 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=1.310 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Glul
Gap=(4,0)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
5 5 2 5 50 200 50 50 50 50 20 50 50 200 20 10 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=1.128 GMD=0.821 Cyp4a14 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Gap=(3,0) Gap=(0,1)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
5 5 5 50 200 50 50 50 50 20 50 50 200 20 140 10 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=1.835 GMD=2.833 GMD=2.284 Stxbp4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Gap=(0,0) Gap=(0,4) Gap=(0,3)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
5 5 50 200 50 50 50 50 20 50 50 200 50 10 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=0.907 GMD=0.562 GMD=0.833 GMD=2.732 D230039L
0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Gap=(3,0) Gap=(0,1) Gap=(0,0) Gap=(3,0) 06Rik
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
2 50 200 50 50 50 50 20 50 50 200 20 140 10 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=1.251 GMD=0.351 GMD=0.589 GMD=2.551 GMD=0.658 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Gas5
Gap=(0,0) Gap=(0,4) Gap=(0,3) Gap=(0,0) Gap=(0,3)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
50 200 50 50 50 50 20 50 50 200 50 10 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8
0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=4.914 GMD=3.931 GMD=3.802 GMD=5.698 GMD=4.339 GMD=3.769 Tpt1 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Gap=(93,0) Gap=(89,0) Gap=(90,0) Gap=(93,0) Gap=(90,0) Gap=(93,0)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
50 50 50 50 50 200 50 200 50 50 50 200 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8

0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=3.875 GMD=2.948 GMD=2.754 GMD=4.821 GMD=3.354 GMD=2.760 GMD=1.480 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Pcna
Gap=(74,0) Gap=(70,0) Gap=(71,0) Gap=(74,0) Gap=(71,0) Gap=(74,0) Gap=(0,19)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
Fraction

0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0

20 20 50 50 50 50 50 50 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8 0.8

0.6 0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=4.435 GMD=4.587 GMD=4.344 GMD=5.415 GMD=4.432 GMD=4.447 GMD=2.315 GMD=2.325 0.4 0.4 0.4 0.4 0.4 0.4 0.4 0.4
D0H4S114
Gap=(57,0) Gap=(54,0) Gap=(54,0) Gap=(59,0) Gap=(54,0) Gap=(59,0) Gap=(0,34) Gap=(0,15)
0.2 0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0 0.0
20 50 50 50 50 20 50 100 300

0.8 0.8 0.8 0.8 0.8 0.8 0.8

0.6 0.6 0.6 0.6 0.6 0.6 0.6
GMD=5.015 GMD=5.015 GMD=4.426 GMD=5.915 GMD=4.872 GMD=4.784 GMD=2.037 GMD=2.277 GMD=2.147 0.4 0.4 0.4 0.4 0.4 0.4 0.4
Gsto1
Gap=(66,0) Gap=(63,0) Gap=(63,0) Gap=(68,0) Gap=(63,0) Gap=(67,0) Gap=(0,25) Gap=(0,6) Gap=(9,0)
0.2 0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0 0.0
50 50 50 50 50 50 100 300

0.8 0.8 0.8 0.8 0.8 0.8

0.6 0.6 0.6 0.6 0.6 0.6
GMD=5.300 GMD=6.336 GMD=5.936 GMD=4.677 GMD=6.155 GMD=6.161 GMD=8.677 GMD=7.739 GMD=6.507 GMD=8.409 0.4 0.4 0.4 0.4 0.4 0.4
Hsd11b1
Gap=(40,0) Gap=(37,0) Gap=(39,0) Gap=(42,0) Gap=(37,0) Gap=(42,0) Gap=(0,51) Gap=(0,32) Gap=(0,17) Gap=(0,26)
0.2 0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0 0.0
50 50 50 200 20 50 100 300

0.8 0.8 0.8 0.8 0.8

0.6 0.6 0.6 0.6 0.6
GMD=3.674 GMD=4.736 GMD=4.628 GMD=4.532 GMD=4.247 GMD=4.734 GMD=8.213 GMD=7.086 GMD=6.236 GMD=8.173 GMD=2.453 0.4 0.4 0.4 0.4 0.4
Csf1
Gap=(0,0) Gap=(0,4) Gap=(0,3) Gap=(1,0) Gap=(0,3) Gap=(0,0) Gap=(0,93) Gap=(0,74) Gap=(0,57) Gap=(0,66) Gap=(0,40)
0.2 0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0 0.0
50 50 200 50 20 100 300

0.8 0.8 0.8 0.8

0.6 0.6 0.6 0.6
GMD=7.010 GMD=8.086 GMD=7.811 GMD=6.336 GMD=7.863 GMD=8.016 GMD=10.799 GMD=9.839 GMD=8.790 GMD=10.539 GMD=2.682 GMD=3.747 0.4 0.4 0.4 0.4
B2m
Gap=(34,0) Gap=(30,0) Gap=(31,0) Gap=(36,0) Gap=(31,0) Gap=(34,0) Gap=(0,57) Gap=(0,39) Gap=(0,23) Gap=(0,32) Gap=(0,6) Gap=(34,0)
0.2 0.2 0.2 0.2
0.0 0.0 0.0 0.0
50 200 20 50 100 300

0.8 0.8 0.8

0.6 0.6 0.6
GMD=4.607 GMD=4.024 GMD=3.538 GMD=5.794 GMD=4.162 GMD=3.802 GMD=1.179 GMD=1.402 GMD=2.462 GMD=1.968 GMD=8.670 GMD=7.881 GMD=10.728 0.4 0.4 0.4
Pfkfb3
Gap=(100,0) Gap=(96,0) Gap=(97,0) Gap=(100,0) Gap=(97,0) Gap=(100,0) Gap=(7,0) Gap=(26,0) Gap=(41,0) Gap=(32,0) Gap=(58,0) Gap=(100,0) Gap=(66,0)
0.2 0.2 0.2
0.0 0.0 0.0
50 200 50 200 100 300

0.8 0.8
0.6 0.6
GMD=17.388 GMD=18.387 GMD=17.706 GMD=15.601 GMD=18.253 GMD=18.130 GMD=18.366 GMD=17.692 GMD=16.389 GMD=16.574 GMD=12.228 GMD=14.311 GMD=11.099 GMD=18.183 Hig1 0.4 0.4
Gap=(35,0) Gap=(31,0) Gap=(32,0) Gap=(35,0) Gap=(32,0) Gap=(35,0) Gap=(0,50) Gap=(0,35) Gap=(0,16) Gap=(0,25) Gap=(0,5) Gap=(35,0) Gap=(1,0) Gap=(0,57)
0.2 0.2
0.0 0.0
20 100 300

0.8
0.6
GMD=17.387 GMD=18.386 GMD=17.705 GMD=15.579 GMD=18.252 GMD=18.130 GMD=18.835 GMD=18.126 GMD=17.125 GMD=17.059 GMD=12.993 GMD=15.084 GMD=12.237 GMD=18.652 GMD=3.992 0.4
Cd72
Gap=(7,0) Gap=(3,0) Gap=(4,0) Gap=(7,0) Gap=(4,0) Gap=(7,0) Gap=(0,74) Gap=(0,58) Gap=(0,42) Gap=(0,51) Gap=(0,33) Gap=(5,0) Gap=(0,29) Gap=(0,81) Gap=(0,24)
0.2
0.0
100 300

GMD=48.292 GMD=49.290 GMD=48.610 GMD=46.521 GMD=49.157 GMD=49.034 GMD=45.360 GMD=46.391 GMD=44.807 GMD=44.467 GMD=43.082 GMD=44.996 GMD=41.294 GMD=45.380 GMD=32.443 GMD=34.026 Egln1
Gap=(137,0) Gap=(133,0) Gap=(134,0) Gap=(137,0) Gap=(134,0) Gap=(137,0) Gap=(44,0) Gap=(63,0) Gap=(80,0) Gap=(71,0) Gap=(97,0) Gap=(137,0) Gap=(103,0) Gap=(37,0) Gap=(98,0) Gap=(121,0)

Position

Figure 5: Graphical output 1 of source code “case-cage.R”.

11
4.2 ChIP-seq: measuring the similarities among histone modification patterns

In this section we demonstrate how GMD is applied to measure the dissimilarities between histone modifications
represented by ChIP-seq reads. Distinctive patterns of chromatin modifications around the TSS are associated
with transcription regulation and expression variation of genes. Comparing the chromatin modification profiles
(originally produced by [1] and [4], and preprocessed by [7]), the sliding option is disabled for fixed alignments
at the TSSs and the flanking regions. The GMD measure indicates how well profiles are co-related to each
other. In addition, the downstream cluster analysis is visualized with function heatmap.3 that use GMD
distance matrix to generate clustering dendrograms and make partitions given variant criteria, including the
‘‘elbow’’ rule (discussed in [7]) or desired number of clusters.

12
 case-chipseq.R
1 require("GMD") # load library
2 data(chipseq_mES) # load data
3 data(chipseq_hCD4T) # load data
4

5 ## pairwise distance and alignment based on GMD metric

6 plot(gmdm(chipseq_mES,sliding=FALSE))
7

8 ## clustering on spatial distributions of histone modifications

9 x <- gmdm(chipseq_hCD4T,sliding=FALSE,resolution=10)
10 heatmap.3(x,revC=TRUE)
11

12 ## Determine the number of clusters by "Elbow" criterion

13 main <- "Heatmap of ChIP-seq data (human CD4+ T cells)"
14 dist.obj <- gmdm2dist(x)
15 css.multi.obj <- css.hclust(dist.obj,hclust(dist.obj))
16 elbow.obj <- elbow.batch(css.multi.obj,ev.thres=0.90,inc.thres=0.05)
17 heatmap.3(dist.obj, main=main, revC=TRUE, kr=elbow.obj$k, kc=elbow.obj$k)
18

19 ## more strict threshold

20 elbow.obj <- elbow.batch(css.multi.obj,ev.thres=0.75,inc.thres=0.1)
21 heatmap.3(dist.obj, main=main, revC=TRUE, kr=elbow.obj$k, kc=elbow.obj$k)
22

23 ## side plots
24 normalizeVector <- function(v){v/sum(v)} # a function to normalize a vector
25 dev.new(width=12,height=8)
26

27 ## summary of row clusters

28 expr1 <- list(quote(op <- par(mar = par("mar")*2)),
29 quote(plot(mhist.summary(as.mhist(i.x)),if.plot.new=FALSE)),
30 quote(par(op))
31 )
32

33 ## summary of row clustering

34 expr2 <- list(quote(tmp.clusters <- cutree(hclust(dist.row),k=kr)),
35 quote(tmp.css <- css(dist.row,tmp.clusters)),
36 quote(print(tmp.css)),
37 quote(tmp.wev <- tmp.css$wss/tmp.css$tss),
38 quote(names(tmp.wev) <- as.character(unique(tmp.clusters))),
39 quote(tmp.wev <- tmp.wev[order(unique(tmp.clusters))]),
40 quote(barplot(tmp.wev,main="Cluster Explained Variance", xlab="Cluster",
41 ylab="EV",col="white",border="black",
42 ylim=c(0,max(tmp.wev)*1.1),cex.main=1)))
43 expr3 <- list(quote(op <- par(mar = par("mar")*2)),
44 quote(plot.elbow(css.multi.obj,elbow.obj,if.plot.new=FALSE)),
45 quote(par(op))
46 )
47

48 heatmap.3(dist.obj, main=main, cex.main=1.25, revC=TRUE, kr=elbow.obj$k, kc=elbow.obj$k,

49 keysize=1,mapsize=4.5,row.data=lapply(chipseq_hCD4T,normalizeVector),
50 plot.row.clusters=TRUE,plot.row.clusters.list=list(expr1),
51 plot.row.clustering=TRUE,plot.row.clustering.list=list(expr2,expr3))
52

Figure 6: Source code of “case-chipseq.R”. 13

 Optimal alignments among distributions (without sliding)

0.007 0.007 0.007 0.007 0.007

0.006 0.006 0.006 0.006 0.006
0.005 0.005 0.005 0.005 0.005
0.004 0.004 0.004 0.004 0.004
H3K27me3
0.003 0.003 0.003 0.003 0.003
0.002 0.002 0.002 0.002 0.002

0.001 0.001 0.001 0.001 0.001

0.000 0.000 0.000 0.000 0.000
200 400 600 800 1000 200 400 600 800 1000 200 400 600 800 1000 200 400 600 800 1000 200 400 600 800 1000

0.007 0.007 0.007 0.007

0.006 0.006 0.006 0.006

0.005 0.005 0.005 0.005
0.004 0.004 0.004 0.004
GMD=118.840
H3K36me3
Gap=(0,0) 0.003 0.003 0.003 0.003
0.002 0.002 0.002 0.002
0.001 0.001 0.001 0.001
0.000 0.000 0.000 0.000
200 400 600 800 1000 200 400 600 800 1000 200 400 600 800 1000 200 400 600 800 1000

0.007 0.007 0.007

0.006 0.006 0.006
0.005 0.005 0.005

0.004 0.004 0.004

GMD=37.253 GMD=82.910
H3K4me1
Gap=(0,0) Gap=(0,0) 0.003 0.003 0.003
0.002 0.002 0.002
0.001 0.001 0.001
Fraction

0.000 0.000 0.000

200 400 600 800 1000 200 400 600 800 1000 200 400 600 800 1000

0.007 0.007
0.006 0.006
0.005 0.005
0.004 0.004
GMD=56.380 GMD=140.037 GMD=77.365
H3K4me2
Gap=(0,0) Gap=(0,0) Gap=(0,0) 0.003 0.003
0.002 0.002
0.001 0.001
0.000 0.000
200 400 600 800 1000 200 400 600 800 1000

0.007
0.006
0.005
0.004
GMD=110.047 GMD=190.394 GMD=135.755 GMD=58.471
H3K4me3
Gap=(0,0) Gap=(0,0) Gap=(0,0) Gap=(0,0) 0.003

0.002
0.001
0.000
200 400 600 800 1000

GMD=46.266 GMD=108.302 GMD=33.494 GMD=99.835 GMD=156.171

H3K9me3
Gap=(0,0) Gap=(0,0) Gap=(0,0) Gap=(0,0) Gap=(0,0)

Position

Figure 7: Graphical output 1 of source code “case-chipseq.R”.

Color Key
150
Count

Heatmap of ChIP−seq data (human CD4+ T cells)

Row Row
0 50

Clusters Clustering
0 5 15
Value

2 3 1
H2AK9ac Row group 2 (n=17)
CTCF k=3 Elbow plot
0.0015

H4K5ac ● ● ● ● ● ● ● ● ●
● ● ●
H4K8ac ●
Explained Variance

●
●
●
H2BK20ac
0.8

●
H2BK12ac 0.77 ●
H2AZ
H3K4ac
0.0000
2

H3K36ac
H4K91ac
0.4

H2BK120ac
H3K4me3 1 116 264 412 560 708 856 ●

H3K18ac
H3K9ac
H3K27ac
0.0

●
H2BK5ac
H4K20me1 5 10 15 20
H3K79me2 Row group 3 (n=7) k
H3K79me3
0.0015

H3K36me3
3

H3K27me1
H3K79me1
H2BK5me1 Cluster Explained Variance
H3K4me2
0.12

H3K9me1
0.0000

H3K4me1
H3K23ac
H4K12ac 1 116 264 412 560 708 856
H4K16ac
H4K20me3
0.08

H3K14ac
H3K36me1
EV
1

H4R3me2
H3R2me1 Row group 1 (n=16)
H3R2me2
0.04

H2AK5ac
H3K27me2
0.004

H3K9me2
H3K9me3
H3K27me3
0.00
H2AK9ac
CTCF
H4K5ac
H4K8ac
H2BK20ac
H2BK12ac
H2AZ
H3K4ac
H3K36ac
H4K91ac
H2BK120ac
H3K4me3
H3K18ac
H3K9ac
H3K27ac
H2BK5ac
H4K20me1
H3K79me2
H3K79me3
H3K36me3
H3K27me1
H3K79me1
H2BK5me1
H3K4me2
H3K9me1
H3K4me1
H3K23ac
H4K12ac
H4K16ac
H4K20me3
H3K14ac
H3K36me1
H4R3me2
H3R2me1
H3R2me2
H2AK5ac
H3K27me2
H3K9me2
H3K9me3
H3K27me3

1 2 3
0.000

Cluster
1 116 264 412 560 708 856

Figure 8: Graphical output 1 of source code “case-chipseq.R”.

14
A Functionality

A.1 An overview

Table 1. Functions of the GMD R package

Function Description

ghist Generalized Histogram Computation and Visualization

gdist Generalized Distance Matrix Computation
css Computing Clustering Sum-of-Squares and
evaluating the clustering by the ‘‘elbow’’ method
heatmap.3 Enhanced Heatmap Representation with Dendrogram and Partition
gmdp Computation of GMD on a pair of histograms
gmdm Computation of GMD Matrix on a set of histograms

15
 A.2 ghist: Generic construction and visualization of histograms

A.2.1 Examples using simulated data

example-ghist.R (fig. 9) is an example on how to construct a histogram object from raw data and make a
visualization based on this.
example-ghist.R
1 ## load library
2 require("GMD")
3

4 ## create two normally-distributed samples

5 ## with unequal means and unequal variances
6 set.seed(2012)
7 v1 <- rnorm(1000,mean=-5, sd=10)
8 v2 <- rnorm(1000,mean=10, sd=5)
9

10 ## create common bins

11 n <- 20 # desired number of bins
12 breaks <- gbreaks(c(v1,v2),n) # bin boundaries
13 x <-
14 list(ghist(v1,breaks=breaks,digits=0),
15 ghist(v2,breaks=breaks,digits=0))
16 mhist.obj <- as.mhist(x)
17

18 ## plot histograms side-by-side

19 plot(mhist.obj,mar=c(1.5,1,1,0),main="Histograms of simulated normal distributions")
20

21 ## plot histograms as subplots,

22 ## with corresponding bins aligned
23 plot(mhist.obj,beside=FALSE,mar=c(1.5,1,1,0),
24 main="Histograms of simulated normal distributions")

Figure 9: Source code of “example-ghist.R”.

16
 Histograms of simulated normal distributions

250
1
2
200

150
Count

100

50

−21 −5 11 27

Bin

Figure 10: Graphical output 1 of source code “example-ghist.R”.

Histograms of simulated normal distributions

250 1
200

150

100

50

0
Count

−21 −5 11 27

250 2
200

150

100

50

−21 −5 11 27

Bin

Figure 11: Graphical output 1 of source code “example-ghist.R”.

17
 A.2.2 Examples using iris data

case-iris.R (fig. 12) is a study on how to obtain and visualize histograms, using Fisher’s iris data set.
case-iris.R
1 ## load library
2 require("GMD")
3

4 ## load data
5 data(iris)
6

7 ## create common bins

8 n <- 30 # the number of bins
9 breaks <- gbreaks(iris[,"Sepal.Length"],n) # the boundary of bins
10

11 ## create a list of histograms

12 Sepal.Length <-
13 list(setosa=ghist(iris[iris$Species=="setosa","Sepal.Length"],breaks=breaks),
14 versicolor=ghist(iris[iris$Species=="versicolor","Sepal.Length"],breaks=breaks),
15 virginica=ghist(iris[iris$Species=="virginica","Sepal.Length"],breaks=breaks)
16 )
17

18 ## convert to a `hist' object

19 x <- as.mhist(Sepal.Length)
20

21 ## get bin-wise summary statistics

22 summary(x)
23

24 ## visualize the histograms

25 plot(x,beside=FALSE,
26 main="Histogram of Sepal.Length of iris",xlab="Sepal.Length (cm)")

Figure 12: Source code of “case-iris.R”.

18
 Histogram of Sepal.Length of iris

8
setosa

4.9 5.5 6.1 6.7 7.3 7.9

8
versicolor

6
Count

4.9 5.5 6.1 6.7 7.3 7.9

8
virginica

4.9 5.5 6.1 6.7 7.3 7.9

Sepal.Length (cm)

Figure 13: Graphical output 1 of source code “case-iris.R”.

19
 A.2.3 Examples using nottem data

case-nottem.R (fig. 14) is a study on how to draw histograms side-by-side and to compute and visualize a
bin-wise summary plot, using air temperature data at Nottingham Castle.
case-nottem.R
1 ## load library
2 require("GMD")
3

4 ## load data
5 data(nottem)
6

7 class(nottem) # a time-series (ts) object

8 x <- ts2df(nottem) # convert ts to data.frame
9 mhist1 <- as.mhist(x[1:3,])
10

11 ## plot multiple discrete distributions side-by-side

12 plot(mhist1,xlab="Month",ylab="Degree Fahrenheit",
13 main="Air temperatures at Nottingham Castle")
14

15 ## make summary statistics for each bin

16 mhist2 <- as.mhist(x)
17 ms <- mhist.summary(mhist2)
18 print(ms)
19

20 ## plot bin-wise summary statistics with

21 ## confidence intervals over the bars
22 plot(ms, main="Mean air temperatures at Nottingham Castle (1920-1939)",
23 xlab="Month", ylab="Degree Fahrenheit")

Figure 14: Source code of “case-nottem.R”.

20
 Air temperatures at Nottingham Castle

70
1920
65 1921
1922
60
Degree Fahrenheit

55

50

45

40

35

Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec

Month

Figure 15: Graphical output 1 of source code “case-nottem.R”.

Mean air temperatures at Nottingham Castle (1920−1939)

60
50
Degree Fahrenheit
40
30
20
10
0

Jan Feb Mar Apr May Jun Jul Aug Sep Oct Nov Dec
Month

Figure 16: Graphical output 2 of source code “case-nottem.R”.

21
 A.3 gdist: Generic construction and visualization of distances

example-gdist.R (fig. 17) is an example on how to measure distances using a user-defined metric, such as
correlation distance and GMD.
example-gdist.R
1 ## load library
2 require("GMD")
3 require(cluster)
4

5 ## compute distance using Euclidean metric (default)

6 data(ruspini)
7 x <- gdist(ruspini)
8

9 ## see a dendrogram result by hierarchical clustering

10 dev.new(width=12, height=6)
11 plot(hclust(x),
12 main="Cluster Dendrogram of Ruspini data",
13 xlab="Observations")
14

15 ## convert to a distance matrix

16 m <- as.matrix(x)
17

18 ## convert from a distance matrix

19 d <- as.dist(m)
20 stopifnot(d == x)
21

22 ## Use correlations between variables "as distance"

23 data(USJudgeRatings)
24 dd <- gdist(x=USJudgeRatings,method="correlation.of.variables")
25 dev.new(width=12, height=6)
26 plot(hclust(dd),
27 main="Cluster Dendrogram of USJudgeRatings data",
28 xlab="Variables")

Figure 17: Source code of “example-gdist.R”.

22
 Cluster Dendrogram of Ruspini data
150
100
Height

50

7
0

61
41
44
45

58

20
46

63

1
5
6
65

17
31

40

2
3
53

11

4
8
72
75
73
74
57
21
22

47
48
54
42
43

14
15
62
66

16

12
13

9
10
37
38

64
68

70
71
25
26

50
52
36
39

32
35
29
30

59
60

67
69
23
24

33
34

49
51
27
28

55
56

18
19
Observations
hclust (*, "complete")

Figure 18: Graphical output 1 of source code “example-gdist.R”.

Cluster Dendrogram of USJudgeRatings data

1.2
1.0

CONT
0.8
0.6
Height

0.4
0.2
0.0

PHYS

RTEN
DILG
INTG

DMNR

CFMG

DECI

PREP

FAMI
ORAL

WRIT

Variables
hclust (*, "complete")

Figure 19: Graphical output 2 of source code “example-gdist.R”.

23
A.4 css: Clustering Sum-of-Squares and the ‘‘elbow’’ plot: determining the
number of clusters in a data set

A good clustering yields clusters where the total within-cluster sum-of-squares (WSSs) is small (i.e. cluster
cohesion, measuring how closely related are objects in a cluster) and the total between-cluster sum-of-squares
(BSSs) is high (i.e. cluster separation, measuring how distinct or well-separated one cluster is from the other).

example-css.R (fig. 20) is an example on how to make correct choice of k using “elbow criterion”. A good
k is selected according a) how much of the total variance in the whole data that the clusters can explain, and
b) how large gain in explained variance we obtain by using these many clusters compared to one less or one
more, the so-called “elbow” criterion.

The optimal choice of k will strike a balance between maximum compression of the data using a single cluster,
and maximum accuracy by assigning each data point to its own cluster. More important, an ideal k should
also be relevant in terms of what it reveals about the data, which typically cannot be measured by a metric
but by a human expert. Here we present a way to measure such performance of a clustering model, using
squared Euclidean distances. The evaluation is based on pairwise distance matrix and therefore more generic
in a way that doesn’t involve computating the “centers” of the clusters in the raw data, which are often not
available or hard to obtain.

24
 example-css.R
1 ## load library
2 require("GMD")
3

4 ## simulate data around 12 points in Euclidean space

5 pointv <- data.frame(x=c(1,2,2,4,4,5,5,6,7,8,9,9),y=c(1,2,8,2,4,4,5,9,9,8,1,9))
6 set.seed(2012)
7 mydata <- c()
8 for (i in 1:nrow(pointv)){
9 mydata <- rbind(mydata,cbind(rnorm(10,pointv[i,1],0.1),rnorm(10,pointv[i,2],0.1)))
10 }
11 mydata <- data.frame(mydata); colnames(mydata) <- c("x","y")
12 plot(mydata,type="p",pch=21, main="Simulated data")
13

14 ## determine a "good" k using elbow

15 dist.obj <- dist(mydata[,1:2])
16 hclust.obj <- hclust(dist.obj)
17 css.obj <- css.hclust(dist.obj,hclust.obj)
18 elbow.obj <- elbow.batch(css.obj)
19 print(elbow.obj)
20

21 ## make partition given the "good" k

22 k <- elbow.obj$k; cutree.obj <- cutree(hclust.obj,k=k)
23 mydata$cluster <- cutree.obj
24

25 ## draw a elbow plot and label the data

26 dev.new(width=12, height=6)
27 par(mfcol=c(1,2),mar=c(4,5,3,3),omi=c(0.75,0,0,0))
28 plot(mydata$x,mydata$y,pch=as.character(mydata$cluster),col=mydata$cluster,cex=0.75,
29 main="Clusters of simulated data")
30 plot.elbow(css.obj,elbow.obj,if.plot.new=FALSE)
31

32 ## clustering with more relaxed thresholds (, resulting a smaller "good" k)

33 elbow.obj2 <- elbow.batch(css.obj,ev.thres=0.90,inc.thres=0.05)
34 mydata$cluster2 <- cutree(hclust.obj,k=elbow.obj2$k)
35

36 dev.new(width=12, height=6)
37 par(mfcol=c(1,2), mar=c(4,5,3,3),omi=c(0.75,0,0,0))
38 plot(mydata$x,mydata$y,pch=as.character(mydata$cluster2),col=mydata$cluster2,cex=0.75,
39 main="Clusters of simulated data")
40 plot.elbow(css.obj,elbow.obj2,if.plot.new=FALSE)

Figure 20: Source code of “example-css.R”.

25
 Clusters of simulated data Elbow plot
k=7

1.0
5 55 55555 6 ● ● ● ● ● ● ● ● ● ● ● ●
55555
5 555
55
6
6666
66
66 ●
● ●
0.98
●

22 2 666
222 66 6
8
●
22
22 666

0.8
Explained Variance
6

0.6
●
mydata$y

4444
4444
4
●

0.4
4444 4
44444
4

4444
444

0.2
1 3
11
1 11111 3333
2

33
1
11111 7
7777777
111

0.0
1 7 ●

2 4 6 8 5 10 15 20

mydata$x k
A "good" k=7 (EV=0.98) is detected when the EV is no less than 0.95
and the increment of EV is no more than 0.01 for a bigger k.

Figure 21: Graphical output 1 of source code “example-css.R”.

Clusters of simulated data Elbow plot

k=5
1.0

4 44 44444 4 4 ● ● ● ● ● ● ● ● ● ● ● ●
44444 444 4444
44
44 ●
● ●
4 44 ●
0.94
22 2 444
222 44 4
8

●
22
22 444
0.8
Explained Variance
6

0.6

●
mydata$y

3333
3333
3
●
0.4

3333 3
3333 33333
4

333
0.2

1 1
11
1 11111 1111
2

11
1
11111 5
5555555
111
0.0

1 5 ●

2 4 6 8 5 10 15 20

mydata$x k
A "good" k=5 (EV=0.94) is detected when the EV is no less than 0.9
and the increment of EV is no more than 0.05 for a bigger k.

Figure 22: Graphical output 2 of source code “example-css.R”.

26
A.5 heatmap.3: Visualization in cluster analysis, with evaluation

A.5.1 Examples using mtcars data

example-heatmap3a.R (fig. 23) is an example on how to make a heatmap with summary visualization of
observations.

27
 example-heatmap3a.R
1 require("GMD") # load library
2 data(mtcars) # load data
3 x <- as.matrix(mtcars) # data as a matrix
4

5 dev.new(width=10,height=8)
6 heatmap.3(x) # default, with reordering and dendrogram
7 heatmap.3(x, Rowv=FALSE, Colv=FALSE) # no reordering and no dendrogram
8 heatmap.3(x, dendrogram="none") # reordering without dendrogram
9 heatmap.3(x, dendrogram="row") # row dendrogram with row (and col) reordering
10 heatmap.3(x, dendrogram="row", Colv=FALSE) # row dendrogram with only row reordering
11 heatmap.3(x, dendrogram="col") # col dendrogram
12 heatmap.3(x, dendrogram="col", Rowv=FALSE) # col dendrogram with only col reordering
13 heatmapOut <-
14 heatmap.3(x, scale="column") # sacled by column
15 names(heatmapOut) # view the list that is returned
16 heatmap.3(x, scale="column", x.center=0) # colors centered around 0
17 heatmap.3(x, scale="column",trace="column")# trun "trace" on
18

19 ## coloring cars (row observations) by brand

20 brands <- sapply(rownames(x), function(e) strsplit(e," ")[[1]][1])
21 names(brands) <- c()
22 brands.index <- as.numeric(as.factor(brands))
23 RowIndividualColors <- rainbow(max(brands.index))[brands.index]
24 heatmap.3(x, scale="column", RowIndividualColors=RowIndividualColors)
25

26 ## coloring attributes (column features) randomly (just for a test :)

27 heatmap.3(x, scale="column", ColIndividualColors=rainbow(ncol(x)))
28

29 ## add a single plot for all row individuals

30 dev.new(width=12,height=8)
31 expr1 <- list(quote(plot(row.data[rowInd,"hp"],1:nrow(row.data),xlab="hp",ylab="",
32 main="Gross horsepower",yaxt="n")),
33 quote(axis(2,1:nrow(row.data),rownames(row.data)[rowInd],las=2)))
34 heatmap.3(x, scale="column", plot.row.individuals=TRUE, row.data=x,
35 plot.row.individuals.list=list(expr1))
36

37 ## add a single plot for all col individuals

38 dev.new(width=12,height=8)
39 expr2 <- list(quote(plot(colMeans(col.data)[colInd], xlab="", ylab="Mean",xaxt="n",
40 main="Mean features",cex=1,pch=19)),
41 quote(axis(1,1:ncol(col.data),colnames(col.data)[colInd],las=2)))
42 heatmap.3(x, scale="column", plot.col.individuals=TRUE, col.data=x,
43 plot.col.individuals.list=list(expr2))
44

45 ## add another single plot for all col individuals

46 dev.new(width=12,height=8)
47 expr3 <- list(quote(op <- par(mar = par("mar")*1.5)),
48 quote(mytmp.data <- apply(col.data,2,function(e) e/sum(e))),
49 quote(boxplot(mytmp.data[,colInd], xlab="", ylab="Value",
50 main="Boxplot of normalized column features")),
51 quote(par(op)))
52 heatmap.3(x, scale="column", plot.col.individuals=TRUE, col.data=x,
53 plot.col.individuals.list=list(expr3))

28
Figure 23: Source code of “example-heatmap3a.R”.
 Color Key

60
Heatmap

Count
30
0
−2 0 1 2 3 4
Value

Cadillac Fleetwood

Lincoln Continental

Chrysler Imperial

Pontiac Firebird

Hornet Sportabout

Duster 360

Camaro Z28

Ford Pantera L

Maserati Bora

Merc 450SL

Merc 450SE

Merc 450SLC

Dodge Challenger

AMC Javelin

Hornet 4 Drive

Valiant

Toyota Corona

Porsche 914−2

Datsun 710

Volvo 142E

Merc 230

Lotus Europa

Merc 240D

Merc 280

Merc 280C

Mazda RX4 Wag

Mazda RX4

Ferrari Dino

Fiat 128

Fiat X1−9

Toyota Corolla

Honda Civic
am

vs

cyl

carb

wt

drat

gear

qsec

mpg

hp

disp
Figure 24: Graphical output 1 of source code “example-heatmap3a.R”.

Color Key
60

Heatmap
Count

Row
30

Individuals
0

−2 0 2 4
Value

Cadillac Fleetwood Gross horsepower

Lincoln Continental
Cadillac Fleetwood ●
Chrysler Imperial
Lincoln Continental ●
Pontiac Firebird
Chrysler Imperial ●
Hornet Sportabout
Pontiac Firebird ●
Duster 360
Hornet Sportabout ●

Camaro Z28 Duster 360 ●

Ford Pantera L Camaro Z28 ●

Maserati Bora Ford Pantera L ●

Merc 450SL Maserati Bora ●

Merc 450SE Merc 450SL ●

Merc 450SLC Merc 450SE ●

Dodge Challenger
Merc 450SLC ●

Dodge Challenger ●
AMC Javelin
AMC Javelin ●
Hornet 4 Drive
Hornet 4 Drive ●
Valiant
Valiant ●
Toyota Corona
Toyota Corona ●
Porsche 914−2
Porsche 914−2 ●

Datsun 710 Datsun 710 ●

Volvo 142E Volvo 142E ●

Merc 230 Merc 230 ●

Lotus Europa Lotus Europa ●

Merc 240D Merc 240D ●

Merc 280 Merc 280 ●

Merc 280C Merc 280C ●

Mazda RX4 Wag

Mazda RX4 Wag ●

Mazda RX4 ●
Mazda RX4
Ferrari Dino ●
Ferrari Dino
Fiat 128 ●
Fiat 128
Fiat X1−9 ●
Fiat X1−9
Toyota Corolla ●
Toyota Corolla
Honda Civic ●

Honda Civic
50 100 150 200 250 300
am

vs

cyl

carb

wt

drat

gear

qsec

mpg

hp

disp

hp

Figure 25: Graphical output 2 of source code “example-heatmap3a.R”.

29
 Color Key

Count
0 40
Heatmap
−2 0 1 2 3 4
Value

Cadillac Fleetwood
Lincoln Continental
Chrysler Imperial
Pontiac Firebird
Hornet Sportabout
Duster 360
Camaro Z28
Ford Pantera L
Maserati Bora
Merc 450SL
Merc 450SE
Merc 450SLC
Dodge Challenger
AMC Javelin
Hornet 4 Drive
Valiant
Toyota Corona
Porsche 914−2
Datsun 710
Volvo 142E
Merc 230
Lotus Europa
Merc 240D
Merc 280
Merc 280C
Mazda RX4 Wag
Mazda RX4
Ferrari Dino
Fiat 128
Fiat X1−9
Toyota Corolla
Honda Civic
am

vs

cyl

carb

wt

drat

gear

qsec

mpg

hp

disp
Mean features
●
Individuals

150
Column

●
Mean
50

● ●
● ● ● ● ● ● ●
0

am

vs

cyl

carb

wt

drat

gear

qsec

mpg

hp

disp
Figure 26: Graphical output 3 of source code “example-heatmap3a.R”.

Color Key
Count
0 40

Heatmap
−2 0 1 2 3 4
Value

Cadillac Fleetwood
Lincoln Continental
Chrysler Imperial
Pontiac Firebird
Hornet Sportabout
Duster 360
Camaro Z28
Ford Pantera L
Maserati Bora
Merc 450SL
Merc 450SE
Merc 450SLC
Dodge Challenger
AMC Javelin
Hornet 4 Drive
Valiant
Toyota Corona
Porsche 914−2
Datsun 710
Volvo 142E
Merc 230
Lotus Europa
Merc 240D
Merc 280
Merc 280C
Mazda RX4 Wag
Mazda RX4
Ferrari Dino
Fiat 128
Fiat X1−9
Toyota Corolla
Honda Civic
am

vs

cyl

carb

wt

drat

gear

qsec

mpg

hp

disp

Boxplot of normalized column features

0.00 0.04 0.08

●
Individuals

Value
Column

●
●

am vs cyl carb wt drat gear qsec mpg hp disp

Figure 27: Graphical output 4 of source code “example-heatmap3a.R”.

30
 A.5.2 Examples using ruspini data

example-heatmap3b.R (fig. 28) is an example on how to make a heatmap with summary visualization of
clusters.
example-heatmap3b.R
1 ## load library
2 require("GMD")
3 require(cluster)
4

5 ## load data
6 data(ruspini)
7

8 ## heatmap on a `dist' object

9 x <- gdist(ruspini)
10 main <- "Heatmap of Ruspini data"
11 dev.new(width=10,height=10)
12 heatmap.3(x, main=main, mapratio=1) # default with a title and a map in square!
13 heatmap.3(x, main=main, revC=TRUE) # reverse column for a symmetric look
14 heatmap.3(x, main=main, kr=2, kc=2) # show partition by predefined number of clusters
15

16 ## show partition by elbow

17 css.multi.obj <- css.hclust(x,hclust(x))
18 elbow.obj <- elbow.batch(css.multi.obj,ev.thres=0.90,inc.thres=0.05)
19 heatmap.3(x, main=main, revC=TRUE, kr=elbow.obj$k, kc=elbow.obj$k)
20 heatmap.3(x, main=main, sub=sub("\n"," ",attr(elbow.obj,"description")), cex.sub=1.25,
21 revC=TRUE,kr=elbow.obj$k, kc=elbow.obj$k) # show elbow info as subtitle
22

23 ## side plot for every row clusters

24 dev.new(width=10,height=10)
25 expr1 <- list(quote(plot(do.call(rbind,i.x),xlab="x",ylab="y",
26 xlim=range(ruspini$x),ylim=range(ruspini$y),)))
27 heatmap.3(x, main=main, revC=TRUE, kr=elbow.obj$k, kc=elbow.obj$k, trace="none",
28 row.data=as.list(data.frame(t(ruspini))),
29 plot.row.clusters=TRUE,plot.row.clusters.list=list(expr1))
30

31 ## side plot for every col clusters

32 dev.new(width=10,height=10)
33 expr2 <- list(quote(plot(do.call(rbind,i.x),xlab="x",ylab="y",
34 xlim=range(ruspini$x),ylim=range(ruspini$y),)))
35 heatmap.3(x, main=main, revC=TRUE, kr=elbow.obj$k, kc=elbow.obj$k, trace="none",
36 col.data=as.list(data.frame(t(ruspini))),
37 plot.col.clusters=TRUE,plot.col.clusters.list=list(expr2))
38

Figure 28: Source code of “example-heatmap3b.R”.

31
 Color Key
Count

Heatmap of Ruspini data

400

Row
A "good" k=4 (EV=0.93) is detected when the EV is no less than 0.9 and the increment of EV is no more than 0.05 for a bigger k. Clusters
0

0 100
Value
2 3 1 4
63
66 Row group 2 (n=23)

150
62 ●●●●●
● ●●
●● ●●
61 ●●●● ● ●● ●●
64 ●
68
74
73

y
75

0 50
72
65
2

71
70
69
67 0 20 40 60 80 120
20 x
11
13
12 Row group 3 (n=17)

150
7
6
8 ● ● ●
●
●
●
4
16
● ● ●● ●●
●●
19 ●● ●

y
18

0 50
17
14
15
10
9
0 20 40 60 80 120
3

3
2
5 x
1
46
48 Row group 1 (n=20)

150
47
57
55
56
58 ●

y
59 ●● ●● ●
60 ● ●● ●
● ●● ● ●●
●●
●

0 50
45 ●
44
54
50
52 0 20 40 60 80 120
53
1

49
51
x
28
27 Row group 4 (n=15)

150
30
29
22
21
23
24

y
41

0 50
42
43
37 ●●● ●●● ●
38 ●●●●●
● ●
●
34
33 0 20 40 60 80 120
4

40
36 x
39
35
32
31
26
25
63
66
62
61
64
68
74
73
75
72
65
71
70
69
67
20
11
13
12
7
6
8
4
16
19
18
17
14
15
10
9
3
2
5
1
46
48
47
57
55
56
58
59
60
45
44
54
50
52
53
49
51
28
27
30
29
22
21
23
24
41
42
43
37
38
34
33
40
36
39
35
32
31
26
25

Col group 2 (n=23) Col group 3 (n=17) Col group 1 (n=20) Col group 4 (n=15)
150

150

150

150

●●●●●●●
●●●
●● ●●
●●●●●● ●
● ● ●
● ● ●
●
●
● ● ●●
●●
●● made by
100

100

100

100
Clusters

●●●
● Function: heatmap.3
Col

●●●●
●● ● ● Package: GMD
●● ● ●
●●●
● ● ●●
50

50

50

50

in R
●●
●●●●●
●
● ●
●● ● ●
●
0

0 40 80 120 0 40 80 120 0 40 80 120 0 40 80 120

x x x x

Figure 29: Graphical output 1 of source code “example-heatmap3b.R”.

32
B Data

B.1 GMD dataset overview

> data(package="GMD")

Data sets in package 'GMD':

cage CAGE Data
cagel CAGE Data
chipseq_hCD4T ChIP-seq Data
chipseq_mES ChIP-seq Data

B.2 CAGE data: cage and cagel

> help(cage)

> data(cage)
> class(cage)

[1] "list"

> length(cage)

[1] 20

> names(cage)

[1] "NA (T01F029805F8)" "Glul (T01F092C2995)"

[3] "Cyp4a14 (T04R06C91673)" "Stxbp4 (T11R05607FD4)"
[5] "D230039L06Rik (T01F0AA465EB)" "Gas5 (T01F09995479)"
[7] "Rab5c (T11R05FBC6C4)" "BC003940 (T11R072A6CB0)"
[9] "Tpt1 (T14F04079189)" "Pcna (T02R07DE319B)"
[11] "D0H4S114 (T18R020553F0)" "Gsto1 (T19F02D03566)"
[13] "Hsd11b1 (T01R0B8305BD)" "Csf1 (T03R0672174D)"
[15] "B2m (T02F0743FF05)" "Alox5ap (T05F08BCF2C4)"
[17] "Pfkfb3 (T02R00AEC2D8)" "Hig1 (T09R0743763C)"
[19] "Cd72 (T04R028B8BC9)" "Egln1 (T08R0769239F)"

> data(cagel)
> names(cagel)

[1] "NA (T17F05912B83)" "Tpt1 (T14F04079189)"

[3] "B2m (T02F0743FF05)" "Grn (T11F0615F289)"
[5] "2600001A11Rik (T12R043A2595)" "Rbbp7 (T0XF091A7ACA)"
[7] "Rpl41 (T10R07AB7138)" "H2afy (T13R034ACF47)"
[9] "Dscr1l1 (T17F02802885)" "Ckap4 (T10R0504CE97)"
[11] "Rab5c (T11R05FBC6C4)" "Pfkfb3 (T02R00AEC2D8)"
[13] "Csf1 (T03R0672174D)" "Ctsb (T14F0348EDBA)"
[15] "Crim1 (T17F04928998)" "3930401E15Rik (T18R02CDD141)"
[17] "Rai17 (T14F014BF473)" "Hig1 (T09R0743763C)"
[19] "Apbb2 (T05R03E329C8)" "Ptn (T06R0230806E)"
[21] "Tmeff2 (T01F030EC757)" "Mrps6 (T16F0583C906)"
[23] "Hsd11b1 (T01R0B8305BD)" "D0H4S114 (T18R020553F0)"
[25] "4930430J02Rik (T09F036E80C6)" "Phtf2 (T05R0125E896)"
[27] "Trpv2 (T11F03B4EBD8)" "Slco3a1 (T07R03AC06B9)"
[29] "Scd1 (T19R029B5186)" "Ctxn (T08R0040864A)"
[31] "5730596K20Rik (T19F006DFC1A)" "Arbp (T05F06BBE13B)"
[33] "Klhl5 (T05F03CCA673)" "Gsto1 (T19F02D03566)"
[35] "NA (T02R07EF5EDA)" "Srpk1 (T17R019F4A41)"
[37] "Nudt7 (T08F06C3B651)" "Tnfrsf10b (T14F03AB1306)"
[39] "Egln1 (T08R0769239F)" "9630050M13Rik (T02F002EC972)"
[41] "BC003940 (T11R072A6CB0)" "Ppia (T11F00604AFF)"
[43] "Alox5ap (T05F08BCF2C4)" "Pcna (T02R07DE319B)"
[45] "Gch1 (T14R02602138)" "Yap1 (T09R0079F3FF)"
[47] "Vrk1 (T12F06010C9B)" "Cd72 (T04R028B8BC9)"
[49] "Wdtc1 (T04R07DAFEDC)" "Centg2 (T01F055392D1)"

33
B.3 ChIP-seq data: chipseq mES and chipseq hCD4T
> help(chipseq)

> data(chipseq_mES)
> class(chipseq_mES)

[1] "list"

> length(chipseq_mES)

[1] 6

> names(chipseq_mES)

[1] "H3K27me3" "H3K36me3" "H3K4me1" "H3K4me2" "H3K4me3" "H3K9me3"

> data(chipseq_hCD4T)
> names(chipseq_hCD4T)

[1] "CTCF" "H2AK5ac" "H2AK9ac" "H2AZ" "H2BK120ac" "H2BK12ac"

[7] "H2BK20ac" "H2BK5ac" "H2BK5me1" "H3K14ac" "H3K18ac" "H3K23ac"
[13] "H3K27ac" "H3K27me1" "H3K27me2" "H3K27me3" "H3K36ac" "H3K36me1"
[19] "H3K36me3" "H3K4ac" "H3K4me1" "H3K4me2" "H3K4me3" "H3K79me1"
[25] "H3K79me2" "H3K79me3" "H3K9ac" "H3K9me1" "H3K9me2" "H3K9me3"
[31] "H3R2me1" "H3R2me2" "H4K12ac" "H4K16ac" "H4K20me1" "H4K20me3"
[37] "H4K5ac" "H4K8ac" "H4K91ac" "H4R3me2"

34
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35