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 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

CONTENTS

Preface to the fifth edition xv

Plan of the book xxi
Introduction to bioinformatics on the web xxii
Acknowledgements xxiii

1 Introduction 1
Life in space and time 4
Phenotype = genotype + environment + life history + epigenetics 4
Evolution is the change over time in the world of living things 5
Biological classification and nomenclature 6
Dogmas: central and peripheral 9
The structure of DNA 9
Transcription and translation 12
The structures of proteins 12
Statics and dynamics 17
Systems biology 17
The human genome 19
Variation in human genome sequences 20
The human genome and medicine 21
Databases in molecular biology 28
Observables and data archives 29
A database without effective modes of access is merely a data graveyard 29
Information flow in bioinformatics 31
Curation, annotation, and quality control 32
The World Wide Web 33
Electronic publication 34
Computers and computer science 34
Programming 35
Après moi, le déluge? Sorry—too late! 38
How much sequencing power is there in the world? 41
How does the amount of data in bioinformatics compare with other large scientific
information archives? 41
Recommended reading 42
Exercises and Problems 43

2 From genetics to genomes 48

The classical genetics background 49
DNA embodies genes 50
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

viii Contents

Maps and tour guides 51

Linkage maps 51
Linkage 51
Chromosome banding 53
High-resolution maps, based directly on DNA sequences 55
Restriction maps 56
DNA sequencing 57
Frederick Sanger and the development of DNA sequencing 57
DNA sequencing by termination of chain replication 58
Automation of DNA sequencing 60
Next-generation sequencing 61
Paired-end reads 66
Life in the fast lane 67
Assembly—computational aspects 68
Pattern matching 68
Suffix trees 68
Fragment assembly 70
Genomics in personal identification 73
DNA ‘fingerprinting’ 74
Personal identification by amplification of specific regions has superseded
the RFLP approach 75
Mitochondrial DNA 76
Analysis of non-human DNA sequences 77
Parentage testing 78
Ethical, legal, and social issues 80
Databases containing human DNA sequence information 80
Use of DNA sequencing in research on human subjects 82
Recommended reading 83
Exercises and Problems 84

3 The panorama of life 88

Genomes, transcriptomes, and proteomes 89
Genes 89
Proteomics and transcriptomics 91
Eavesdropping on the transmission of genetic information 93
Genome-sequencing projects 93
Genomes of prokaryotes 94
The genome of the bacterium Escherichia coli 95
The genome of the archaeon Methanocaldococcus jannaschii 97
The genome of one of the simplest organisms: Mycoplasma genitalium 98
Metagenomics: the collection of genomes in a coherent environmental sample 99
The human microbiome 101
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

Contents ix

Genomes of eukarya 102

Gene families 103
The genome of Saccharomyces cerevisiae (Baker’s yeast) 103
The genome of Caenorhabditis elegans 105
The genome of Drosophila melanogaster 105
The genome of Arabidopsis thaliana 107
The genome of Homo sapiens (the human genome) 108
Protein-coding genes 109
Repeat sequences 109
RNA 110
Single-nucleotide polymorphisms and haplotypes 110
Systematic measurements and collections of single-nucleotide
polymorphisms 113
Genetic diversity in anthropology 114
DNA sequences and languages 116
Evolution of genomes 116
Please pass the genes: horizontal gene transfer 118
Comparative genomics of eukarya 119
Recommended reading 120
Exercises and Problems 121

4 Alignments and phylogenetic trees 123

Introduction to sequence alignment 124
Dotplots and sequence alignments 130
Measures of sequence similarity 132
Scoring schemes 132
Derivation of substitution matrices: PAM matrices 133
Computing the alignment of two sequences 135
Variations and generalizations 135
Approximate methods for quick screening of databases 135
The dynamic programming algorithm for optimal pairwise sequence
alignment 137
Significance of alignments 141
Multiple sequence alignment 143
Applications of multiple sequence alignments to database searching 143
Profiles 146
PSI-BLAST 147
Complete pairwise sequence alignment of human PAX-6 protein and Drosophila
melanogaster eyeless 151
Hidden Markov Models 152
Phylogeny 154
Determination of taxonomic relationships from molecular properties 155
Use of sequences to determine phylogenetic relationships 159
Use of SINES and LINES to derive phylogenetic relationships 161
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

x Contents

Phylogenetic trees 162

Clustering methods 164
The maximum-likelihood method 165
Reconstruction of ancestral sequences 165
Pyruvate decarboxylase: synthesis, activity, and crystal structure of predicted ancestor 167
The problem of varying rates of evolution 168
Bayesian methods 169
Are trees the correct way to present phylogenetic relationships? 169
Computational considerations 170
Putting it all together 171
Recommended reading 171
Exercises and Problems 172

5 Structural bioinformatics and drug discovery 177

Introduction 178
Protein stability and folding 180
The Sasisekharan–Ramakrishnan–Ramachandran plot
describes allowed mainchain conformations 180
The sidechains 181
Protein stability and denaturation 183
Protein folding as a process 185
Applications of hydrophobicity 187
Coiled-coiled proteins 187
Description of the variety of protein structures 190
Superposition of structures, and structural alignments 192
Evolution of protein structures 197
Classifications of protein structures 199
SCOP 199
Protein structure prediction and modelling 201
A priori and empirical methods 202
Critical Assessment of Structure Prediction 203
Secondary structure prediction 204
Homology modelling 205
Fold recognition 205
Conformational energy calculations and molecular dynamics 207
ROSETTA 209
Protein structure prediction from contact maps derived from correlated mutations
in multiple sequence alignments 210
Design of novel proteins 213
Drug discovery and development 215
The lead compound 216
Improving on the lead compound: quantitative structure–activity relationships 217
Bioinformatics in drug discovery and development 218
Molecular modelling in drug discovery 219
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

Contents xi

Recommended reading 225

Exercises and Problems 228

6 Scientific publications and archives: media, content, access,

and presentation 233
The scientific literature 234
Access to scholarly publications 235
Open access 236
The Public Library of Science 237
Traditional and digital libraries 237
How to populate a digital library 238
The information explosion 239
The web: higher dimensions 239
New media: video, sound 240
Searching the scientific literature 240
Bibliography management 241
Databases 242
Database contents 242
Database quality control 243
The literature as a database 244
Database organization 244
Annotation 246
Markup languages 248
Database access 250
Links 250
Database interoperability 251
Data mining 251
Programming languages and tools for database construction and access 255
Traditional programming languages 255
Scripting languages 256
Program libraries specialized for molecular biology 256
Java—computing over the web 256
Natural language processing 257
Natural language processing in mining the biomedical literature 258
Biomedical applications of text mining 260
Hypothesis generation 264
A glaucoma-related network derived by text mining 265
Recommended reading 268
Exercises and Problems 269

7 Artificial intelligence and machine learning 271

What are artificial intelligence and machine learning? 272
Classification and clustering 273
Binary classifier 276
Receiver Operating Characteristic (ROC) curves 277
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

xii Contents

Artificial neural networks 279

Self-organizing maps 281
Decision trees 281
Support vector machines (SVMs) 286
Kernel methods 286
Clustering 287
Clustering by graph spectral theory 291
Recommended reading 293
Exercises and Problems 293

8 Introduction to systems biology 296

Introduction 297
Networks and graphs 298
Connectivity in networks 299
Dynamics, stability, and robustness 301
Some sources of ideas for systems biology 302
Complexity of sequences 302
Shannon’s definition of entropy 303
Complexity of sequences 304
The relationship between complexity, randomness, and compressibility 305
The Burrows-Wheeler Transform 305
Inverting the Burrows-Wheeler Transform 306
The Burrows-Wheeler Transform brings repeats together, facilitating compression 306
Use of the Burrows-Wheeler transform for searching for patterns in strings 306
Complexity of other types of biological data 308
Static and dynamic complexity 308
Predictability and chaos 309
Analysis and comparison of networks 310
Analysis of graphs by matrix algebra 311
Graph isomorphism 312
Recommended reading 314
Exercises and Problems 314

9 Metabolic pathways 317

Introduction 318
Classification of protein function 320
The Enzyme Commission 320
The Gene OntologyTM Consortium protein function classification 320
Prediction of protein function 321
Catalysis by enzymes 324
Active sites 325
Cofactors 325
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

Contents xiii

Protein–ligand binding equilibria 326

Enzyme kinetics 327
Measures of effectiveness of enzymes 328
How do enzymes evolve new functions? 329
Control over enzyme activity 329
Structural mechanisms of evolution of altered or novel protein functions 329
Pathways and limits in the divergence of sequence, structure,
and function 334
Evolution by gene duplication 335
Databases of metabolic pathways 337
The Kyoto Encyclopedia of Genes and Genomes (KEGG) 339
Evolution and phylogeny of metabolic pathways 341
Pathway comparison 341
Alignment of metabolic pathways 343
Comparing linear metabolic pathways 343
Comparing non-linear metabolic pathways: The pentose phosphate pathway
and the Calvin-Benson cycle 346
Dynamics of metabolic networks 347
Robustness of metabolic networks 347
Dynamic modelling of metabolism 347
Simulation of metabolic pathways in Plasmodium falciparum 351
The Human Metabolome Database supports clinical applications to the study
of inborn errors of metabolism, and to cancer 352
Recommended reading 353
Exercises and Problems 353

10 Control of organization and organization of control 355

Transcriptomics 356
The ENCODE Project 357
Determination of RNA sequences 358
RNAseq v. microarrays 358
DNA microarrays 359
RNAseq 363
The Genotype-Tissue Expression (GTEx) project 366
Expression patterns in different physiological states 367
Variation of expression patterns during the life cycle of Drosophila
melanogaster 368
Different life stages make different demands on different genes 370
Protein complexes and aggregates 373
Properties of protein–protein complexes 373
Protein interaction networks 375
Components of the primosome assembly in Bacillus subtilis 378
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

xiv Contents

Regulatory networks 380

Signal transduction and transcriptional control 380
Structural biology of regulatory networks 382
Examples of relatively simple regulatory control networks 383
Regulation of the lactose operon in E. coli 383
The genetic switch of bacteriophage λ 385
The diauxic shift in Saccharomyces cerevisiae 389
Logical structure of regulatory networks 391
The transcriptional regulatory network of E. coli 391
The transcriptional regulatory network of Saccharomyces cerevisiae 392
Adaptability of the yeast regulatory network 393
Recommended reading 396
Exercises and Problems 396

Conclusions 399
Index 400
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

PREFACE TO THE FIFTH EDITION

On June 26, 2000, the sciences of biology and to prevent Celera—or anyone else—from applying
medicine changed forever. Prime Minister of the for patents.
United Kingdom Tony Blair and President of The academic groups lined up against Celera were
the United States Bill Clinton held a joint press a collaborating group of laboratories based primar-
conference, linked via satellite, to announce the ily but not exclusively in the UK and USA. These
completion of the draft of the Human Genome. The included the Wellcome Trust Sanger Institute in Eng-
New York Times ran a banner headline: ‘Genetic land; Washington University in St. Louis, Missouri;
Code of Human Life is Cracked by Scientists’. The the Whitehead Institute at the Massachusetts Institute
sequence of 3 billion bases was the culmination of of Technology in Cambridge, Massachusetts; Baylor
over a decade of work, during which the goal was College of Medicine in Houston, Texas; the Joint
always clearly in sight and the only questions were Genome Institute at Lawrence Livermore National
how fast the technology could progress and how Laboratory in Livermore, California; and the RIKEN
generously the funding would flow. The Table shows Genomic Sciences Center, now in Yokahama, Japan.
some of the landmarks along the way. Both sides could dip into deep pockets. Celera had
Next to the politicians stood the scientists. The its original venture capitalists; its current parent com-
late John Sulston (later Sir John, CH, FRS), Direc- pany, the PE Corporation; and, after going public,
tor of the Wellcome Trust Sanger Institute in the anyone who cared to take a flutter. The UK Medical
UK, had been a key player since the beginning of Research Council and The Wellcome Trust supported
high-throughput sequencing methods. He had grown the Sanger Institute. The US National Institutes of
with the project from the earliest ‘one man and a Health and Department of Energy supported the US
dog’ stages to the large international consortium. academic labs.
In the US, appearing with President Clinton were On June 26, 2000 the contestants agreed to declare
Francis Collins, director of the US National Human the race a tie, or at least a carefully out-of-focus
Genome Research Institute, representing the US pub- photo finish.
licly funded efforts; and J. Craig Venter, President The sequencing of the human genome ranks
and Chief Scientific Officer of Celera Genomics Cor- with the Manhattan project that produced atomic
poration, representing the commercial sector. It is weapons during the Second World War, and the
difficult to introduce these two without thinking, ‘In space program that sent people to the moon, as one
this corner . . . and in this corner . . . ’. Although never of the great bursts of technological achievement of
actually coming to blows, there was certainly intense the last century. These projects share a grounding in
competition, in the later stages a race. fundamental science, and large-scale and expensive
The race was more than an effort to finish first and engineering development and support. For biology,
to receive scientific credit for priority. It was a race neither the attitudes nor the budgets will ever be
after which the contestants would be tested not for the same. Soon a ‘one man and a dog project’ will
whether they had taken drugs, but whether they and refer only to an afternoon’s undergraduate practical
others could discover them. Clinical applications experiment in sequencing and comparison of two
were a prime motive for support of the Human mammalian genomes.
Genome Project. Once the courts had held that The human genome is only one of the many
gene sequences were patentable—with enormous complete genome sequences known. Taken together,
potential payoffs for drugs and diagnostic tools based genome sequences from organisms distributed widely
on them—the commercial sector rushed to submit among the branches of the tree of life give us a
patents on sets of sequences that they determined, sense, only hinted at before, of the very great unity
and the academic groups rushed to place each bit of in detail of all life on Earth. These results have
sequence that they determined into the public domain changed our perceptions, much as the first pictures
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

xvi Preface to the fifth edition

Landmarks in the Human Genome Project

1953 Watson–Crick structure of DNA published.
1975 F. Sanger, and independently A. Maxam and W. Gilbert, develop methods for sequencing DNA.
1977 Bacteriophage ϕX-174 sequenced: first ‘complete genome’.
1980 US Supreme Court holds that genetically modified bacteria are patentable. This decision was the basis for
patenting of genes.
1981 Human mitochondrial DNA sequenced: 16 568 base pairs.
1984 Epstein–Barr virus genome sequenced: 172 281 base pairs
1990 International Human Genome Project launched: target horizon 15 years.
1991 J. Craig Venter and colleagues identify active genes via expressed sequence tags, sequences of initial
portions of DNA complementary to messenger RNA.
1992 Complete low-resolution linkage map of the human genome.
1992 Beginning of the Caenorhabditis elegans sequencing project.
1992 Wellcome Trust and UK Medical Research Council establish the Sanger Centre for large-scale genomic
sequencing, directed by J. Sulston.
1992 J. Craig Venter forms The Institute for Genome Research (TIGR), associated with plans to exploit sequencing
commercially through gene identification and drug discovery.
1995 First complete sequence of a bacterial genome, Haemophilus influenzae, by TIGR.
1996 High-resolution map of human genome: markers spaced by ∼600 000 base pairs.
1996 Completion of yeast genome, first eukaryotic genome sequence.
May 1998 Celera claims to be able to finish human genome by 2001. Wellcome responds by increasing funding to
Sanger Centre.
1998 Caenorhabditis elegans sequence published.
September 1, 1999 Drosophila melanogaster genome sequence announced, by Celera Genomics; released Spring 2000.
1999 Human Genome Project states goal: working draft of human genome by 2001 (90% of genes sequenced to
>95% accuracy).
December 1, 1999 Sequence of first complete human chromosome published.
June 26, 2000 Joint announcement of complete draft sequence of human genome.
2003 Fiftieth anniversary of discovery of the structure of DNA. Announcement of completion of human genome
sequence.

of Earth from space engendered a unified view of Computing is an essential component of the first
our planet. three of these; CRISPR is spawning bioinformatics
support.
Molecular biology has seen four major break- Where will this lead? We can study the present as
throughs since the previous edition of this book thoroughly as we wish, and the past as extensively as
appeared: we can. What of the future? Molecular biologists used
to be like astronomers, in that we could observe our
• Explosive growth in next-generation genome
subjects but not affect them. This is no longer true:
sequencing
we now have the ability, and the challenge, of direct
• Consistent successful prediction of protein struc- control over living things, including but not limited
ture from amino acid sequence to ourselves.
• Achievement of atomic resolution in cryo-electron A gratifying consequence of this progress is its con-
microscopy tributions to medicine, agriculture, and technology.
• CRISPR for genome editing. A better understanding of life processes empowers us
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

Preface to the fifth edition xvii

to deal with them when they go wrong, and even to lengthier answers or in some cases calculations. The
improve them when they do not. third category, ‘weblems,’ require access to the World
The human genome is fundamentally about Wide Web. Weblems are designed to give readers
information, and computers were essential both practice with the tools required for further study and
for the determination of the sequence and for the research in the field.
applications to biology and medicine that are flowing Examples and problems appear in the text.
from it. Computing contributed not only the raw Weblems are on the Online Resource Centre, a
capacity for processing and storage of data, but also website companion to the book.
the mathematically sophisticated methods required Some computing, based on the widely available
to achieve the results. The marriage of biology and language PERL, is introduced in this book. PERL
computer science has created a new field called is a relatively simple but extremely effective
bioinformatics. programming language. It is one of the languages
Today bioinformatics is an applied science. popular in the bioinformatics community. Similar
Computers have been an essential component of
the projects that determine sequence, structure, and
other types of data. We use computer programs to PERL, PYTHON, and RUBY
make inferences from the data archives of molecular
biology and medicine, to make connections among
As languages, PERL, PYTHON, and RUBY have roughly
them, and to derive useful and interesting predictions.
equivalent ranges of expression. However, applica-
This book is aimed at students and practising
tions of bioinformatics have begun to coalesce around
scientists who need to know how to access the
PYTHON: PYTHON arguably has a richer literature,
data archives, including but not limited to those of
both in training material and in available program mod-
genomes and proteins, how to use the tools that
ules (see, for example, http://rosalind.info/problems/
have been developed to work with these archives, locations/).
and the kinds of questions that these data and tools Nevertheless sample programs in the text are in PERL.
can answer. In fact, there are a lot of sources of this There are two reasons for this. The main one is that PERL
information. Sites treating topics in bioinformatics programs are more self-contained. PYTHON programs
sprawl out all over the Web. Our challenge is to select tend to invoke external ‘black boxes’ and are therefore
an essential core of this material and to describe it, less perspicuous. It is also true that PYTHON is divided
together with the necessary biological background, between PYTHON 2 and PYTHON 3—which to choose?
clearly and coherently, at an introductory level. Unfortunately they are not fully compatible. It is likely
It is assumed that the reader already has some that PYTHON 3 will eventually take over, but a large
knowledge of modern molecular biology, and some amount of legacy software exerts a viscous drag. Also,
facility at using a computer. The purpose of this book presenting PERL programs in the book has the advantage
is to build on and develop this background. It is that readers can be asked to provide PYTHON or RUBY
suitable as a textbook for advanced undergraduates versions as exercises!1
or beginning postgraduate students. Many worked- PERL, PYTHON, and RUBY each has an extensive
out examples are integrated into the text, and refer- repertoire of available components utilizable as ‘stand-
ences to useful websites and recommended reading alone’ modules, or for combination into novel programs:
are provided. A useful list of biomedical courses on- https://bioperl.org, https://biopython.org and https://
line appears at https://www.science.co.il/biomedical/ bioruby.org, and ‘Cookbooks’: Christiansen, T. & Tork-
ington, N. (2009). Perl cookbook 2nd edn; Beazley, D.
Courses.php.
& Jones, B.K. (2013). Python cookbook 3rd edn; and
Problems test and consolidate understanding, pro-
Carlson, L. & Richardson, L. (2015). Ruby cookbook 2nd
vide opportunities to practise skills, and explore addi-
edn; all published by O’Reilly Media, Sebastopol, CA.
tional topics. Three types of problems supplement the
texts of the chapters. Exercises are short and straight- 1 For a comment about PERL to PYTHON conversion see

forward applications. Problems also require no http://code.activestate.com/lists/python-list/292576/

information not contained in the text, but require
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

xviii Preface to the fifth edition

languages include PYTHON and RUBY (see Box on The most striking change in attitude has been
previous page); each of these has its adherents. a refocus on integration; that is, of trying to see
Examples of simple programs appear in the context life processes as unified systems. As I wrote at the
of biological problems. Many simple programming end of Introduction to Protein Science: Architecture,
tasks are assigned as exercises, problems, or weblems Function and Genomics, ‘During the last century,
at the ends of the chapters. molecular biologists have been taking living things
Bioinformatics has grown, explosively, since the apart. Our task now is to understand how to put
first edition of this book appeared. Underlying this them back together.’ We have large amounts of data.
surge are the multiplication and growth of data Now we want to see how they interrelate. At the heart
streams. In addition to genomics, these include, but of life processes are complicated patterns of interac-
are not limited to, transcriptomics, metabolomics, tion among the components, in space and in time.
and regulatory networks. The human genome, To understand these patterns, the field has moved
available in draft form when the first edition towards combining information into networks, and
appeared, is now complete. Large-scale human analysing their structures and dynamics. Many tools
genome sequencing projects are underway. Joining are now available for storing, visualizing, analysing,
these results are the complete genomes of hundreds and comparing networks. Contemporary bioinfor-
of other eukaryote species, very large numbers of matics could not do without them.
prokaryotes, and many other organelle and viral Research and applications require that the data be
sequences. The extension to metagenomics—the available in useful form. It is not enough to make the
survey of distributions of sequences in a region of data public. The information must be subjected to
the land or ocean, or the human body—is a recent quality control, annotation, and a logical structure
development but a copious producer of data. These must be imposed on it to make information retrieval
data illuminate one another. One story that they tell possible. For this we are indebted to the institutions
is about unsuspected underlying unities of all living that archive, curate, organize, and distribute the data.
things, despite the obvious and profound differences A trend has seen mergers of these groups into col-
in morphology and lifestyle. laborative projects spanning the continents. In accord
These genome data are the harvest of new and with the need to integrate the study of different types
more powerful DNA sequencing methods. Of course of data, we are moving in the direction of a large-
particular attention is focused on human genomes: scale unified biological data archive and repository.
several hundred thousand individual genomes are Conversely, individual scientists will be able to define
known. The goal of the US$1000 human genome has ‘virtual databanks’ tailoring access to the information
been achieved. This enterprise continues to grow: it to suit particular needs and interests.
is likely that during the lifetimes of many readers, Major changes in information distribution involve
human genome sequencing will be nearly universal the transition from paper to electronic libraries. The
in many countries. implications for scientific research are only a part
Genomic sequences are supplemented by other of the great social revolution that has flowed from
data streams. Patterns of gene expression, and the development of the Web; comparable to, if not
networks of regulatory interactions, show how cells exceeding, the one impelled by the printing press
and organisms implement the information in the 500 years ago. Only connect.
DNA. The potential for the life of an organism is The natural habitat of bioinformatics is the Web.
contained in its genome, but it would be impossible To achieve their scientific goals most projects begin by
to deduce a biography from it. Genomes are not identifying the relevant data on one or more websites;
formulae or scripts. It is in the RNAs and proteins, analysing them, usually applying tools available on
and their interactions amongst themselves and the Web; and reaching robustly justified conclusions;
with DNA, that we must seek the set of life’s ultimately leading to a publication also distributed on
activities, tragic and comic, contingent on and the Web. However, a serious problem with the Web
responsive to developmental programmes and/or the is its volatility. Sites come and go, leaving trails of
environment. dead links in their wake. Databases proliferate. Many
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

Preface to the fifth edition xix

new databases represent different ways of ‘slicing and available tools rather than creating new ones.
dicing’ information from other resources. Changes Many programs are freely available. As with
in a source database may trickle down slowly—or the data, the challenges are often to recombine
not at all—to derived resources. The resulting lack available programs. Constructing a cart requires
of consistency is a serious problem. not reinventing wheels, but assembling them.
It is necessary to try to find a few gateways that
Practitioners of bioinformatics must assimilate these
are stable: not only continuing to exist but also kept
attitudes and reflect them in their training.
up-to-date in both their contents and links. I have
Accordingly, there is somewhat more mention of
suggested some such sites, but many others are just
topics from computer science in this edition than in
as good. The problem is not to create a long list
the previous ones. This is in part because of critical
of useful sites—this has been done many times, and
comments received, but also because it is easier to use
is relatively easy—but to create a short one—this
these tools than before. Instead of needing to write a
is much harder! The site https://www.science.co.il/
sophisticated program, it is often possible to find an
biomedical/databases/ is comprehensive but useful.
available distributed one, or even a web server.
All research in contemporary biology and medicine
Indeed, one of the arguments for the suggestion
depends on data, and on programs to retrieve and
that sophisticated programming skills are not gener-
analyse them. There is consensus that all biomedical
ally required is the great panoply of freely available
scientists must achieve a minimum of programming
programs, written by acknowledged professionals.
skills, but there is vigorous debate over what this
What is essential is developing skill in using these
minimum level should be. In the preface to the fourth
programs, and in intelligent interpretion of the results
edition I wrote: ‘The point of view expressed in this
that they produce.
book is that molecular biologists based primarily
This is the goal of the problems and projects in the
in a “wet” lab must dip no more than their toes
the text and in the Online Resource Centre. Many of
into the stream; those based primarily at a computer
them are ‘weblems’ based on data and facilities on the
must wade in up to their waist perhaps; but only
web. Some are programming exercises. Some of these
those specializing in computer science and software
involve modifying programs. Such challenges can be
development must undergo total immersion.’ Today,
more focused than writing programs from scratch.
this appears as a gross understatement, given that:
Moreover, such exercises better exemplify situations
1. The growth in the amounts, and kinds, of that arise in actual research.
data. Traditional input streams, such as DNA Some of the exercises, problems, and weblems,
sequences; and novel ones, such as tissue- although not requiring any programming, can be
dependent inventories of RNAs (transcriptomics), solved more easily by writing short programs. Read-
produce torrential output. There is danger in ers are encouraged to try this approach whenever
adopting too-casual an attitude. appropriate.
The minimal computing skills essential for a
2. A revolution in approach to data has been the
biomedical scientist would also include facility
recognition that the structure of data is as impor-
with using social media for communication (it is
tant as the values (it would be going a bit too far
assumed that readers are familiar with Facebook and
to say more important).
YouTube, but there are others that are in use for
3. Different ways to recombine data provide novel communication among scientists), and the ability to
approaches. An example would be to combine create a website. Studying from this book and the
genome sequences with signatures of protein func- Online Resource Centre affords an opportunity to
tion to assign roles to gene products. practise these skills. You might, for instance, ‘turn in’
4. The data have elicited new sets of tools for the answers to homework assignments by gathering
analysis and application. The breadth of the them into a Web page. Questions about statements
software literature is so encompassing that that you and the other students found unclear in
research applications tend to involve the selecting your instructor’s lectures—or, conceivably, even in
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

xx Preface to the fifth edition

this book—could be shared and discussed in a blog. Bioinformatics is one of them. I have also written
Indeed, there is now a trend to integrating websites about genomics, and about proteins, in companion
and social media. However, there are security issues. volumes also published by Oxford University
Your instructor might be unhappy if everyone copied Press: Introduction to Protein Science: Architecture,
the answers to the exercises from the first student Function, and Genomics and Introduction to
to post them. A class taught from this book would Genomics. As a result, this book is focused more
afford a fine opportunity to explore the possibilities tightly on the applied science of bioinformatics.
and challenges. Readers are urged to put the books together
There are many different possible points of for a more rounded appreciation of the pageant
view from which to present molecular biology. of life.
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

PLAN OF THE BOOK

• Chapter 1 sets the stage and introduces all of the paper to electronic form. This transition has many
major players: DNA and protein sequences and consequences, both intellectual and practical. It
structures, genomes and proteomes, databases and has had profound effects on research in bio-
information retrieval, the World Wide Web, and informatics.
computer programming. Before developing indi- • Chapter 7 treats the topics of artificial intelligence
vidual topics in detail it is important to see the and machine learning. Few activities, even outside
framework of their interactions. science, have escaped applications of these meth-
• Chapter 2 presents the background of genetics ods. They are playing a role of growing importance
and genomes, and the development of DNA in molecular biology.
sequencing. • Chapter 8 introduces systems biology. The key idea
• Chapter 3 contains a survey and some important of systems biology is integration: how do all the
examples of genome sequencing. pieces fit together? How do they interact? How
• Chapter 4 treats the analysis of relationships among do the individual molecules and processes together
sequences: alignments and phylogenetic trees. create a whole that so far transcends the pieces in
These methods underlie some of the major compu- self-sufficiency?
tational challenges of bioinformatics: detecting • Chapter 9 describes metabolic pathways. The
distant relatives, understanding relationships activities of individual enzymes are the subject
among genomes of different organisms, and tracing matter of classical biochemistry. Understanding
the course of evolution at the species and molecular their controls has been a goal of molecular biology,
levels. revealing a variety of mechanisms at the levels
• Chapter 5 moves into three dimensions, treating of transcription, translation, post-translational
protein structure and folding. Sequence and struc- modifications, and the interaction of inhibitors
ture must be seen as full partners, with bioinfor- and allosteric effectors with enzymes themselves.
matics developing methods for moving back and The integration of these controls is a development
forth between them as fluently as possible. Under- of systems biology, as a continuation of Chapter 8.
standing protein structures in detail is essential • Chapter 10 deals with more general control
for determining their functions and mechanisms mechanisms, including gene expression. Control
of action, and for clinical and pharmacological of gene expression is involved in responses to
applications. stimuli and changes in the cell’s environment,
• Chapter 6 describes the current state of the and governs short- and long-term developmental
scientific literature as it makes the transition from processes.
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

INTRODUCTION TO BIOINFORMATICS ON THE WEB

Bioinformatics is intimately bound up with the World 3. Animations of structural diagrams.

Wide Web. This book is closely coordinated with 4. World Wide Web resources, to supplement treat-
its own website: http://www.oxfordtextbooks.co.uk/ ments of specific topics. Some of these sites imple-
orc/leskbioinf5e/. ment methods, such as sequence alignment, or
This site contains: homology modelling of protein structures. Others
1. References to all sites mentioned in the book, provide curated lists of links to other websites spe-
in context, so that the reader can link to them cialized to particular subjects, such as expression
directly instead of needing to type their locations. databases.
2. In previous editions, the weblems appeared in 5. In general, all material from the book that the
the text. These are now in the Online Resource reader would find useful to have in computer-
Centre. They have been developed and now fea- readable form, including data for exercises and
ture challenges with a range of difficulties, from problems, and all programs, now appears in the
relatively straightforward exercises to extended Online Resource Centre.
projects.
 OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi

ACKNOWLEDGEMENTS

I am grateful to many colleagues for discussions and T. Madden, J. Magré, M. McFall-Ngai, J. McInerney,
advice during the preparation of this book, and to the P. Miller, C. Mitchell, J. Moult, E. Nacheva, C.
universities of Uppsala, Umeå, Rome ‘Tor Vergata’, Notredame, C. Ouzounis, A.P. Pandurangan, H.
Bologna, and Cambridge for the opportunity to try Parfrey, D. Parkinson, A. Pastore, M. Peitsch,
out this material. D. Penny, J. Pettitt, C.A. Praul, C. Ravarani, S.
I thank S. Adhya, D.J. Abraham, S. Aparicio, Reikine, F.W. Roberts, G.D. Rose, P.B. Rosenthal,
M.M. Babu, T. Baglin, D. Baker, S. Balaji, M. B. Rost, B. Santhanam, E.J. Simon, M. Segal,
Bashton, A. Bateman, A. Bench, J.M. Bollinger, V. O. Skovegaard, G. Slodkowicz, E.L. Sonnham-
Bonazzi, M. Brand, A. Brazma, A. Buckle, C. Cantor, mer, R. Srinivasan, R. Staden, J. Sulston, I.
R.W. Carrell, S. Chavali, C. Chothia, D. Crowther, Tickle, A. Tramontano, A.A. Travers, A.R. Venki-
T. Dafforn, A. P. Diz, I. Dodd, R.B. Eckhardt, taraman, G. Vriend, P. Welsch, J.C. Whisstock,
J.G. Ferry, R. Foley, A. Friday, M. Galperin, M. Wildersten, A.S. Wilkins, S.H. White,
M.B. Gerstein, T. Gibson, J. Gough, J. Irving, B. V.E. Womble, and E.B. Ziff for advice and critical
Jorden, J. Karn, K. Karplus, P. Klappa, A.S. Kon- reading.
agurthu, E.V. Koonin, M. Krichevsky, P. Lawrence,
E.L. Lesk, M.E. Lesk, V.E. Lesk, V.I. Lesk, A. Lister, A.M.L.
L. Lo Conte, D.A. Lomas, A.D. MacKerell Jr, Cambridge, July 2018
OUP CORRECTED PROOF – FINAL, 23/4/2019, SPi
Other documents randomly have
different content
another’s intentions in declining her invitation, and found that each
had acted without knowing what the other meant to do. Don’t you
think she must have decided that in the minds of some people the
prayer meeting was an important place, and the engagement to
attend it was not to be lightly broken?
M ERRY CHRISTMAS to all my Blossoms! Glad am I to be able to
give you this happy greeting once more. And to the many new
faces which I greet for the first time, the wish is just as hearty; may
each of you have this month, not only the merriest, but, in the truest
sense of the word, the best Christmas you ever enjoyed in your
lives.
Shall I tell you how to make sure of this? I dare say you know,
but I will just repeat the thought to keep it before your minds. In the
first place, let each of you make a Christmas gift, very costly and
very precious, to the best friend you have. Now I see many
sorrowful faces, and hear low, regretful voices: “You would like to,
but you have no money to spend, or at best but very little, and
cannot afford anything costly.” Is that what I hear you say?
Mistaken, every one of you. The most costly and precious thing in
the world, is the strong true heart which has its home in your body,
my boy—or my girl. I want you, this Christmas month, to give it
away to the Lord Jesus Christ.
No matter if you have done it before. I am glad to know that
many of you have. Just renew the gift. Choose some quiet hour, and
go alone, and, on your knees, say: “Lord Jesus, I give myself anew
to Thee, in return for Thy great Christmas gift to me. I give Thee my
time, and my strength and my will. I ask Thee to direct me all day,
and every day, in the way in which Thou wouldst have me go. And I
promise to use my tongue to speak for Thee, and my hands to work
for Thee, and my feet to do errands for Thee, and my heart to love
Thee.”
I wonder how many will use this prayer, and mean each word in
their hearts? Just so many will be sure to have a happy Christmas,
and a happy year.
One other thing: Some of our Blossoms have been transplanted
during the year that is past. God has sent his angels and gathered
them to bloom in His upper garden, where flowers never wither.
Now, while we are glad for them, shall we not remember the homes
from which they have gone? The dear fathers and mothers and
brothers and sisters, who cannot, sometimes, keep the tears from
coming, because they miss their darlings so? Will not every Pansy
Blossom ask the dear Lord to help and comfort these sorrowing
hearts?
Now, just one thing more: What can you do on Christmas Day
which will make somebody else happier than he or she could
possibly have been if you had not done it?
Think it out, my Blossoms, “something for each of you now to
do.” Then, set to work and do it; then write and tell me all about it,
and I’ll weave all your letters into a story for next year’s Pansy.
Lovingly,
Pansy.

Annie and Florence. No doubt your game, “Jack Throws,” is a

good one but as it has a diagram, and as our printer cannot always
prepare such, it may fail to appear. Sorry.
Alice L. Snow. Send a copy of your Queer Story for examination.
You did capitally on the geographical puzzle. “A Christian since
eleven years of age.” So glad I am of this. I trust the Saviour is glad,
too. Is it your constant aim to make Him glad?
Myrtie A. B. Your Queer Story almost, not quite, correct. Don’t be
discouraged. The effort has done you great good, making you a
better speller. And you don’t have to go “three miles” to the Queer
Story school. You must teach your puss better manners when you
are working at your lessons.
Mamie Fuller. You must keep some of the writing you do now
while you are six years old, and put it by the side of what you do
when you are sixty. I guess you will see great progress. And I hope
you will come very near the Lord Jesus in that time.
C. R. Richmond. A picnic in a beautiful grove; five hundred
present; a brass band; oranges, ice-cream, lemonade; talks by Revs.
King and Adams; a bountiful dinner following, etc., etc. It seems as
though Master Colin and his companions must have gone home with
sunny hearts and faces, and in love with the S. S. Did you?
Minnie Locket. When a dear little Pansy writes as well as ever he
or she can, they need not ask me to excuse their penmanship or
anything else about the writing. All I care to know is that they do
their very best. No one can do more, and so there is no room for an
excuse. Do your best, Minnie, every time, and you will not be
ashamed.
Jessie P. Davis. Whenever your Pansies fail to come, write to D.
Lothrop Co., 32 Franklin Street, Boston, Mass. Do not think that any
Pansy can be so small as not to be missed. The baby is the smallest
body in the house. Don’t you think it would be missed?
H. C. Withey. The monkey game you send must be very funny,
but its length and the difficulty of “doing” it here will explain why it
does not appear in the Pansy. Perhaps you will try again, and send
something very short and easily played. The Pansies will be
interested in anything of that sort from Africa.
 Lolo Keeling. Don’t despair whatever may come. Carry your
failures to Jesus. Try again. Triumph will come.
M. Nellie Lindsey. Thank you for remembering to write to me
many times, even if you did “forget to send them off.” And what in
the August magazine pleased you so much besides that letter?
Fannie W. Ambler. Let me commend you for plain writing. The
Queer Story is well done, though with some mistakes. Try again, my
dear.
M. Lillie Read. Your Queer Story is much like Fannie’s, almost, but
not quite, right. Never mind; there’s another for you. Don’t be afraid
of it. Don’t be cast down over any failure, but up and at it again.
Train the baby to be a true Pansy. I’m so glad you make any
progress.
Nellie Wright. “Nine years old.” Said and done a thousand things
or more in that time. Now, suppose you live to be ninety,—my dear
uncle has just died at ninety-two,—and you keep on saying and
doing, then how many thousand will it be? But all the better if it be
all for Jesus’ sake. I like Florida, and you must like Kansas, which
gives you “lots of corn and fruit.”

BIBLE READING FOR DECEMBER.

(Christ.)

Dec. 1. Ps. xxiv: 7-10; 1 Cor. ii: 8.

“ 2. Mi. v: 2; Matt. ii: 3-6.
“ 3. Is. lx: 6; Matt. ii: 7-12.
“ 4. Hos. xi: 1; Matt. ii: 13-15.
“ 5. Jer. xxxi: 15; Matt. ii: 16-18.
“ 6. Is. xi: 2 and xlii: 1; Matt. 3: 13-17.
“ 7. Is. ix: 6; Luke ii: 11.
“ 8. Is. liii: 4; Matt. viii: 16, 17
“ 9. Is. liii: 5; 1 Pet. ii: 24.
“ 10. Is. liii: 7; Matt. xxvii: 12-14.
“ 11. Is. liii: 9; Matt. xxvii: 57-60.
“ 12. Is. liii: 10; Luke iii: 6.
“ 13. Matt. iv: 18-22.
“ 14. Matt. iv: 23-25.
“ 15. Matt. ix: 18, 19, 23-26.
“ 16. Matt. xi: 25-30.
“ 17. 2 Cor. v: 14, 15, 17.
“ 18. Rom. viii: 1, 2, 17, 18.
“ 19. Rom. viii: 35, 37-39.
“ 20. Rom. xv: 1-3.
“ 21. Rom. v: 1-2.
“ 22. Rom. v: 7-9.
“ 23. John xviii: 33-40.
“ 24. Luke ii: 8-14.
“ 25. Luke ii: 15-20.
“ 26. John xix: 1-7.
“ 27. John xix: 8-12.
“ 28. John xix: 13-16.
“ 29. John xix: 17-22 and xx: 31.
“ 30. Mark xvi: 19; Lu. xxiv: 51; Acts i: 9.
“ 31. Acts i: 10, 11; Matt. 24: 42-46.
 EXTRACTS FROM PANSY LETTERS.
Dear Pansy:
From a tiny child, May, if told to do anything, would do it just a
little differently. If told to put a thing on the table, she would say, “I
dess will put it on lounge,” and when I answered, “No; on the table,”
she would skip along, laughing, “I dess will put it on chair.” But doing
a little differently from mamma’s way and God’s way will end in
many a heartache Pray that mother and child may yield impliably to
the dear Father’s will.
May and her Mamma.
P. S. May sends six cents for the organ, earned by washing and
wiping dishes.

Dear Pansy:
This is one of my compositions:
A PAPER OF NEEDLES.

First, a coil of steel wire, twenty-four inches long.

This cut through the middle by scissors into little
curved bundles, long enough for two needles.
Straighten and point them—on a grindstone. They are
pointed at both ends. They must now have their eyes
opened. Needles, like puppies and kittens, are born
blind. They are stamped with a heavy die that leaves
the print of two needles’ heads and eyes at the center
of the wire. Then the eyes are opened with a double
punch.
 Harry B. Hayes.

Dear Pansy:
My fault is that I am ready to give, but not to take advice. I am
president of a society. One member chooses another to write about;
others read or recite selections. I attend Shurtleff Grammar School.
Nellie F. Treat.

Dear Pansy:
When I read answers to letters, it seems to hit me. I will try to be
more patient with brother and sister. I am a church member, and I
very much wish my cousin were. It is hard sometimes to talk about
Christ. I talk about other things readily. Why?
Blanche E. Treat.

Dear Pansy:
As to how we spend our evenings: we play games and speak
pieces, and we have lots of books and papers. We like The Pansy.
Papa got The Pansy for us and we did not know it till we got the mail.
Mamma gave us “Young People at Home,” which you wrote, and
which we like much. Maud and Mabel Davis, my sisters, are writing;
so I thought I would. Mamma does not like to have me tease my
little brother Romie. I will try not to do so.
Alice.

Dear Pansy:
Badges received in time to organize. They are lovely. They will
help us to remember our faults. We meet at each other’s homes
weekly and read, sew, sing, pray, and play The Pansy games.
“Monteagle” is splendid. Hope the trip will help Dilly get well, and
she will help naughty Hart get better. What a good woman Mrs.
Hammond is to have such a bad boy. I guess he’ll come out right.
Mabel S. Kagey.

Dear Pansy:
I send a small sum for the organ. I sent sixty cents to D. Lothrop
Company, and received “A Girl’s Room.” I would not part with it for
twice that. Every girl should have it. The book gives me an idea
what to do with things. I am much interested in “Around the Family
Lamp.”
Emma Fisk.

Dear Pansy:
Our mission band is “The Cheerful Givers.” Mrs. Prof. Jewett is the
teacher. She is nice. We like her—at least I do, and we all think the
others ought to. We have mite boxes to put our pennies into, to buy
maps of the world. I am not failing to mind my badge.
Mabel Hicks.

Dear Pansy:
I am eight and cannot write well. I tried last week. The words
were hard to spell. I got tired. Tears came, so did mamma, and said,
“Wait, I will help you.” I have taken your paper since I was a bit of a
girl, and love it, and can hardly wait till it comes.
I have many, many, many naughty traits. Send me a badge. It
may help me. Mamma and my good auntie read The Pansy to me,
and they enjoy it as much as I. Papa died before I was three years
old. I want to be good “For Jesus’ sake.”
 Annie T. Dana.

Dear Pansy:
Your answer to my last did me a world of good. I have it in my
album, as one of my most precious treasures. I’m determined to
grow up a useful Christian woman—thanks to the “Whisper Motto”
and the influence of Pansy and my mother. Mamma has no ambition
for her children but that they become genuinely good. When we are
determined to be so, she wishes us to join some church. There are
six, and as merry and happy as the “Little Peppers” of whom dear
Margaret Sidney wrote so sweetly.
Blanche Crawford.

Dear Pansy:
I enclose five cents. I earned it for the organ fund by learning the
multiplication table.
Minnie Lockwood.

Dear Pansy:
We have almost finished a bed-quilt and are making holders to
sell; in that way to raise money to buy cotton and lining. Every week
a word is given out. We learn a Bible verse with that word, to repeat
at the next meeting.
Gertie Currier.

Dear Pansy:
Please print my letter, as it is the first I have written to any paper.
I got a good many Christmas gifts, among them a canary. I named
her Soldie. I want to join the P. S. My fault is getting angry. I mean
to try real hard and break that horrid fault. I would like a badge. I
think you are real lovely. I would like to see you. I am your constant
reader and loving little friend. No one helped me about my story.
Eliza W. Holland.

ELIZA’S STORY.
(Ella’s Lesson.)

Ella Smith was six years old. Her mother told her to
take care of the baby while she went up street. Ella
went into her mother’s room for the baby. The bureau
drawer being open, she thought she would take a
peep. Naughty Ella! in it lay three large oranges. How
good they looked! She said to herself, “I will just touch
them.” They felt so mellow she took two large bites.
When her mother came, she punished Ella. Never did
she peep into her mother’s drawer again.

Dear Pansy:
I am trying to overcome my fault, which is putting off, and not
starting the moment I am spoken to. I like the whisper motto. I have
your picture and think very much of it. I have a pansy bed, and
when I look at it, it makes me think of the Pansy Society.
Ula Cook.

Dear Pansy:
I think the games published in The Pansy are lovely. We children,
this spring, in our yard, planted some seeds, and in three days they
began to show themselves. We planted the seeds in June and now
they have buds on them.
Rita E. Boardman.
Dear Pansy:
I have five dolls, Mabel, Phœbe, Sallie, Mollie, and Nannie. I love
Phœbe the best. I have had her almost two years. I have two cats,
Mrs. Kitty Clyde and her son Tom. We have a dog, too. His name is
Nero.
Rosalie T. Canfield.

Dear Pansy:
Perhaps some of the Pansies would like to hear of my two little
horned toads, about an inch and one half long. I have them in a
wire-screen cage. I have a big black beetle in the cage with them,
and they don’t seem to like him very well. I feed them with cornmeal
and flies.
Ollie Cutter Clarke.

Dear Pansy:
Thank you ever so much for the badge. I think it has helped me
some, but I left it in Denver when I came to Leadville to spend the
summer.
Orlena Beggs.

Dear Pansy:
I lend my Pansies to my friends to read. I found ninety-eight
mistakes in the Queer Story of August Pansy.
Sadie M. Knight.
Dear Pansy:
Willie Hicks, a dear little fellow in Africa, six months only, has
gone to his heavenly home. Each Angola station has its
representative in Heaven; but with Melville Cox, we say, “Though a
thousand fall, let Africa be redeemed.” My bed is a bamboo frame, a
canvas cot laced to it, set on low horses. I am learning how to use
tools. I want to make a desk and case for my little library. I have no
Pansy books. If I had money they would be here soon.
Herbert C. Withey.

Dear Pansy:
I walked in my garden to-day;
Many wee faces looked up
From their shady retreat.
Some had eyes dark or blue;
Some, curls of golden hue;
Dressed were some in velvets rare,
Or quaint, gay frocks,
These babies dear.
I asked them to come and live with me;
Gayly they laughed,
“Pansies are we.”
E. Edwards.

Dear Pansy:
I am so ashamed of myself I don’t like to write to you. I beg your
pardon. I have not worn my badge much, and I have not written to
you. It is all because I am so neglectful. I will try to wear my badge
more.
Mamie Thompson.
Dear Mrs. Alden:
We have a beautiful location. Looking south, we see the little
valley with Keiser Creek, like a thread, bright and clear; on either
side, the rocky hills, with pines, separated by deep ravines. Away
beyond the foot hills, overtopping all, is the Yellowstone or Snowy
Range, lifting up their white tops on the hottest days, as if in
derision. Rocks are everywhere. “The Rockies” are rightly named.
Several miles down the river is “Pompey’s Pillar,” on the rocky face of
which is carved the name of Wm. Clark, 1806, the explorer. Not far
is the battle-ground, on which the gallant Custer and his men lost
their lives by the Indians.
Lue J. Roseau.

Dear Mrs. Alden:

The Carmans gave us a concert. The church was full. Every one
was delighted. They stayed all night at our house and sang. So we
had a free concert. It makes me think of an angel to look at Miss
Nellie while she sings. Davie is such a sweet, manly boy. I wish there
were more such boys. I don’t know of any. I like Mr. and Mrs.
Carman, too. They are all beautiful singers. Davie is fourteen, and
superintends a mission Sabbath-school. While reading Docia’s
Journal, I decided for Christ.
Edith M. Hillbrant.

Dear Pansy:
When your kind letter came, Auntie Alice Ferree was here from
Kansas City. She used to live in Greensburg, Ind., and knew you, and
wishes to be remembered. I showed my badge to Gov. St. John. He
sends kindest regards.
Lena Pugh.
Dear Pansy:
I counted one hundred and sixty-six mistakes in the Queer Story.
I have written to ask you to write a letter and put it in The Pansy. I
could not find A Sevenfold Trouble, in one of the Pansies. Call me
Aurelia from Mass. There are so many Lizzies.
Lizzie A. Potter.

Dear Pansy:
I found one hundred and fifty-seven mistakes in the Queer Story.
[Send a copy corrected.—Ed.] I have two pets, a bird and a kitten.
The kitten is black, yellow, and white. Her name is Pansy. My bird is
cardinal; his name, Mac. Good-by!
Mabel Dynam.

E. Smith’s letter, with badge, sent to Roberts, Ill., has returned.

A letter signed Jonnae J., 2617, Park Ave., St. Louis, Mo.,
received.
Henry P. Austin’s letter, sent to South Paris, Me., with badge,
comes back.
S. H. Sterling writes from Philadelphia, Pa., without street and
number.
The above persons should immediately write, giving full name,
street, and number, and write very plainly.
 THE QUEER STORY.

P RESCILA ALDON lived in a verry valuble pavilian in Tenessee. She

was a buisy gerl, driveing her Shettling pones from hous too
hous and carying lillies, raisens, robbins, rabits, aggs and menny
other plesent an prety presants two sutch as wer in nead.
She wasent a fraid nuther off takeing a litle troubble whar thar
was opertunity of makeing her nabours hapy.
She desided to oregonize a socity of Pansys. Consequensely she
was going an comming hear an thare verry ofen and writeing leters,
hopeing two recieve manny menbers like other socitys. She securred
meny honary nembers two. So she ocupied herself til Febwary, wen
the aniversary of the socity came. Then it was reccomended that the
anniversity should be selebrated by an entetainment of adreses,
resiting peaces, musec, bone fiers, ice creems, punkin pise and
chickins and that all Pansys should ware thare bages and collors and
that no teaseing or plaging boy be aloud too com til he promiced too
lieve everry folt to home an bring towells and sope to cleen the
durty dishes.
Sence sum wouldent concent, thay wer not permited two com.
But the selebration was a grate sucess an Prescila was hapy.
[Mabel L. Thomas and Maude Lincoln have
mastered the mistakes of the Queer Story.]
 THE STENOGRAPH.

A CONTRIVANCE, by M. M. Bartholomew of New York, weighing

about three pounds, costing forty dollars, about the size and
shape of a man’s hand, with five different keys and a roll of narrow
paper, something like a spool of silk, designed for “fast writing,” is
the Stenograph; so named, because that is the meaning of the word
in Greek.
This queer little creature can make its mark, that’s all. But it can
do it so quickly and put it in so many different places and orders
when its fingers are touched by wire and spry human fingers that it
can write all your pen can write and do it in a fraction of the time,
while the operator’s eyes are looking away into the face of the
speaker whose words are being taken down.
Then, you can learn to “make it talk” in three months!
Suppose, now, some of you Pansies manage in some way to own
a Stenograph and teach the dear to mind the moment you speak to
it. It would be a delight to you to play upon it as upon your piano.
Besides, it would probably make music for your pocket. There is
money in it. You would be in demand at conventions and other
places as a reporter of speeches and sermons. And you could
command good pay, if you were a good stenographer.
Who knows but some of you will get your “bread and butter” by
means of this queer little creature?
C. M. L.
 NEW BOOKS.
(Published by D. Lothrop Company.)
True Manliness. By Thomas Hughes. Price $1. I want to say a word
about this book to a certain dear little girl I know who has a whole
dollar with which to buy papa’s Christmas present, and who cannot
decide what to buy. She knows papa likes books better than almost
anything else, but then, how should a young girl know what book to
select for a man who has gray hair?
My darling, I feel sure papa would like this very book. It is the
reason I selected it from a large number of others, to tell the Pansies
about, because I thought of you and your dollar. A very pretty book
in a deep maroon binding, with three hundred pages of reading
matter, and all of it put in paragraphs so that a busy man, who has
but a few minutes to read, can seize this volume, get some sweet
and helpful thoughts from it, during the five minutes when he is
waiting for the mail, or for the street-car, and then go on his busy
way. Just the thing for your hurried papa, don’t you see? In fact the
book belongs to a set, named “The Spare Minute Series,” gotten up
for just such waitings as I have described.
“Would the book be ‘nice’ for you to read?”
Well, no, little girlie, I’m inclined to think it would be rather
“grown-up” for you, because, you see, it was written to help your
grown-up papa and mamma. There are bits in it that you would like;
and your fifteen-year-old brother would often read words in it that
would please and help him. I open the book at random, and find
these words under the heading of “Courage.”
“After all, what would life be without fighting, I should like to
know? From the cradle to the grave, fighting, rightly understood, is