Microarrays for an Integrative Genomics

Microarrays for an Integrative Genomics 1

Foreword 2

Preface 3

Acknowledgments 4

Chapter 1: Introduction 7

1.1 The Future Is So Bright 7

1.2 Functional Genomics 9

1.2.1 Informatics and advances in enabling technology 12

1.2.2 Why do we need new techniques? 14

1.3 Missing the Forest for the Dendrograms, or One Aspect of Integrative Genomics 15

1.3.1 Sociology of a functional genomics pipeline 18

1.4 Functional Genomics, Not Genetics 19

1.4.1 In silico analysis will never substitute for in vitro and in vivo 21

1.5.1 Biological caveats in mRNA measurements 23

1.5.2 Sequence-level genomics 27

1.5.3 Proteomics 29

1.5 Basic Biology 29

Chapter 2: Experimental Design 32

2.1 The Safe Conception of a Functional Genomic Experiment 32

2.1.1 Experiment design space 32

2.1.2 Expression space 34

2.1.3 Exercising the expression space 38

2.1.4 Discarding data and low-hanging fruit 44

2.2 Gene-Clustering Dogma 48

2.2.1 Supervised versus unsupervised learning 49

2.2.2 Figure of merit: The elusive gold standard in functional genomics 51

Chapter 3: Microarray Measurements to Analyses 55

3.1 Generic Features of Microarray Technologies 55

3.1.1 Robotically spotted microarrays 58

3.1.2 Oligonucleotide microarrays 60

3.2 Replicate Experiments, Reproducibility, and Noise 70

3.2.1 What is a replicate experiment? A reproducible experimental outcome? 71

3.2.2 Reproducibility across repeated microarray experiments: Absolute expression level and fold difference 72

3.2.3 Cross-plat form (technology) reproducibility 75

3.2.4 Pooling sample probes and PCR for replicate experiments 76

3.2.6 Sources and examples of noise in the generic microarray experiment 78

3.2.7 Biological variation as noise: The Human Genome Project and irreproducibility of expression measurements 86

3.2.8 Managing noise 87

3.3 Prototypical Objectives and Questions in Microarray Analyses 90

3.3.1 Two examples: Inter-array and intra-array 91

3.4 Preprocessing: Filters and Normalization 93

3.4.1 Normalization 94

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