Introduction to Pharmaceutical Biotechnology: Basic Techniques and Concepts

by Saurabh Bhatia

Overview

Animal biotechnology is a broad field including polarities of fundamental and applied research, as well as DNA science, covering key topics of DNA studies and its recent applications. In Introduction to Pharmaceutical Biotechnology, DNA isolation procedures followed by molecular markers and screening methods of the genomic library are explained. Interesting areas like isolation, sequencing and synthesis of genes, with the broader coverage on synthesis of genes, are also described. The book begins with an introduction to biotechnology and its main branches, explaining both the basic science and the applications of biotechnology-derived pharmaceuticals, with special emphasis on their clinical use. It then moves on to historical development and scope of biotechnology with an overall review of early applications that scientists employed long before the field was defined.

Product Details

ISBN-13:	9780750313001
Publisher:	Iop Publishing Ltd
Publication date:	07/31/2018
Pages:	589
Product dimensions:	7.30(w) x 10.39(h) x 0.92(d)

Table of Contents

Preface xiv

Acknowledgments xv

Author biographies xvi

1 History, scope and development of biotechnology 1-1

1.1 Introduction 1-1

1.2 Branches of biotechnology 1-5

1.3 Biotechnology and its various stages of development 1-6

1.3.1 Old and new biotechnology 1-17

1.3.2 Ancient biotechnology (pre-1800) 1-17

1.3.3 Classical biotechnology 1-19

1.3.4 Modern biotechnology 1-21

1.4 Scope and importance of biotechnology 1-27

1.4.1 Biotechnology in medicine 1-27

1.4.2 Industrial biotechnology 1-28

1.4.3 Biotechnology and the environment 1-29

1.4.4 Biotechnology and agriculture 1-29

1.5 Biotechnology techniques 1-29

1.5.1 Bioreactors 1-29

1.5.2 Cell fusion 1-29

1.5.3 Liposome-based delivery 1-31

1.5.4 Cell or tissue culture 1-32

1.5.5 Genetic engineering 1-32

1.5.6 DNA fingerprinting 1-32

1.5.7 Cloning 1-32

1.5.8 Artificial insemination and ET technology 1-33

1.5.9 Stem cell technology 1-33

1.6 Applications of biotechnology 1-34

1.6.1 Basic applications of biotechnology 1-35

1.6.2 Most common applications 1-35

1.7 Biotech research: 2015-2016 1-47

References 1-57

2 Modern DNA science and its applications 2-1

2.1 Introduction 2-1

2.1.1 Genes: units of inheritance 2-1

2.2 The Human Genome Project 2-3

2.3 DNA synthesis begins at replication origins 2-4

2.4 Gene expression 2-4

2.4.1 One gene, one product 2-4

2.5 Structure of DNA 2-6

2.5.1 Basic structural features of DNA 2-7

2.6 DNA replication 2-7

2.6.1 Leading strand 2-8

2.6.2 Lagging strand 2-9

2.7 DNA supercoiling 2-9

2.8 Repair and recombination 2-9

2.9 Types of DNA damage 2-11

2.9.1 Base excision repair (BER) 2-13

2.9.2 Mismatch repair (MMR) 2-13

2.9.3 Nucleotide excision repair (NER) 2-14

2.9.4 Double-strand break (DSB) repair 2-14

2.10 DNA recombination 2-14

2.10.1 Types and examples of recombination 2-15

2.11 DNA isolation is the commencement of molecular marker analysis 2-16

2.11.1 DNA extraction protocols 2-17

2.12 Types of molecular markers 2-18

2.12.1 Hybridization-based molecular markers 2-18

2.12.2 PGR-based markers 2-22

2.13 Genomic library screening methods 2-34

2.13.1 Colony hybridization 2-34

2.13.2 Chromosome walking 2-35

2.13.3 Blotting techniques 2-36

2.13.4 Southern blot analysis 2-36

2.13.5 Northern blot analysis 2-36

2.13.6 Western blot analysis 2-37

2.13.7 Dot blot technique 2-37

2.13.8 Techniques for the detection of specific proteins 2-38

2.13.9 Electrophoresis techniques 2-38

2.14 Triplex DNA, TFOS, PNAs, RNA-DNA hybrids and DSRNA/RNAI 2-44

2.15 Isolation, sequencing and synthesis of genes 2-46

2.15.1 Isolation of ribosomal RNA 2-46

2.15.2 Isolation of genes coding for specific proteins 2-46

2.15.3 Isolation of genes (with known or unknown products) using DNA or RNA probes 2-47

2.15.4 Isolation of genes (with known or unknown products) using DNA or RNA probes 2-48

2.15.5 Use of transposable elements (transposon tagging) 2-49

2.15.6 T-DNA insertion mutagenesis for isolation of plant genes 2-50

2.15.7 Promoter, enhancer and gene trap for isolation of genes 2-50

2.15.8 Mutation complementation 2-51

2.15.9 Differential screening and differential display technique for isolation of genes 2-51

2.15.10 Subtractive hybridization for gene isolation 2-52

2.15.11 Map-based cloning for gene isolation 2-52

2.15.12 Isolation of novel genes 2-52

2.15.13 Sequencing of a gene or a DNA fragment 2-53

2.16 Synthesis of genes 2-54

2.16.1 Chemical synthesis of tRNA genes 2-54

2.16.2 Synthesis of the gene for yeast alanyl tRNA 2-56

2.16.3 Synthesis of a gene from true precursor tRNA 2-56

2.16.4 Mass spectrometry for genomics and proteomics 2-56

2.17 Genomics and proteomics research 2-60

References 2-61

3 Introduction to genetic engineering 3-1

3.1 Introduction 3-1

3.2 Gene transfer technologies 3-2

3.2.1 Electro poration 3-2

3.2.2 Microinjection 3-3

3.2.3 Biolistics or microprojectiles for DNA transfer 3-3

3.2.4 Liposome-mediated gene transfer 3-4

3.2.5 Calcium-phosphate-mediated DNA transfer 3-4

3.2.6 DNA transfer by DEAE-dextran method 3-5

3.2.7 Transfer of DNA by polycation-DMSO 3-6

3.2.8 Poly ethylene-gly col-mediated transfection 3-7

3.2.9 Gene transfer through peptides 3-7

3.2.10 Gene transfer by retroviruses 3-8

3.3 Plasmids 3-9

3.4 Different hosts and protein expression technologies 3-10

3.4.1 rDNA technology 3-12

3.5 Gene cloning 3-15

3.6 Transfection methods and transgenic animals 3-16

3.6.1 Gene transfer or transfection 3-17

3.6.2 Transfection of fertilized eggs or embryos 3-17

3.6.3 Transfer of whole nuclei (or split embryos) 3-17

3.6.4 DNA microinjection into the egg 3-18

3.6.5 Virus-mediated gene transfer to embryo 3-19

3.6.6 Transfection of cultured mammalian cells 3-20

3.6.7 Targeted gene transfer 3-21

3.6.8 Transgenic animals in biotechnology 3-21

3.7 Applications of genetic engineering in biotechnology 3-30

3.8 Mammalian cell line characterization 3-32

3.9 In vitro fertilization (IVF) and embryo transfer in humans and domestic animals 3-33

3.10 IVF in humans and embryo transfer 3-34

3.10.1 Types and causes of infertility 3-34

3.10.2 Evaluation and assessment of patients 3-37

3.10.3 IVF fertility treatment 3-38

3.10.4 Development of ovarian follicles in natural menstrual cycles 3-39

3.10.5 Development of ovarian follicles in stimulated cycles 3-39

3.10.6 Development of ovarian follicles during a controlled cycle 3-40

3.10.7 Ovarian stimulation protocols for IVF 3-40

3.10.8 Spontaneous luteinizing hormone (LH) surge 3-43

3.10.9 Administration of hCG for controlled ovulation 3-43

3.10.10 Equipment and technique for laparoscopy 3-44

3.10.11 Oocyte culture and IVF culture of oocytes 3-45

3.10.12 Preparation of semen 3-46

3.10.13 In vitro fertilization 3-47

3.11 Embryo transfer (ET) in humans 3-48

3.11.1 Time of ET 3-48

3.31.1 ET technique 3-48

3.12 Superovulation, IVF and embryo culture in farm animals 3-50

3.13 ET in cattle 3-52

3.13.1 ET technique in cattle 3-52

3.13.2 Technique for freezing embryos in cattle 3-54

3.13.3 Benefits of ET in cattle 3-55

References 3-55

4 Applications of stem cells in disease and gene therapy 4-1

1.1 Introduction

4.2 Types of gene therapy 4-2

4.2.1 Somatic gene therapy 4-2

4.2.2 Gennline gene therapy 4-3

4.3 Gene therapy strategies 4-3

4.3.1 Gene augmentation therapy (GAT) 4-3

4.3.2 Targeted killing of specific cells 4-4

4.3.3 Targeted inhibition of gene expression 4-5

4.3.4 Targeted gene mutation correction 4-5

4.4 Methods of gene therapy 4-5

4.1.1 Ex vivo gene therapy 4-6

4.1.2 In vivo gene therapy 4-7

4.5 Gene therapy approaches 4-8

4.5.1 Conventional gene therapy 4-10

4.5.2 Non-classical gene therapy 4-10

4.6 Vectors for gene therapy 4-10

4.6.1 Viral-mediated gene delivery 4-10

4.6.2 Non-viral-mediated gene therapy 4-11

4.7 Target sites for gene therapy 4-19

4.7.1 Target cells for gene transfer 4-19

4.8 Gene therapy strategies for cancer 4-20

4.8.1 Tumor necrosis factor gene therapy 4-20

4.8.2 Suicide gene therapy 4-20

4.8.3 Two gene cancer therapy 4-21

4.8.4 Gene replacement therapy 4-21

4.9 Gene therapy for AIDS 4-22

4.9.1 REV and ENV genes 4-22

4.9.2 Genes of HIV proteins 4-22

4.9.3 Gene to inactivate gpl20 4-22

4.10 Oligonucleotide therapies: anttgene and antisense therapy 4-22

4.10.1 Antisense therapy for cancer 4-23

4.10.2 Antisense therapy for AIDS 4-25

4.11 Antisense oligonucleotides as therapeutic agents 4-26

4.12 Chimeric oligonucleotides in gene correction 4-27

4.13 Aptamers as therapeutic agents 4-28

4.14 Ribozymes as therapeutic agents 4-29

4.15 The future of gene therapy 4-29

4.16 Stem cell research 4-30

4.16.1 Stem cell classification 4-30

4.16.2 Historical background of stem cell research 4-32

4.16.3 Ethical issues associated with cell lines 4-33

4.16.4 Applications of stem cell research 4-33

4.16.5 Human ES ceils 4-34

References 4-35

5 Transgenic animals in biotechnology 5-1

5.1 Introduction 5-1

5.2 Major objectives of gene transfer 5-2

5.3 Cloning vectors 5-2

5.3.1 Fish vectors in molecular genetics and biotechnology 5-5

5.3.2 P element vectors/transposon 5-5

5.3.3 Baculovirus as versatile vectors for protein expression in insects 5-7

5.4 Efficient and versatile mammalian virus vectors 5-8

5.4.1 SV40 vectors 5-8

5.4.2 BPV (bovine papillomavirus) DNA vectors 5-14

5.4.3 Retrovirus vectors and their use 5-14

5.4.4 Vaccinia virus vectors: new approach for producing recombinant vaccines 5-16

5.4.5 Adenovirus vectors (for gene therapy, vaccination and cancer gene therapy) 5-16

5.5 Mammalian artificial chromosome (MAC) vectors for somatic gene therapy 5-17

5.6 DNA constructs 5-18

5.6.1 Promoter sequences 5-19

5.6.2 Selectable reporter or marker genes 5-19

5.7 Transfection methods: an approach towards mammalian cell transfection 5-24

5.7.1 Calcium phosphate-mediated transfection of eukaryotic cells 5-24

5.7.2 Transfection using DEAE-dextran 5-25

5.7.3 Lipofection (lipid-mediated DNA transfection method) 5-26

5.7.4 Bacterial protoplast fusion 5-28

5.7.5 Gene transfer by electroporation 5-28

5.7.6 Retrovirus-mediated gene transfer 5-30

5.7.7 Basics of DNA microinjection 5-31

5.8 Xenopus oocytes as a heterologous expression system 5-34

5.9 ES cell-mediated gene transfer 5-35

5.10 Targeted gene transfer or gene therapy in mammals 5-37

5.10.1 Gene disruption by. HR in mammals 5-39

5.10.2 Gene targeting/replacement 5-40

5.11 Transgene integration, organization and expression 5-45

5.12 Transgene recovery in mammahan cells 5-46

5.13 Cloned protein expression in mammalian cells 5-48

5.13.1 Expression vectors for mammalian cells 5-50

5.13.2 Enhanced production of recombinant proteins 5-56

5.13.3 Scale-up of protein purification (stages in downstream processing) 5-60

References 5-63