Gene Editing, Epigenetic, Cloning and Therapy

By Amin Elsersawi Ph.D.

This book is really helpful for someone who wants to start learning about genes and DNA. It is a well-written book describing all the introductory materials one would need to become current with genomes and genomics topics. It begins with an introduction to DNA and genes in chapter 1 and goes on from there through epigenetic in chapter 2, including acetylation, methylation, ubiquitylation of protein, deimination, and proline isomerization. It goes through gene editing in chapter 3, which includes good description of TALENs, ZFNs, and CRISPR/Cas systems. Chapter 4 includes cloning using artificial embryo twinning, somatic cell nuclear transfer, and asexual reproduction. Chapter 5 is about the material on basic stem cells of embryonic stem cells and adult stem cells. Chapter 6 discusses techniques and technology of gene therapy and cloning therapy. Chapter 7 includes descriptions on cell division, mitosis, meiosis, biological life cycle, parthenogenesis, bacterial conjugation, DNA fingerprints, genetic relationship between individuals and surname studies.

The book includes many diagrams and a glossary and an index. For a serious book on DNA and genes, this book is quite readable. It is a user-friendly textbook so that many readers will find it helpful to read some chapters more than once. The book is a valuable introduction to the extremely important field of genes and genomics.

• Language: English
• Publisher: AuthorHouse
• Release date: Aug 5, 2016
• ISBN: 9781524621988

Amin Elsersawi Ph.D.

Amin Elsersawi is a citizen of Canada since 1985. He received his Ph.D. degree in electrical engineering with emphasis in power electronic from Bradford University, U.K. He is a professional engineer registered with the Professional Engineering Society of Ontario Canada Dr. Elsersawi is currently retired. He previously served as general director for notable power generation and distribution energy utility. Prior to that, he was a chief of electrical engineering for the Public Work and Government Services Canada. He published more than 100 papers and reports in engineering and mathematic, biology, astronomy, and chemistry. He spent more than 15 years in teaching at universities and colleges. His main research interests are twentieth-century engineering, astronomy and chemistry. He is the author of the book Chemistry, Biology and Cancer: the Bond.

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© 2016 Amin Elsersawi. All rights reserved.

No part of this book may be reproduced, stored in a retrieval system, or transmitted by any means without the written permission of the author.

Published by AuthorHouse 08/04/2016

ISBN: 978-1-5246-2199-5 (sc)

ISBN: 978-1-5246-2198-8 (e)

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Because of the dynamic nature of the Internet, any web addresses or links contained in this book may have changed since publication and may no longer be valid. The views expressed in this work are solely those of the author and do not necessarily reflect the views of the publisher, and the publisher hereby disclaims any responsibility for them.

Contents

Introduction

CHAPTER 1: DNA and GENES

1.1 Chromosomes

1.1.1 DNA and RNA

1.1.2 Genes and Genomes

1.1.3 Alleles

1.2 Protein Synthesis

1.3 Coding

1.4 Control Gene Expression

1.5 Gene Silencing

1.5.1 Gene silencing and aging

1.6 Replication and Cell Division

1.7 Sister and nonsister chromatids

1.8 Molecule Inheritance

1.9 RNA Splicing

1.10 Gene Mutation

1.10.1 Causes

1.11 Gene Expression

1.12 Genetic Coding

1.13 Nucleic Acid Hybridation

1.13.1 Monohybrid and dihybrid cross

1.14 Gene/Chromosome Abnormalities

1.14.1 Genetic Disorders

1.15 Gene Therapy

1.15.1 Gene therapy and aging

1.15.2 Hayflick limit and gene therapy

1.16 Traits

1.16.1 Mendelian Inheritance

1.16.2 Molecule Inheritance

1.17 Inherited Traits

1.17.1 Acquired traits

1.18 Essential Genes

1.19 Genetic Polymorphism

CHAPTER 2: EPIGENETICS

2.1 Introduction

2.2 Molecular Basis of Epigenetics DNA Modification and Histone Modification

2.3 Histone Modification and Effect

2.3.1 Epigenetic Modifications and Mechanisms

2.3.2 Acetylation

2.3.3 Effect of Epigenetic and Genetic Alteration

2.3.4 Methylation

2.3.5 Histone Protein and Non Histone Protein

2.3.6 Mechanisms of DNA Methylation and Demethylation, DNA Replication

2.3.7 DNA Methylation and Cancer

2.3.8 Chromatin-Binding Proteins

2.3.9 Phosphorylation of proteins

2.3.10 Ubiquitylation of Protein

2.3.11 Sumoylation

2.3.12 ADP Ribosylation

2.3.13 Deimination

2.3.14 Biotinylation

2.3.15 Propionylation and Butyrylation

2.3.16 N-formylation

2.3.17 Proline Isomerization

CHAPTER 3: GENE EDITING

3.1 Transcription activator-like effector nucleases (TALENs)

3.1.1 TALE Structure

3.2 Zinc Finger Nucleases (ZFNs)

3.2.1 Why Zinc?

3.3 CRISPR/Cas system

3.4 Genetic Manipulation

3.4.1 Growth Hormone Deficiency

3.4.2 Blood clot factor VII

3.4.3 Hepatitis Virus (HV) (A and B strands)

3.4.4 Insulin production

3.4.5 Crop resistance to herbicide

3.4.6 Producing a new protein or enzyme

3.4.7 Introducing a novel trait (Canada)

CHAPTER 4: CLONING

4.1 Artificial embryo twinning

4.2 Somatic cell nuclear transfer

4.3 Asexual reproduction

4.3.1 Binary Fission

4.3.2 Budding

4.3.3 Vegetative Reproduction

4.3.4 Spore

4.3.5 Fragmentation

4.3.6 Agamogenesis

4.4 Alternation between Sexual and Asexual Reproduction

4.5 Inheritance of asexual reproduction in sexual species

4.6 Cloning

4.6.1 Artificial Cloning

4.6.2 Fusion Cell Cloning

4.6.3 Natural Cloning

4.7 Gene Cloning

4.7.1 Human Cloning

4.7.2 Ethical Issues of Cloning

4.7.3 Religious Views

4.7.4 Cloning extinct and endangered species

CHAPTER 5: STEM CELLS

5.1 Unique Properties of Stem Cells

5.2 Embryonic stem cells

5.3 Adult Stem Cells

5.4 Induced Pluripotent Stem Cells

5.5 Current Arguments and Counterarguments Regarding Human Reproductive Cloning

5.6 Summary

CHAPTER 6: GENE THERAPY AND CLONING THERAPY

6.1 Gene Therapy

6.2 Somatic Gene Therapy

6.3 Germline Gene Therapy

6.4 Cloning Therapy

6.5 Conclusion

CHAPTER 7: CELL BIOLOGY

7.1 Cell Division

7.1.1 Mitosis

7.1.2 Meiosis

7.1.3 Why Meiosis?

7.1.5 Biological life cycle

7.1.5.1 Parthenogenesis

7.2 Bacterial Conjugation

7.3 DNA Fingerprinting

7.4 Genetic Relationship between Individuals

7.4.1 Surname studies

7.4.2 Direct to consumer paternity testing

7.4.3 The genetic genealogy revolution

7.5 The Genographic Project

7.6 Human Genome Project

7.11 Frozen Ark

Glossary

INTRODUCTION

This book is really helpful for someone who wants to start learning about genes and DNA. It is well written book describing all the introductory materials one would need to become current with genomes and genomics topics. It begins with an introduction to DNA and genes in Chapter 1, and goes on from there through epigenetic in chapter 2, including acetylation, methylation, ubiquitylation of protein, deimination and proline isomerization. It goes through gene editing in chapter 3 which includes good description of TALENs, ZFNs and CRISPR/Cas systems. Chapter 4 includes cloning using artificial embryo twinning, somatic cell nuclear transfer, and asexual reproduction. Chapter 5 is the material on basic stem cells of embryonic stem cells and adult stem cells. Chapter 6 discusses techniques and technology of gene therapy and cloning therapy. Chapter 7 includes descriptions on cell division, mitosis, meiosis, biological life cycle, parthenogenesis, bacterial conjugation, DNA fingerprints, genetic relationship between individuals and surname studies.

The book includes many diagrams and glossary and an index at the front. For a serious book on DNA and genes this book is quite readable it is a user-friendly textbook, so that many readers will find it helpful to read some chapters more than once. The book is a valuable introduction to the extremely important field of genes and genomics.

A highly informative book, I recommend it highly to anyone interested in the subject."

CHAPTER 1

DNA and GENES

1.1 Chromosomes

In sexually reproducing organism two types of cell division are needed. One is for the processes of growth, repair and asexual reproduction and it is called mitosis. Mitosis produces daughter cells that are diploid and genetically identical to the parent cell.

When the organism wants to make gametes (eggs and sperms) a different mechanism is required. Gametes are not diploid like all the other body cells, but instead they only have one member of each homologous pair of chromosomes. In order to make a haploid daughter cell, a second type of cell division, meiosis, is needed, Figure (1).

Figure (1): Mitosis and meiosis

image002.jpg

Chromosomes are thread-like structures located inside the nucleus of a cell.. Each chromosome is made of protein and a single molecule of deoxyribonucleic acid (DNA). Passed from parents to offspring, DNA contains the specific instructions that make each type of living organism unique.

The nucleus of each cell in our bodies contains approximately 1.8 meters of DNA in total, although each strand is less than one millionth of a centimeter thick. This DNA is tightly packed into structures called chromosomes, which consist of long chains of DNA and associated proteins. In eukaryotes, DNA molecules are tightly wound around proteins – called histone proteins - which provide structural support and play a role in controlling the activities of the genes. A strand 150 to 200 nucleotides long is wrapped twice around a core of eight histone proteins to form a structure called a nucleosome, Figure (2).

Figure (2): Nucleosome, hisone and DNA

image004.jpg

The chromosomes - and the DNA they contain - are copied as part of the cell cycle, and passed to daughter cells through the processes of mitosis and meiosis.

Chromosomes end with two telomeres. If telomeres are shortened, cells and human will age. If telomerase activity is high, or if the telomeres are extended, chromosome (and DNA) is maintained, and cellular senescence is delayed. In contrast, if telomeres are damaged or defected, cancer and certain inherited disease may be developed. Scientists found that when a cell is about to divide, the DNA molecules, which contain the four bases that form the genetic code, are copied, base by base, by DNA polymerase enzymes. However, for one of the two DNA strands, a problem exists in that the very end of the strand cannot be copied. Therefore, the chromosomes should be shortened every time a cell divides, Figure (3).

Figure (3): Shortened DNA and thus chromosome due to damaged or shortened telomere

image006.jpg

Luckily, these problems were solved when this year’s Nobel Laureates (Elizabeth Blackburn, Jack Szostak, and Carol Greider) discovered how the telomere functions and found the enzyme that copies it.

During his experiment, Jack Szostak observed that a linear DNA molecule, a type of minichromosome, is rapidly degraded when introduced into yeast cells. In fact, all DNA molecules are degraded by time after replication, but putting the DNA molecules in yeast cells, they delay faster. The question is how to delay or prevent the degradation. If this is achieved, the span of life would be elongated and the aging process would be delayed.

Elizabeth Blackburn mapped DNA sequences. When studying the chromosomes of Tetrahymena, a unicellular ciliate organism, she identified a DNA sequence that was repeated several times at the ends of the chromosomes. The function of this sequence, CCCCAA (C for cytosine and A for adenine, both are nucleotides of the DNA), was unclear. Blackburn and Szostak decided to perform an experiment that would implement their discoveries jointly. From the DNA of Tetrahymena, Blackburn isolated the CCCCAA sequence. Szostak coupled it to the minichromosomes and put them back into yeast cells. Results were amazing - the telomere DNA sequence protected the minichromosomes from degradation, Figure (4). This was the start of the reversal of aging.

Figure (4): Szostack and Blackburn joint experiment

image008.jpg

Blackburn and her graduate student Carol Greider started to investigate if the formation of telomere DNA could be due to an unknown enzyme. Greider discovered signs of enzymatic activity in a cell extract. Greider and Blackburn named the enzyme telomerase. The telomerase consists of RNA nucleotides CCCCAA and protein. Telomerase extends telomere DNA, providing a platform that enables DNA polymerases to copy the entire length of the chromosome without missing the very the very end portion, Figure (5).

Figure (5): Effect of telomerase on cell division

image010.jpg

Telomeres safeguard the chromosome ends from DNA repair and degradation activities. With gradual shortening of the telomeres and chromosomes cells grew poorly and eventually stopped dividing. This could lead to premature cellular ageing – senescence. Scientists concluded that functional telomeres could prevent chromosomal damage and delay cellular senescence. They showed that the senescence of human cells is also delayed by telomerase.

1.1.1 DNA and RNA

Deoxyribonucleic acid or DNA is a part of chromosomes and includes information needed to develop all features of life. The DNA is found in every cell and passed down from generation to generation. DNA carries the code to control the characteristic in all life forms.

Deoxyribonucleic acid or DNA is a part of chromosomes and includes information needed to develop all features of life. The DNA is found in every cell and passed down from generation to generation. DNA carries the code to control the characteristic in all life forms.

DNA is a very long macromolcule that is the main component of chromosomes made up of molecules called nucleotides. Each nucleotide is composed of a nitrogen-containing nucleobase - either cytosine (C), guanine (G), adenine (A), or thymine (T)—as well as a sugar called deoxyribose and a phosphorate group. The nucleotides are joined to one another in a chain by covalent bonds between the sugar of one nucleotide and the phosphate of the next, resulting in an alternating sugar-phospate backbone. According to base pairing rules rules (A with T, and C with G), hydrogen bonds bind the nitrogenous bases of the two separate polynucleotide strands to make double-stranded DNA. Each DNA sequence that contains instructions to make a protein is known as a gene.

The entire human genome contains about 3 billion bases and about 20,000 genes.

DNA is usually a double-helix and has two strands running in opposite directions.. Each chain is a polymer of subunits called nucleotides (hence the name polynucleotide). Nucleotides are attached together to form two long strands that spiral to create a structure called a double helix. If you think of the double helix structure as a ladder, the phosphate and sugar molecules would be the sides, while the bases would be the rungs. The bases on one strand pair with the bases on another strand: adenine pairs with thymine, and guanine pairs with cytosine, Figure (6).

Figure (6): Structure of DNA

image012.jpg

Although most DNA is packaged in chromosomes within the nucleus, mitochondria also have a small amount of their own DNA. This genetic material is known as mitochondrial DNA or mtDNA. Mitochondrial DNA (mtDNA or mDNA) is the DNA located in mitochondria, cellular organelles within eukaryotic cells that convert chemical energy from food into a form that cells can use.

The number of mitochondria in a cell can vary widely by organism, tissue, and cell type. For instance, red blood cells have no mitochondria, whereas liver cells can have more than 2000

Mitochondria have been described as the powerhouse of the cell because they generate most of the cell’s supply of adenosine triphosphate (ATP), used as a source of chemical energy. In addition to supplying cellular energy, mitochondria are involved in other tasks, such as signaling, cellular differentiation, and cell death, as well as maintaining control of the cell cycle and cell growth.. Each cell contains hundreds to thousands of mitochondria, which are located in the fluid that surrounds the nucleus (the cytoplasm). The number of mitochondria in a cell can vary widely by organism, tissue, and cell type. For instance, red blood cells have no mitochondria, whereas liver cells can have more than 2000.

In addition to producing energy, mitochondria store calcium for cell signaling activities, generate heat, and mediate cell growth and death. The number of mitochondria per cell varies widely; for example, in humans, erythrocytes (red blood cells) do not contain any mitochondria, whereas liver cells and muscle cells may contain hundreds or even thousands.

In addition to energy production, mitochondria play a role in several other cellular activities. For example, mitochondria help regulate the self-destruction of cells (apoptosis). They are also necessary for the production of substances such as cholesterol and heme (a component of hemoglobin, the molecule that carries oxygen in the blood).

The human mitochondrial DNA (mtDNA) is a double-stranded, and contains 37 genes coding for two rRNAs, 22 tRNAs and 13 polypeptides. Mitochondrial DNA contains 37 genes, all of which are essential for normal mitochondrial