Lecture 1 - Molecular diagnosis of genetic diseases | Biotechnology (Zoology) by ILLL, DU - Biotechnology Engineering (BT) PDF Download

 Page 1


Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
 
 
Lesson Prepared Under MHRD project “National 
Mission on Education Through ICT” 
 
Zoology 
Biotechnology 
Lesson: Molecular diagnosis of genetic diseases 
(Cystic fibrosis, Huntington’s disease, Sickle cell 
anemia) 
Lesson Developer: Dr. Jaspreet Kaur 
College/Dept: S.G.T.B. Khalsa College 
University of Delhi 
Lesson Reviewer: Dr. Ravi Toteja 
College/Dept: Zoology, Acharya Narendra Dev 
College 
University of Delhi 
 
 
 
 
 
 
 
Page 2


Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
 
 
Lesson Prepared Under MHRD project “National 
Mission on Education Through ICT” 
 
Zoology 
Biotechnology 
Lesson: Molecular diagnosis of genetic diseases 
(Cystic fibrosis, Huntington’s disease, Sickle cell 
anemia) 
Lesson Developer: Dr. Jaspreet Kaur 
College/Dept: S.G.T.B. Khalsa College 
University of Delhi 
Lesson Reviewer: Dr. Ravi Toteja 
College/Dept: Zoology, Acharya Narendra Dev 
College 
University of Delhi 
 
 
 
 
 
 
 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
Table of Contents 
 
? Introduction  
? Molecular diagnosis of Genetic diseases 
? Technology of Molecular Diagnostics 
? Cystic Fibrosis or mucoviscidosis 
? Symptoms and Causes 
? Pathogenesis 
? Structure of CFTR 
? Function of CFTR 
? Common Mutations leading to Cystic 
Fibrosis 
? Methods used in CFTR testing 
? Huntington’s Disease 
? Symptoms and Causes 
? Structure of Huntingtin 
? Mutation that causes Huntingtin’s 
disease 
? Methods used in testing Huntington’s 
Disease 
? Sickle Cell Anaemia 
? Symptoms and Causes 
? Structure of Hemoglobin 
? Mutation which causes Sickle cell anemia 
? Methods used in testing Sickle Cell 
Anaemia 
 
? Summary 
? Exercise/ Practice 
? Glossary 
? References/ Bibliography/ Further Reading 
Page 3


Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
 
 
Lesson Prepared Under MHRD project “National 
Mission on Education Through ICT” 
 
Zoology 
Biotechnology 
Lesson: Molecular diagnosis of genetic diseases 
(Cystic fibrosis, Huntington’s disease, Sickle cell 
anemia) 
Lesson Developer: Dr. Jaspreet Kaur 
College/Dept: S.G.T.B. Khalsa College 
University of Delhi 
Lesson Reviewer: Dr. Ravi Toteja 
College/Dept: Zoology, Acharya Narendra Dev 
College 
University of Delhi 
 
 
 
 
 
 
 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
Table of Contents 
 
? Introduction  
? Molecular diagnosis of Genetic diseases 
? Technology of Molecular Diagnostics 
? Cystic Fibrosis or mucoviscidosis 
? Symptoms and Causes 
? Pathogenesis 
? Structure of CFTR 
? Function of CFTR 
? Common Mutations leading to Cystic 
Fibrosis 
? Methods used in CFTR testing 
? Huntington’s Disease 
? Symptoms and Causes 
? Structure of Huntingtin 
? Mutation that causes Huntingtin’s 
disease 
? Methods used in testing Huntington’s 
Disease 
? Sickle Cell Anaemia 
? Symptoms and Causes 
? Structure of Hemoglobin 
? Mutation which causes Sickle cell anemia 
? Methods used in testing Sickle Cell 
Anaemia 
 
? Summary 
? Exercise/ Practice 
? Glossary 
? References/ Bibliography/ Further Reading 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
  
 
 
Introduction 
A genetic disorder can be defined as any mutation in gene which changes its 
instructions to make protein or improper functioning of the protein or its complete 
absence, is termed as Moreover, these disorders are categorized as: 
? Single-gene disorders: where a mutation affects only one gene. 
Example, Sickle cell anemia. 
? Chromosomal disorders: where chromosomes (or parts of 
chromosomes) are missing or changed. Example, Down syndrome.  
? Complex disorders: where mutations affect two or more genes. 
Example, Colon cancer. 
? Mitochondrial disorders: where mutations affect the non-chromosomal 
DNA of mitochondria. Example, an eye disease called Leber's hereditary 
optic atrophy; a type of epilepsy called MERRF which stands for Myoclonus 
Epilepsy with Ragged Red Fibers. 
 
In comparison to the infectious diseases, genetic disorders represent a major 
health problem since they may develop in later life and are usually incurable. 
Thus, molecular diagnosis of these disorders, especially at the prenatal stage 
becomes extremely important to prevent their transmission to next generation. 
Molecular Diagnosis of Genetic Diseases 
In order to efficiently deliver effective care to a patient, the first and foremost 
thing for a health care practitioner is the ability to accurately identify the cause of 
patient‘s problem, i.e., make the diagnosis. 
Now, as the name suggests, ?Molecular diagnostics? is a comprehensive term 
defining different diagnostic tests that assess a person‘s health literally at a 
molecular level, i.e., detecting and measuring specific genetic sequences (also 
called biological markers or biomarkers) in deoxyribonucleic acid (DNA) or 
ribonucleic acid (RNA) or the proteins they express. Molecular diagnostics help to 
determine whether a specific person is predisposed to have a disease, or actually 
have a disease, or whether a certain treatment option is likely to be effective for 
a specific disease. 
In the nineteenth century, before the advent of molecular diagnostics, the only 
available tools were microscopy and histopathology or cellular pathology. For 
Page 4


Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
 
 
Lesson Prepared Under MHRD project “National 
Mission on Education Through ICT” 
 
Zoology 
Biotechnology 
Lesson: Molecular diagnosis of genetic diseases 
(Cystic fibrosis, Huntington’s disease, Sickle cell 
anemia) 
Lesson Developer: Dr. Jaspreet Kaur 
College/Dept: S.G.T.B. Khalsa College 
University of Delhi 
Lesson Reviewer: Dr. Ravi Toteja 
College/Dept: Zoology, Acharya Narendra Dev 
College 
University of Delhi 
 
 
 
 
 
 
 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
Table of Contents 
 
? Introduction  
? Molecular diagnosis of Genetic diseases 
? Technology of Molecular Diagnostics 
? Cystic Fibrosis or mucoviscidosis 
? Symptoms and Causes 
? Pathogenesis 
? Structure of CFTR 
? Function of CFTR 
? Common Mutations leading to Cystic 
Fibrosis 
? Methods used in CFTR testing 
? Huntington’s Disease 
? Symptoms and Causes 
? Structure of Huntingtin 
? Mutation that causes Huntingtin’s 
disease 
? Methods used in testing Huntington’s 
Disease 
? Sickle Cell Anaemia 
? Symptoms and Causes 
? Structure of Hemoglobin 
? Mutation which causes Sickle cell anemia 
? Methods used in testing Sickle Cell 
Anaemia 
 
? Summary 
? Exercise/ Practice 
? Glossary 
? References/ Bibliography/ Further Reading 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
  
 
 
Introduction 
A genetic disorder can be defined as any mutation in gene which changes its 
instructions to make protein or improper functioning of the protein or its complete 
absence, is termed as Moreover, these disorders are categorized as: 
? Single-gene disorders: where a mutation affects only one gene. 
Example, Sickle cell anemia. 
? Chromosomal disorders: where chromosomes (or parts of 
chromosomes) are missing or changed. Example, Down syndrome.  
? Complex disorders: where mutations affect two or more genes. 
Example, Colon cancer. 
? Mitochondrial disorders: where mutations affect the non-chromosomal 
DNA of mitochondria. Example, an eye disease called Leber's hereditary 
optic atrophy; a type of epilepsy called MERRF which stands for Myoclonus 
Epilepsy with Ragged Red Fibers. 
 
In comparison to the infectious diseases, genetic disorders represent a major 
health problem since they may develop in later life and are usually incurable. 
Thus, molecular diagnosis of these disorders, especially at the prenatal stage 
becomes extremely important to prevent their transmission to next generation. 
Molecular Diagnosis of Genetic Diseases 
In order to efficiently deliver effective care to a patient, the first and foremost 
thing for a health care practitioner is the ability to accurately identify the cause of 
patient‘s problem, i.e., make the diagnosis. 
Now, as the name suggests, ?Molecular diagnostics? is a comprehensive term 
defining different diagnostic tests that assess a person‘s health literally at a 
molecular level, i.e., detecting and measuring specific genetic sequences (also 
called biological markers or biomarkers) in deoxyribonucleic acid (DNA) or 
ribonucleic acid (RNA) or the proteins they express. Molecular diagnostics help to 
determine whether a specific person is predisposed to have a disease, or actually 
have a disease, or whether a certain treatment option is likely to be effective for 
a specific disease. 
In the nineteenth century, before the advent of molecular diagnostics, the only 
available tools were microscopy and histopathology or cellular pathology. For 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
example, clinicians categorized cancer cells according to their pathology, that is, 
according to their appearance under a microscope. Despite being the first line 
tools for quickly identifying the presence of infection, these methods have their 
significant limitations. For example, errors may be encountered at all stages of 
histopathological diagnosis, like processing of tissues, difficulty in histological 
detection and misinterpretation, etc. Thus, genetic information from molecular 
diagnostics provide critical additional information that help in correct diagnosis of 
a disease (Fig. 1). 
 
Figure 1: Traditional and improved method of molecular 
diagnostics 
Source:http://www.cancer.gov/cancertopics/understandingcancer/moleculardiagn
ostics/AllPages 
 
Technology of Molecular Diagnostics 
A number of techniques have been employed for molecular diagnostics of a 
particular disease. Some of the powerful tools are described briefly below: 
 
? Immobilization of DNA by Southern or dot blotting or in situ 
hybridization (isH) or Fluorescent in situ hybridization (FisH): —a 
labeled DNA or RNA strand that hybridizes with the target, complementary 
sequence and hence identifies and quantifies the target sequence in the 
sample (e.g., blood, tissue, saliva, etc.); 
Page 5


Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
 
 
Lesson Prepared Under MHRD project “National 
Mission on Education Through ICT” 
 
Zoology 
Biotechnology 
Lesson: Molecular diagnosis of genetic diseases 
(Cystic fibrosis, Huntington’s disease, Sickle cell 
anemia) 
Lesson Developer: Dr. Jaspreet Kaur 
College/Dept: S.G.T.B. Khalsa College 
University of Delhi 
Lesson Reviewer: Dr. Ravi Toteja 
College/Dept: Zoology, Acharya Narendra Dev 
College 
University of Delhi 
 
 
 
 
 
 
 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
 
 
 
Table of Contents 
 
? Introduction  
? Molecular diagnosis of Genetic diseases 
? Technology of Molecular Diagnostics 
? Cystic Fibrosis or mucoviscidosis 
? Symptoms and Causes 
? Pathogenesis 
? Structure of CFTR 
? Function of CFTR 
? Common Mutations leading to Cystic 
Fibrosis 
? Methods used in CFTR testing 
? Huntington’s Disease 
? Symptoms and Causes 
? Structure of Huntingtin 
? Mutation that causes Huntingtin’s 
disease 
? Methods used in testing Huntington’s 
Disease 
? Sickle Cell Anaemia 
? Symptoms and Causes 
? Structure of Hemoglobin 
? Mutation which causes Sickle cell anemia 
? Methods used in testing Sickle Cell 
Anaemia 
 
? Summary 
? Exercise/ Practice 
? Glossary 
? References/ Bibliography/ Further Reading 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
  
 
 
Introduction 
A genetic disorder can be defined as any mutation in gene which changes its 
instructions to make protein or improper functioning of the protein or its complete 
absence, is termed as Moreover, these disorders are categorized as: 
? Single-gene disorders: where a mutation affects only one gene. 
Example, Sickle cell anemia. 
? Chromosomal disorders: where chromosomes (or parts of 
chromosomes) are missing or changed. Example, Down syndrome.  
? Complex disorders: where mutations affect two or more genes. 
Example, Colon cancer. 
? Mitochondrial disorders: where mutations affect the non-chromosomal 
DNA of mitochondria. Example, an eye disease called Leber's hereditary 
optic atrophy; a type of epilepsy called MERRF which stands for Myoclonus 
Epilepsy with Ragged Red Fibers. 
 
In comparison to the infectious diseases, genetic disorders represent a major 
health problem since they may develop in later life and are usually incurable. 
Thus, molecular diagnosis of these disorders, especially at the prenatal stage 
becomes extremely important to prevent their transmission to next generation. 
Molecular Diagnosis of Genetic Diseases 
In order to efficiently deliver effective care to a patient, the first and foremost 
thing for a health care practitioner is the ability to accurately identify the cause of 
patient‘s problem, i.e., make the diagnosis. 
Now, as the name suggests, ?Molecular diagnostics? is a comprehensive term 
defining different diagnostic tests that assess a person‘s health literally at a 
molecular level, i.e., detecting and measuring specific genetic sequences (also 
called biological markers or biomarkers) in deoxyribonucleic acid (DNA) or 
ribonucleic acid (RNA) or the proteins they express. Molecular diagnostics help to 
determine whether a specific person is predisposed to have a disease, or actually 
have a disease, or whether a certain treatment option is likely to be effective for 
a specific disease. 
In the nineteenth century, before the advent of molecular diagnostics, the only 
available tools were microscopy and histopathology or cellular pathology. For 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
 
example, clinicians categorized cancer cells according to their pathology, that is, 
according to their appearance under a microscope. Despite being the first line 
tools for quickly identifying the presence of infection, these methods have their 
significant limitations. For example, errors may be encountered at all stages of 
histopathological diagnosis, like processing of tissues, difficulty in histological 
detection and misinterpretation, etc. Thus, genetic information from molecular 
diagnostics provide critical additional information that help in correct diagnosis of 
a disease (Fig. 1). 
 
Figure 1: Traditional and improved method of molecular 
diagnostics 
Source:http://www.cancer.gov/cancertopics/understandingcancer/moleculardiagn
ostics/AllPages 
 
Technology of Molecular Diagnostics 
A number of techniques have been employed for molecular diagnostics of a 
particular disease. Some of the powerful tools are described briefly below: 
 
? Immobilization of DNA by Southern or dot blotting or in situ 
hybridization (isH) or Fluorescent in situ hybridization (FisH): —a 
labeled DNA or RNA strand that hybridizes with the target, complementary 
sequence and hence identifies and quantifies the target sequence in the 
sample (e.g., blood, tissue, saliva, etc.); 
Molecular diagnosis of genetic diseases (Cystic fibrosis, Huntington’s 
disease, Sickle cell anemia) 
 
Institute of Lifelong Learning, University of Delhi 
? Amplification of DNA using the polymerase chain reaction (PCR): 
the process of exponentially increasing the amount of a specific DNA or 
RNA sequence found in a sample until there are so many copies that they 
can be detected and measured; 
? Restriction endonuclease digestions: identifying RFLPs (Restriction 
Fragment Length Polymorphisms) that are linked to the disease gene. 
? Microarrays: which measure the expression of a large number of genes, 
or detect single nucleotide polymorphisms (SNPs), or genome regions;  
? Sequencing: a technique used to map out the sequence of nucleotides 
that comprise a strand of DNA. 
The next section describes the use of the above listed techniques in some 
genetic diseases as test examples in order to appreciate the impact of 
molecular diagnostics. 
 
Cystic Fibrosis or mucoviscidosis 
It is the most common and life threatening autosomal recessive disorder, 
meaning that the abnormal or mutated gene is located on one of the autosomes 
and two copies of the gene are necessary to have the trait or disease, one 
inherited from the mother, and one from the father (Fig. 2).