Medical Genetics

Genetics plays an important role in the field of medical sciences. There arises a genetic base to a disease, the development, reproduction, and other processes in the body. The gene or a blueprint of life occurs in every cell and is an important inheriting factor. Hence, it is important to understand genes and their role in various diseases, cancers, developmental stages, and interactions with various components. Some of the genes encode for proteins. The genes get passed on from one generation to the next generation. They also carry any minute changes or mutations. The diverse population is a sign of variation. Due to recent advances in health research, more and more diseases showed a genetic base. The science of genetics helps in detecting many diseases. It helps in their treatment or prevention. When you try to understand the molecular intricacies of the genetic material, you will be amazed to see an entirely different world. Thousands of reactions keep occurring at a molecular level.

DNA, the blueprint of life consists of nucleotide base pairs attached to the phosphodiester bonds. Small units of DNA form genes and exhibit specific functions. Various disorders arise due to gene mutations. Hence, they are known as genetic disorders. Detection of genetic disorders involves modern genetic diagnostic techniques. There involves the tremendous importance of genetics in medical sciences. To understand its role in health and disease, we must know the basic concept of genetics. 

The review covers the following topics in detail:

1.     What are genes and various branches of genetics?

2.     Chromosomes and disorders associated with the genes

3.     Disorders inherited due to faulty genes

4.     The role of gene mutations in biochemical disorders

5.     Detection of genetic disorders

6.     Genetic counseling

7.     Gene Cloning and DNA analysis

8.     Gene therapy

Image 1: Medical genetics

Various branches of genetics:

Human genetics involves several branches such as cytogenetics, molecular genetics, biochemical genetics, cancer genetics, Immunogenetics, and developmental genetics. Cytogenetics is the study of chromosomes and various disorders associated with the same. A chromosome is a specialized structure consisting of DNA-protein complex packed in a condensed manner. Cytogenetic techniques help in studying these structures. Molecular genetics involves the study of genes at the molecular levels such as single base pair changes, mutations, and other studies. Biochemical genetics involves genes controlling the enzyme production. Cancer genetics involves studying cancer genes and mutations associated with cell cycle control. Different antigen-antibody interactions studies involve immunogenetics. Developmental genetics helps to study the genetic control of the development. Hence there are many such branches of genetics. The field of genetic science is very vast. Half the rate of first trimester abortions involves chromosomal abnormalities. Congenital malformations, childhood blindness, deafness, and mental retardation mostly occur due to gene mutations.

Chromosomes and disorders associated with the same:

Every cell in the human body constitutes chromosomes. The chromosome complement in humans consists of autosomes and sex chromosomes. There are 22 pairs of autosomes and a pair of sex chromosomes. They are known as X and Y chromosomes respectively. A normal male has 22 pairs of autosomes, an X, and a Y chromosome. A normal female has 22 pairs of autosomes and two X chromosomes. Karyotyping involves arranging the chromosomes as per the groups resulting in a photomicrograph. This photomicrograph or a karyotype consists of autosomes and sex chromosomes arranged in groups. Structural and numerical abnormalities in the chromosomes result in genetic disorders. They are known as chromosomal anomalies or chromosomal abnormalities.

Disorders due to an abnormal number of chromosomes are known as numerical anomalies. They arise due to non-disjunction of chromosomes. Monosomies (one chromosome less), trisomies (one chromosome extra), and many other conditions consist of chromosomal anomalies. Down’s syndrome is an example of trisomy. It occurs due to trisomy of the 21^st chromosome. Disorders associated with the structural abnormalities of chromosomes are known as structural anomalies. Deletions, duplications, inversions, and translocations arise due to the structural anomalies of the chromosomes. They also result in conditions such as mosaicism.

Disorders inherited due to faulty genes:

Monogenic or single gene disorders are the genetic disorders arising due to the inheritance of a mutated gene. Two main types of inheritance include autosomal inheritance and sex-linked inheritance. Autosomal inheritance involves autosome related disorders or traits. Inheritance of the traits due to the expression of genes present on the sex chromosomes is known as a sex-linked inheritance. Autosomal dominant inheritance manifests the trait even if the mutant gene occurs in a single dose. For example, Huntington’s disease arises due to abnormal CAG nucleotide repeat. Autosomal recessive inheritance manifests the trait even if the gene is present in the double dose (homozygous). An example includes sickle-cell anemia. Sex-linked inheritance is either X-linked or Y-linked. The X-linked inheritance occurs in a dominant or a recessive form. Very few cases report Y-linked inheritance.

Polygenic or multifactorial inheritance depends on many genes. The traits are known as quantitative traits and depend on many factors. Abnormalities in mitochondrial DNA (mtDNA) occur due to mitochondrial inheritance. The genes do not behave as dominant or recessive in polygenic inheritance. They exhibit an additive effect on the trait.

Biochemical genetics:

This area of genetics deals with the genetic control of the metabolic pathways. According to the one gene-one enzyme hypothesis proposed by Beadle and Tatum, metabolic processes occur in various steps controlled by enzymes. Each enzyme gets coded by one gene. Inborn error of metabolism arises due to enzyme unavailability or insufficiency occurring as a result of related gene mutations. The inborn error of metabolism follows Mendelian inheritance pattern. PKU, a classic example of the inborn error of metabolism, arises due to the deficiency of phenylalanine hydroxylase.

Image 2: The role of genetics in medicine

Detection of genetic disorders:

Cytogenetic and molecular genetic studies or tests detect various anomalies. The prenatal diagnosis helps to detect abnormalities in the fetus before birth. Examples of prenatal tests include Amniocentesis, chorionic villus sampling, fetoscopy, ultrasonography, maternal serum screening, and fetal blood sampling. Non-invasive prenatal tests also help in detecting the fetal DNA through maternal serum testing. Karyotyping helps to detect chromosomal abnormalities. FISH and other hybridization techniques detect minute changes in the DNA. SNP genotyping involves PCR, agarose gel electrophoresis, and gel documentation systems.

Genetic counseling:

It is a specialized session with a genetic counselor. The genetic counselor is an expert in genetics and a trained professional who guides the couples and patients suspected with the genetic disorders. Genetic counseling helps in the risk assessment of hereditary diseases, repeated abortions, and stillbirth cases. It also helps in reducing the risk of having a baby with the genetic and other ailments including the inborn errors of metabolism.

Gene cloning and DNA analysis:

Gene cloning involves cloning the gene of interest for incorporation into the desired vector. It has a wide range of applications in recombinant DNA technology. It helps in expressing the desired products such as proteins, vitamins, and other molecules. Peptide vaccines and hormones get developed using gene cloning. DNA analysis using techniques such as DNA fingerprinting help to solve parental issues. Analyzing DNA and genotyping techniques help in detecting gene mutations.

Gene therapy:

It helps in replacing the gene that has lost its function. The first step involves defective gene identification and cloning the normal gene in place of the defective gene. The normal gene insertion includes a vector. The first gene therapy gained success in case of a child having ADA deficiency associated with severe combined immunodeficiency syndrome (SCID). 

References:

[1] Emery’s elements of Medical Genetics, Peter D. Turnpenny

[2] Chromosome Abnormalities and Genetic Counseling,  R.J. MKinlay Gardner, Grant R Sutherland, Lisa G. Shaffer
[3] Medical Genetics, G. Bradley Schaefer, James N. Thompson

Copyright, 2018 All Rights Reserved

Prenatal diagnosis

Genetic disorders are difficult to treat.  It is very important to diagnose a genetic disorder. The prenatal diagnosis helps to detect genetic abnormalities before birth. Prenatal diagnosis involves various techniques such as ultrasonography, fetal blood sampling, fetoscopy, maternal serum screening, testing cell-free DNA, amniocentesis, and chorionic villus sampling.

What is the prenatal diagnosis?

The prenatal diagnosis serves in detecting the genetic abnormalities in a child before birth. Thus, it helps the parents in decision making.

The high-risk couples get a chance to decide.  Following couples get the benefit of prenatal diagnosis:

1.     The family history of genetic disorders

2.     Women above 35 years expecting a baby

3.     The family history of neural tube defects

4.     The first child has chromosomal abnormalities or any genetic condition

5.     Couples having a first child with Down's syndrome

6.     Diabetics

7.     Carrier mothers

Period of screening	Prenatal screening tests	Estimation of risk
First trimester	Blood tests	·        Estimate the level of pregnancy-associated placental protein A ·        Beta-human chorionic gonadotropin ·        Help in estimating the risk of Down’s syndrome
	Ultrasonography	·        Detection of the abnormalities associated with the neck and back of the fetus.
	Cell-free fetal DNA (cfDNA testing)	·        Determine chromosomal abnormalities.
	Chorionic villus sampling, amniocentesis	·        Amniocentesis helps in estimating the alpha-fetoprotein. ·        Help in the detection of chromosomal abnormalities.
Second trimester	Blood tests	·        Alpha-fetoprotein ·        Estriol ·        HCG ·        Inhibin A ·        Detection of neural tube defects ·     Complications associated with miscarriages
	Ultrasonography Amniocentesis Triple screening	·        Determine the presence of more than one fetus ·        Determine  defects

Table: Estimation of risk using prenatal diagnosis

Amniocentesis:

Amniocentesis involves chromosomal abnormalities to get detected by a collection of the amniotic fluid. It involves obtaining the cells of the developing fetus for analysis. The procedure gets performed between 14-16 weeks of pregnancy.

The amniotic fluid analysis helps in detecting neural tube defects by estimating alpha-fetoprotein with a raised level in the amniotic fluid. The amniotic fluid is a protective fluid that surrounds the developing fetus. It serves as a cushion against shock. The amniotic sac provides a favorable environment for fetal development. Amniocentesis is a very specialized procedure that requires an expert. The procedure is extremely risky and needs proper concentration and careful examination. The amniotic fluid is collected using a syringe needle injection through the uterine wall and amniotic sac. This fluid consists of fetal cells. These cells are analyzed further for the chromosome studies. The fetal cells are cultured in the laboratory. Various tests such as DNA tests and biochemical tests for enzyme deficiencies are carried out.

Image: Amniocentesis

Chorionic villus sampling:

Chorionic villus sampling is a form of biopsy. It involves sampling of the fetal tissue which contains a large number of fetal cells. It detects chromosomal disorders as well as single gene disorders. The procedure is carried out during 10 to 11 weeks of the gestation period. The risk of abortions reduces with this type of detection. This procedure is carried out at early stages of pregnancy. Chorionic villus sampling between the 8^th and 12^th weeks of pregnancy is ideal. The amniocentesis is ideal after the 12^th week of gestation. A membrane layer surrounding the fetus is known as a chorion.

Non-Invasive Prenatal Testing:

The reason for conducting a genetic test involves the detection of any mutations occurring in the fetus before birth. Prenatal diagnosis is an important application of genetic testing. Hence, we check whether the fetus is at risk. Two main procedures such as prenatal diagnosis and chorionic villus sampling involve a lot of risk for the mother and the baby. They are very painful too. Hence, it led to the development of non-invasive prenatal testing (NIPT). The maternal plasma shows the presence of fetal DNA. It is a cell-free DNA floating in the plasma of the expected mother. The abbreviation of cell-free DNA is known as cfDNA. Hence, it is possible to collect plasma samples of the expected mothers for analyzing cfDNA. NIPT is a simple, harmless, and painless procedure. It detects gene mutations and chromosomal abnormalities. The embryos with chances of serious genetic disorders get removed before the implantation stage.

Fetoscopy:

It is a kind of endoscopy that enables to visualize the fetus.  A fetoscope is a fibro-optic self-illuminated instrument that gets inserted in the amniotic cavity under local anesthesia. Cleft lip, cleft palate, facial malformations, limb defects, and skin disorders get detected with the help of fetoscopy.

Ultrasonography:

The 12^th week of pregnancy is an ideal period for performing ultrasonography test. However, the first and the second trimesters may involve the test. It is an ultrasound-based imaging technique involved in the purpose of diagnosis.  The high-frequency sound waves help in generating images. Ultrasounds help in confirming the pregnancy. It helps in checking the fetal heartbeat and help in detecting any abnormality in the fetus. It detects an ectopic pregnancy. It is a condition in which the fetus does not attach to the uterus. It is possible to determine the sex of the baby using ultrasonography. During the second trimester, it helps to determine the possible characteristics of the Down’s syndrome or any other physical abnormalities in the baby.

Maternal serum screening:

The indication of the level of alpha-fetoprotein uses maternal serum screening. The raised level of alpha-fetoprotein indicates neural tube defects, including spina bifida and anencephaly.

Fetal blood sampling:

It involves a prenatal diagnosis of hemophilia, thalassemia, sickle cell disease, immune deficiency disorders, and chromosomal analysis. It requires a small amount of blood from the fetus or the unborn baby.

Conclusion:

Although prenatal testing serves for the detection of many genetic conditions, still many babies are born with the genetic conditions every year. The reason involves the lack of proper knowledge regarding familial disorders, pregnancy risks, and many other environmental conditions. It is important for the physicians to be familiar with the risks associated with the genetic disorders and the genetic tests available for the same. The knowledge of prenatal testing and genetic counseling helps to prevent the birth of the babies with the defects and help in decision making.

References:
[1] Human Genetics, 3/e, Gangane
[2] Chromosome Abnormalities and Genetic Counseling, R.J. McKinlay Gardner, Grant R Sutherland, Lisa G. Shaffer
[3] Essential Medical Genetics, Edward S. Tobias, Michael Connor, Malcolm Ferguson-Smith
[4] Thompson & Thompson Genetics in Medicine, Robert L. Nussbaum, Roderick R. McInnes, Huntington F. Willard

                                     © Copyright, 2018 All Rights Reserved.

Chromosome analysis

The chromosome analysis involves many conditions such as congenital malformations, mental retardation, repeated abortions, sex determination, and prenatal diagnosis. The procedure of chromosome analysis involves techniques such as karyotyping and FISH. A microscopic examination of chromosomes for detection of abnormalities or novel mutations involves chromosome analysis. It helps to detect monosomy, trisomy, infertility issues, repeated abortions, hereditary syndromes, and chromosomal aberrations. It helps in detecting the disorder or a disease. It helps in the genetic counseling and consultation. It is possible to diagnose hematological disorders as well. Modern FISH techniques detect cytogenetic abnormalities in haemato-lymphoid malignancies and hereditary cancers. Let us know the technique in detail.

Karyotyping:

It is a procedure to obtain karyotype of an individual. A karyotype is nothing but a chromosome complement of an individual. It depicts a photomicrograph of the metaphase chromosome arranged in a standard sequence. The review article describes the procedure in detail.

The human peripheral blood lymphocytes culturing and processing helps to obtain a karyotype. A sterile syringe is used to collect the sample. It involves the collection of 5ml of the nervous blood in a heparinized tube. Addition of heparin prevents blood clotting. The procedure primarily involves the addition of the heparinized whole blood sample into the culture medium consisting of RPMI-1640, fetal calf serum, phytohaemagglutinin, and antibiotics. The culture media and the fetal calf serum nourish the lymphocytes. Phytohaemagglutinin stimulates cell division in lymphocytes. Addition of the antibiotics prevents the infection. The culture vial involves incubation of 3 days at 37⁰C temperature.

The incubation of the culture vials leads to the division of the lymphocytic cells. The addition of a mitotic inhibitor (colchicine) at the end of the third day, helps a lot by preventing the spindles. It arrests the cells in metaphase. Visibility of the chromosomes is best during the metaphase. Then the culture vials are kept undisturbed for two hours. After 2 hours the lymphocytes are centrifuged. Then the cells undergo saline treatment. The saline creates a hypotonic treatment. The cells swell under the hypotonic environment. Followed by re-centrifugation, the procedure requires removal of the supernatant. After discarding the supernatant, the cells get fixed in a fixative such as Carnoy’s fixative. A careful dropping of few drops of the cell suspension on a clean, pre-chilled, grease free slide enables the chromosomes to get dispersed. The processing of the slides and staining them helps to visualize the chromosomes under a microscope. Karyotyping software helps to arrange the chromosomes in groups thereby generating a photomicrograph of chromosomes known as a karyotype.

Observation of the chromosomes:

A karyotype reflects the differences in the absolute sizes of the chromosomes. It helps to study the DNA duplication. The differences in the relative sizes of the chromosomes arise due to the interchange of the chromosomal segments. The chromosomal segments may interchange in unequal lengths. A karyotype helps to observe various types of chromosomal abnormalities known as deletions, duplications, translocations, and inversions. For example, a translocation arises due to differences in the Centromeric position. Grouping of the chromosomes helps in identifying the correct number of the chromosomes. Also, the karyotyping enables to study the number and the position of the satellites.

Giemsa banding:

It commonly helps in analyzing the chromosomes. Giemsa banding technique first involves treatment of the chromosomes with a denaturing agent known as trypsin. It helps in denaturing the proteins. A stain known as Giemsa helps to stain the slides. Staining with the Giemsa stain creates a unique banding pattern to the chromosomes. The pattern shows light and dark bands. A specific banding pattern helps to visualize the long and short arms of the chromosome. Giemsa stain is a complex stain specific to the phosphate groups of the DNA. The light and dark stripes appearing on the chromosomal arms depict the bands. They appear after staining the cell preparation.

The heterochromatin depicts a dark band. The euchromatin depicts a light band. Heterochromatin is rich in the repetitive DNA sequences. Other types of banding techniques include R-banding, C-banding, Q-banding, T-banding and silver staining.

Three main types of karyotyping include classical karyotyping, spectral karyotyping, and virtual karyotyping. The above method of karyotyping involves Giemsa banding belonging to the classical karyotyping. The spectral karyotyping helps to visualize all the chromosomes in different colors. This type of karyotyping is known as SKY technique. A digital karyotype helps to quantify the DNA copy number.

Image : Giemsa banding of chromosomes

Fluorescence in situ hybridization (FISH):

A metaphase chromosome consists of a DNA packed into it in a highly condensed form. The DNA sequence helps to detect the defects. The FISH technique uses a single-stranded DNA probe. It has a unique ability to anneal with the complementary DNA sequence on the chromosome. A DNA probe is a single-stranded sequence. It gets labeled radioactively. The DNA probe detects the DNA fragments with similar sequences. The hybridization of the DNA sequence with the probe sequence enables it to be visualized using autoradiography.

The FISH technique involves different types of probes. The Centromeric probes are nothing but the DNA sequences found in and around the centromere. They are specific to a particular chromosome and have a repetitive sequence. A chromosome-specific unique sequence probe identifies sub-microscopic deletions and duplications. A whole chromosome paint helps to visualize an entire chromosome. Examination of slides under a microscope detects the presence of a hybridized fluorescent signal. It also detects the absence of the chromosomal material if there is no signal. Not only metaphase chromosomes but also non-metaphase chromosomes such as interphase chromosomes hybridize with the fluorescent labeled probes.

Chromosome painting is the next version of FISH. In this technique, the hybridization probe involves a mixture of DNA molecules specific for different regions of a single chromosome.    

References:
[1] Medical genetics, G.P. Pal
[2] Molecular Cytogenetics: Protocols and Applications, Yao-Shan Fan  
[3] Human Chromosomes: Structure, Behavior, and Effects, Eeva Therman, Millard Susman
[4] The AGT Cytogenetics Laboratory Manual, Marilyn S. Arsham, Margaret J. Barch, Helen J. Lawce  

                                      © Copyright, 2018 All Rights Reserved.

The First Gene Therapy

Gene therapy is a new type of treatment strategy. It involves the replacement of a defective gene with a normal gene. It helps to restore the lost gene function in the body. The first gene therapy in humans was carried out in the year 1990 by French Anderson. The therapy helped in treating a child suffering from ADA deficiency. The deficiency of ADA involves an enzyme deficiency known as adenosine deaminase. The WBCs become functionally inactive in ADA deficiency. The disease leading to the ADA deficiency is known as SCID or severe combined immunodeficiency. The gene therapy proved to be successful in the child suffering from SCID. However, only the somatic gene therapy was successful. It was because the somatic gene therapy helped in treating the disease. With time, the advancements in the gene therapy kept on progressing. However, the first gene therapy set an example of new treatment mode. The idea of gene therapy helped the healthcare sector in advancing and treating rare diseases and disorders with no drug treatment.

The gene therapy encompasses many types of genetic engineering applications for inserting the relevant genes in humans. Among all, the somatic cell gene therapy technically sounds simple and convenient. The children with the ADA deficiency die early due to the lack of an essentially functional immune system. They fail to fight the recurrent microbial infections. Most of the patients suffering from ADA deficiency die due to imbalanced platelets or WBCs due to viral or bacterial infections.

Image 1: Gene therapy

Severe Combined Immunodeficiency (SCID):

Before understanding the gene therapy in SCID patients, we must know more about this immunodeficiency syndrome. SCID stems from a defective lymphoid development. It mainly affects the T-cells, B cells and the natural killer cells (NK cells). Although SCID is an immunodeficiency disorder, it mainly arises due to the gene defects. The circulating lymphocytes in SCID individuals are very less in number. Thus, the entire immune system gets hampered. The failure of the T-cell response and development directly affects the thymus. The thymus is a specialized primary lymphoid organ, important for the immune system. The T-cell maturation mainly occurs in the thymus. They mainly help in building the adaptive immunity and fight with the foreign invaders. The thymus poorly develops in the individuals with the condition such as SCID. The development of the thymus gets affected in this condition. The individuals with SCID experience a low adaptive immunity and hence face the difficulty in fighting the invading microbes. The T-cell count is lower than the expected range. Although the individuals with SCID exhibit normal levels of the myeloid and erythroid cells, the impact of the depleted lymphoid cells is extremely high. Hence, it leads to severe complications.

While designing the gene therapy for the individuals with SCID, the WBCs were thoroughly studied. Since the root of the disease lies in the nucleus of the WBC, it was thus important to study the genes and deduce a therapy. The platelets and erythrocytes lack a nucleus. The white blood cells show a well-defined nucleus. The SCID infants and children always suffered due to recurrent infections, opportunistic infections, chronic diarrhea, pneumonia and skin infections, mouth and throat lesions. The patients with SCID show a highly compromised immune system. Even the vaccines revert and lead to severe infections. Another feature of SCID involves a defective antibody response arising due to affected B-lymphocytes. SCID affects one in a lakh newborn babies.

Although the somatic gene therapy in ADA deficiency was easy to define theoretically, it was an extremely difficult and careful activity conducted by a group of experts in medicine, immunology, and genetics. 

Following are the types of severe combined immunodeficiency:

Type of SCID	Genetic Condition
X-linked IL-2RγChain deficiency	Mutations arise in a gene encoding a common gamma chain for an interleukin receptor protein.
JAK-3 deficiency	JAK 3 gene mutation leads to JAK 3 enzyme deficiency.
CD45 deficiency	Mutant CD45 alleles, uniparental disomy
IL-7R αchain deficiency	IL-7R gene mutations encoding an IL-R alpha chain.
CD3 δchain deficiency	Mutations in gene encoding CD3 delta chain
Adenosine deaminase deficiency (ADA)	Defective adenosine deaminase enzyme production.
Artemis deficiency	Lack of Artemis gene.
RAG1 and RAG 2 deficiency	Mutations in RAG-1 and RAG-2 genes result in prevention of VDJ recombination.

Table: Types of SCID

The first gene therapy was carried out in a girl suffering from SCID due to ADA deficiency. The enzyme known as adenosine deaminase catalyzes the conversion of adenosine to inosine. Hence, the deficiency of adenosine deaminase results in accumulation of adenosine. It interferes with the purine metabolism and DNA synthesis. It also results in the accumulation of toxic metabolites in the T cells and B cells. It is an autosomal recessive disease since it follows a pattern of autosomal recessive inheritance. The chromosome 20q13 possesses the defect. It means that the mutant allele is present in a double dose.

The gene therapy in ADA deficiency consisted of following steps:

Step 1: Collection of the patient's blood sample

Dr. French Anderson and his colleagues decided to conduct a clinical trial in ADA deficient girls. The blood sample collected from the affected girl involved careful analysis. The patient and her parents signed the consent form for the therapy. Analysis followed the culturing of the cells.

Step 2: Filtration of WBCs

The process of filtration helped to retain the white blood cells. Hence the white blood cells were filtered out of the red blood cells and platelets. The WBCs stored carefully helped in analyzing them.

Step 3: Alteration of the viral vector

The viral vector used in the therapy was known as an Adenovirus. It belongs to the class of retroviruses. The therapy involved a careful manipulation of viral genes for preventing its replication in the cells. It must only do the job of delivering the life-saving ADA gene.

Step 4: Culturing WBCs mixed with a viral vector having ADA gene

The cells grew in large numbers in tissue culture plates. The process involved mixing the cells with the genetically engineered virus. The virus inserted the ADA gene into the target cell’s DNA.

Step 5: Injection of WBCs containing the life-saving ADA gene

After screening the cells containing the therapeutic gene, the further procedure involved returning the cells with the ADA gene to the patient’s body. Hence, the correct gene gets inserted into the patient’s genome. 
References:
[1] Human Genetics, 3/e, Gangane
[2] Medical genetics, G.P. Pal

© Copyright, 2018 All Rights Reserved.