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DNA sequencing technology

DNA Sequencing
UE Gestion de l’innovation
Teacher: Christophe Lécuyer
Anya Rahmoune
L3 Sciences de la Vie-Gestion
II – History of DNA sequencing
III – Pharmacogenomics
IV – Direct-to-consumer genetic testing: case study of 23andme
IV – Genetic sequencing, ethics and society
Since its discovery, DNA has fascinated scientist and general public alike. Indeed, a large part of a person’s
information can be found in its DNA: their biological sex, their ancestry, their physical appearance, sometimes
even their medical condition. Genetic makeup is part and parcel of who we are. It’s why DNA sequencing was
crucial to the pursuit of research in Life Sciences. Some scientists go as far as to say DNA sequencing
technology is such an important innovation, it is akin to that of the microscope. Subsequently, this technology
was embraced by many industries in the Life Sciences field: medicine, virology, evolutionary biology, forensics,
etc. and even spawned new ones, two of which we will study in this essay: pharmacogenomics and direct-toconsumer genetic testing.
The question is, how did DNA sequencing technology came to be and how did it become essential and
ubiquitous in the Life Sciences industry, adopted as it is by every laboratory and company in this field? To
understand how this came to be, we will start with a retelling of the discovery of genetic testing technology.
I – History of DNA sequencing
DNA sequencing as an innovation was very much incremental. Several milestones had to be reached to get
to the technology we have today. Listing each one would be too long, and it is not the point of this essay, so
we’ll only name the most important ones.
An important figure in DNA sequencing is Frederick Sanger, a British biochemist who received two Nobel
Prizes: the first for “his work on the structure of proteins, especially that of insulin”, the second, he shared
with Walter Gilbert for “their contributions concerning the determination of base sequences in nucleic acids”
– that is to say, DNA sequencing. Sanger did not discover DNA sequencing methods immediately, he first had
to sequence a protein, insulin, then RNA, and finally DNA.
Sequencing of Insulin
Frederick Sanger obtained his Ph.D in 1943 in Cambridge, working on the metabolism of the amino acid
lysine in the animal body. The same year, he got his first job, a position in the Biochemistry department of
Cambridge offered by A.C. Chibnall. Chibnall was interested in insulin, of which he determined, with
colleagues, the amino acid composition of its bovine form. The structure of insulin, that is to say, the exact
order of these amino acids, was however still a mystery to biologists.
Chibnall put Sanger up to the task of investigating the end groups of the
protein, which would later lead to research on its structure. Sanger tried several
techniques and molecules before he managed, thanks to his connections, to get
his hands on dinitrofluorobenzene (DNFB), synthesised at the time for the war
effort. It worked, and he published his first paper. It was not the discovery of the
entire sequence yet, but it was a start.
Sanger later found insulin was made of two chains, that he called chain A et
chain B. With Hans Tuppy and later Ted Thompson, Austrian and Australian
biochemists, he gradually pieced together the complete structure of insulin from
overlapping sequences.
The only thing missing was the emplacement of the disulfide bonds1 that
joined the two chains. Using the new technique of paper electrophoresis2, Sanger located the disulfide
bridges, thus finally completing the structure of insulin.
Sanger therefore proved proteins are well-ordered molecules and that, by analogy, genes must be, too. For
his work, Sanger received his first Nobel Prize in Chemistry in 1958.
Sequencing of RNA
At the time, nucleic acids, that is to say, DNA and RNA, were still a mystery to scientists. Sanger only got
interested in RNA because DNA remained too difficult to sequence yet. We know now DNA and RNA are made
of pieces called nucleotides, of which there are five types: A (adenine), C (cytosine), G (guanine), T (thymine)
and U (Uracil). We know DNA is exclusively composed of A, C, G, and T, and mRNA3 can be a perfect mirror to
DNA, with the only difference being every T is replaced by a U in the sequence. Today, we also know, for every
gene of the human genome, the exact order of these nucleotides.
Disulfide bonds : a disulfide refers to a functional group with the structure R−S−S−R′. The linkage is also called an SS-bond or
sometimes a disulfide bridge. Source : Wikipedia.
Paper electrophoresis : technique used to separate small charged molecules such as amino acids and small proteins.
mRNA : single-stranded RNA molecule that corresponds to the genetic sequence of a gene and is read by the ribosome in the
process of producing a protein. Source : Wikipedia.
However, at the time, DNA molecules were too big and complicated for scientists to sequence, with no
known technique to identify the structure. With RNA, on the other hand, it was more manageable. Even if the
simplest RNA viruses contained a few thousand nucleotides, still too big to sequence, tRNAs4, a different and
newly-discovered type of RNAs, contained fewer than a hundred.
The appeal was obvious. Sanger’s goal was to not only work out sequence methods for RNA and be the first
to sequence one, but also to reveal the genetic code. Indeed, since mRNA reflects the DNA structure,
sequencing a mRNA corresponding to a protein whose sequence is known would reveal the genetic code.
However, Robert Holley from Cornell University published first his sequence analysis of the 77
ribonucleotides of alanine tRNA from a species of yeast in 1965 and so beat him to it. By 1967, Sanger's group
had determined the nucleotide sequence of a small RNA of 120 nucleotides from bacteria E. coli.
Sequencing of DNA
DNA sequencing was Sanger’s ultimate goal, the consecration of his career. The problem, at the time, was
the unavailability of small-sized DNA or ways to break long DNA into smaller, more manageable pieces.
Sanger chose to work on bacteriophage ØX174 DNA because of its convenience: the phage was singlestranded and his colleague John Sedat knew how to grow it. He wanted to use DNA polymerase5 to develop
his DNA sequencing method, but DNA polymerase requires a primer6, and such a primer would have to be
synthesized. Thanks to Hans Kössel, a German molecular biologist who worked alongside Khorana, an IndianAmerican biochemist who pioneered DNA synthesis methods at MIT, Sanger had access to octanucleotide
primers. So he used an octanucleotide primer and E. coli DNA polymerase.
In the early 1970s, Sanger hired a new technician, Alan Coulson, 20 year old. Because of their shared
reserved nature, a sharp contrast to the outgoing personality of other members of Sanger’s team, Coulson
and Sanger get on marvellously and worked well together.
Sanger and Coulson went on to invent what they called “the plus and minus method” of DNA sequencing.
By 1977, they had published the whole 5400 base pair sequence of bacteriophage ØX174. They also
discovered the number of proteins produced by ØX174 to be much higher than the number of its genes –
which meant that the one gene-one enzyme hypothesis of Beadle and Tatum was false. Gene sequences
overlapped, they just happened to start at different points in the DNA sequence.
This method outperformed other techniques, but it was still too laborious. Inspired by Fred Arthur
Kornberg’s work posing dideoxythymidine triphosphate (ddTTP) as a substrate for DNA polymerase, Sanger
developed another method, the dideoxy chain-termination method, or what is known today as the Sanger
method. In parallel, Maxam and Gilbert were working on another method, perfected around the same time
Sanger invented his. However, the Sanger method used fewer toxic chemicals, lower amounts of radioactivity
and was easier and reliable, so it became the favoured method from the 1980s to the 2000s7.
In 1980, Sanger received his second Nobel Prize in Chemistry, which he shared with Gilbert, for “their
contributions concerning the determination of base sequences in nucleic acids”.
Sanger’s method is only the first generation of DNA sequencing method. After that come the second and
third generations, quicker, more precise and more reliable, each time needing less genetic material to work
tRNA : necessary component of translation from mRNA to proteins.
DNA polymerase : enzymatic complex intervening in DNA replication during cell multiplication.
Primer : short single-stranded nucleic acid utilized by all living organisms in the initiation of DNA synthesis. Source : Wikipedia
Source : DNA sequencing, Wikipedia.
Shendure, Jay & Balasubramanian, Shankar & Church, George & Gilbert, Walter & Rogers, Jane & Schloss, Jeffery & Waterston,
Robert. (2017). DNA sequencing at 40: Past, present and future. Nature. 550. 10.1038/nature24286.
II – Pharmacogenomics
We can raise two relevant definitions of pharmacogenomics:
The scientific paper authored by Theodora Katsila and George P. Patrinos, defines it as follows:
“Pharmacogenomics aims to develop strategies for individualizing therapy to optimize drug efficacy and
minimize toxicity on the basis of our improved understanding of how genomic variants influence drug
According to the European Ubiquitous Pharmacogenomics program, “Pharmacogenomics is the study of
genetic variability affecting an individual’s response to a drug. Clinical application of pharmacogenomics
knowledge will result in less ‘trial and error’ prescribing and more efficacious, safer and cost-effective drug
Those are both good definitions. Nonetheless, to be more accurate, we need to explain yet another word:
pharmacogenetics. These two words, pharmacogenomics and pharmacogenetics, co-exist and are, while
slightly different, more or less interchangeable when it comes to designate what is essentially the same field.
Pharmacogenetics is the “study of variability in drug responses due to heredity”11, that is to say, how one’s
genes affect one’s response to a drug. Pharmacogenomics, on the other hand, is a broader term: the whole
genome is studied in relation to drug response – indeed, genes compose barely 2% of the genome, the rest is
noncoding DNA, mitonchondrial or chloroplast DNA.
An estimated 70% of response variability to drugs has a genetic origin, related to absorption, distribution,
metabolism, action mechanism, biological effects, underlying pathological state, etc. Such a high number can
only encourage scientists to look into it and the pharmacogenomics field.
To understand precisely what DNA sequencing technology’s role is in individualized therapy, we can use
the examples of the FSH receptor, asthma and cancer therapy.
The FSH, Follicle Stimulating Hormone, is a multifunction hormone targeting the gonads, reproductive
glands (ovaries and testicles). In the female body, the FSH receptor is situated at the surface of the ovaries.
Several variants of the FSH receptor have been identified. The genotype of the receptor determines the
ovarian response to FSH. The allelic combination serin/serin is associated with higher levels of FSH, which can
be explained by lower sensitivity to FSH: the body compensates a low sensitivity to the hormone with higher
levels of it. On the opposite, the mutated asparagin/asparagin receptor is related to higher sensitivity and is
then associated with ovarian hyperstimulation. Thus, depending on her genome, a patient can either be
exposed to a wrong response (abnormally high levels of FSH) or to hyperstimulation. In consequence, if the
patient ever needs medication for hormone-related issues, knowing her genome helps determining the most
suitable course of action to treat her, the best medication with the best dosage, increasing chances of success
and reducing drug-related risks in case of medication intake.
For asthma, we know there is a variability in the patient’s response to the 3 categories of treatments:
Beta2-adrenergic receptor agonists, antileukotrienes and inhaled corticoids. For every therapeutic class,
research shows the existence of polymorphisms in patients’ genes, such as β2-receptor, the 5-lipoxygenase
gene promoter or the corticotropin releasing factor receptor, are associated with different responses to the
different treatments.
In the case of cancer, researchers are looking into the genetic markers for the different types of cancer, but
also for a way to diminish the toxicity of chemotherapy who is, by nature, not an individualised treatment,
and thus has very harmful side effects.
Source : Whole genome sequencing in pharmacogenomics.
Source : The Ubiquitous Pharmacogenomics program website.
Source : Pharmacogenetics and pharmacogenomics.
III – Direct-to-consumer genetic testing: case study of 23andme
23andme is an American biotechnology company created in April 2006. It proposes direct-to-consumer
genetic testing with two offers: “Ancestry Personal Genetic Testing” and “Health + Ancestry Service”. The
former determines the client’s genetic ancestry and the latter uses known genetic markers to uncover which
diseases the client risk developing in future.
As of now, the company mainly operates in the United States of America, in part because the Health
Service, due to applicable regulations, is allowed only in the USA, Canada and the United Kingdom, and not
everywhere internationally.
The price of DTC genetic testing started at 999 $ in 2007, fell down to 399 $ in 2008 to finally attain the
more affordable price it is now, 99 $. To achieve such affordability, the product was sold at a loss to build
valuable consumer database for more accurate results in the future. Today, in the US, for 99 $ and a shipping
fee, anyone can obtain a genetic analyse of their ancestry. After ordering, a kit arrives home with the tools
and one only has to provide a sample of their saliva and mail it back to the lab. Results are accessible online
after 3 to 5 weeks.
Of course, the Ancestry test also allows clients to find their biological siblings or egg or sperm donor, if they
were conceived this way.
Title: 23andme’s genetic testing tool kit
In 2013, the Personal Genome Service Genetic Health Risk service had to be withdrawn from the market
due to lack of approval before it was finally authorized the Food and Drugs Administration in 2015 for
detecting the Bloom Syndrome, a “rare autosomal recessive disorder characterized by short stature,
predisposition to the development of cancer, and genomic instability.”12
Source : Wikipedia.
In 2017, 23andme was allowed to test for 10 diseases and conditions, described as follow by the FDA13:
- Parkinson’s disease, a nervous system disorder impacting movement;
- Late-onset Alzheimer’s disease, a progressive brain disorder that destroys memory and thinking
- Celiac disease, a disorder resulting in the inability to digest gluten;
- Alpha-1 antitrypsin deficiency, a disorder that raises the risk of lung and liver disease;
- Early-onset primary dystonia, a movement disorder involving involuntary muscle contractions
and other uncontrolled movements;
- Factor XI deficiency, a blood clotting disorder;
- Gaucher disease type 1, an organ and tissue disorder;
- Glucose-6-Phosphate Dehydrogenase deficiency, also known as G6PD, a red blood cell
- Hereditary hemochromatosis, an iron overload disorder; and
- Hereditary thrombophilia, a blood clot disorder.
The Health service is supposed to help clients make informed decisions about their lifestyle or start a
discussion with a health professional; however, the FDA warns: the presence or absence of genetic marker for
these diseases does not mean the client will or won’t ultimately develop a condition. Other factors,
environmental and lifestyle-related, are also determining and not to be neglected.
A major shift of the access point for genetic testing from physicians to the internet might occur in the next
decades, which would not only facilitate pharmacogenomics and thus, improve targeting of drugs, but also
have economic effects on public health care. Overall, DTC genetic testing might lower health care costs by
encouraging healthier behaviours and prevention strategies with an increase in early detection and
However, DTC genetic testing could also raise the cost if “it results in more tests, follow-on screenings and
interventions, or if negative test results give a false sense of security and either leads to under-use of regular
check-ups and other preventive measures, or is viewed as permission to resume, continue or commence
smoking, poor diet, or a sedentary lifestyle—all behaviours that contribute to a multitude of chronic, multibillion-dollar-a-year diseases.
In light of the potential for overuse of DTC genetic testing, it will be important to determine under what
circumstances genetic testing is appropriate and beneficial.”14
Another societal aspect to consider is that physicians are, for the most part, not prepared or equipped to
interpret whole genome analysis results. The lack of knowledge and expertise may result in mistrust in one’s
physician, seeing him or her as outdated and behind the times. This is due to medical schools not including
advanced genetics and genomics in their curriculum. In a survey15 done in Canadian and American schools, it
turned out less than half incorporated genetics classes in 3rd and 4th year, when clinical practice starts, and
most of the genetics classes focused on the theoretical aspect and not the practical applications. It will take
decades before producing enough physicians who are as comfortable with genetic results reading as they are
with other aspects of their work.
Source : FDA allows marketing of first direct-to-consumer tests that provide genetic risk information for certain conditions - Food
and Drug Administration, 6 avril 2017.
Source : Committee on Science, Technology, and Law, Forum on Drug Discovery, Roundtable on Translating Genomic-Based
Research for Health, National Research Council, Institute of Medicine - Direct-to-Consumer Genetic Testing: Summary of a
Thurston V, Wales P, Bell M, Torbeck L, Brokaw J., The current status of medical genetics
instruction in U.S. and Canadian medical schools, Academic Medicine, 2007; 82 (5):441-445
IV – Genetic sequencing, ethics and society
Genetic sequencing has been surrounded by controversy since the general public became aware of its
existence. Like every new technology, fear of its misuse arose. One does not have to look farther than media
in pop culture: the very well-known movie Gattaca, produced by Andrez Niccol in 1998, describes a world in
which genetically inferior individuals are discriminated against, while those of perfect genetics get to live a life
of privilege.
This fear, for the most part dissipated in the last decades, is born of the misconception that genetics
determine a person’s whole being, instead of being understood as the complex machinery it is, influenced by
exterior factors such as environmental or biological ones. We hear every day of the discovery of new genes
supposedly responsible for illness as varied as obesity, alcoholism or even serial-killer inclination.
Nonetheless, one cannot simply measure their intelligence, athletic ability, or creativity with DNA analysis.
For an employer, for example, it will never be as easy as undertaking a genetic test to determine a candidate’s
intelligence; a relevant aptitude test is better-suited for that purpose.
Another facet of the debate is prenatal genetic sequencing. Some worry parents will resort to abortion or
genetic manipulation if the baby does not match their ideals. This concern, which has been around for
decades now, predict this new future is incoming. As of right now, except in the case of medical reasons, this
has not happened yet, but it might be because we still do not possess affordable technology for such
advanced analysis on foetuses.
In the case of genetic testing done by private companies like 23andme, the suspicion of sale of personal
data to third-parties quickly arose.
This matter is furthermore complicated by the fact that not everyone is able to consent – for example,
minors, or unaware test subject’s relatives – but their lives could nonetheless be affected by the results. “In
recent years, in China, for approximately $900, parents in Chongqing can send their children—ages three to
12 years old—to a five-day camp for DNA testing to identify their gifts and talents, so they can focus on those
strengths from an early age. According to the director of the Chongqing Children’s Palace, “Nowadays,
competition in the world is about who has the most talent. We can give Chinese children an effective,
scientific plan at an early age.””16 On the opposite, in the United Kingdom, genetic testing on minors is only
recommended for medical reasons and if delaying the test until their coming-of-age might have negative
effects on their health.
Privacy concerns have yet to be addressed by companies offering DTC testing or competent, relevant
scientific and political entities.
Ultimately, the main concern remains the misreading of the results: without professional assistance, clients
can delude themselves into thinking they are perfectly healthy when they are not, or vice-versa.
Those are all questions of ethical, scientific and medical nature that our society will have to learn to answer
in the next decades, as DTC genetic testing goes on to be more affordable and accessible.
Source : Committee on Science, Technology, and Law, Forum on Drug Discovery, Roundtable on Translating Genomic-Based
Research for Health, National Research Council, Institute of Medicine - Direct-to-Consumer Genetic Testing: Summary of a
In conclusion, DNA sequencing is omnipresent: it is in every Life Sciences field, in every laboratory, every
company. It is the result of decades of research and the combined efforts of hundreds of scientists. Frederick
Sanger played an important role, as the biochemist responsible for the sequencing of inulin, then RNA and
finally DNA. His method, though improved upon, has been used continuously for decades and still is, for it is
reliable and cost-effective.
Amongst others, two new fields emerged out of the birth of this technology: pharmacogenomics and DTC
genetic testing, each one with its advantages and challenges.
The fear surrounding DNA sequencing technology, justified or not, is well and truly present. Bioethicists
warn against the dangers and perversions of this technology, while, on the other extreme, others commend
its usefulness and potential for humanity’s betterment. As with most things, the truth is probably somewhere
in the middle. Nonetheless, if humanity wants to progress with this technology, it will have to carefully take
into consideration many factors and be vigilant.
Hugues, J. N. “Apport de La Pharmacogénomique Dans l’individualisation Des Réponses
Thérapeutiques.” Gynecologie Obstetrique & Fertilite, vol. 36, Dec. 2008, pp. 6–7. EBSCOhost,
J.M. Drazen, E.K. Silverman, T.H. LeeHeterogeneity of therapeutic responses in asthma. Br. Med.
Bull., 56 (4) (2000), pp. 1054-1070
Wikipedia contributors. "DNA sequencing." Wikipedia, The Free Encyclopedia. Wikipedia, The Free
Encyclopedia, 18 Apr. 2020. Web.
Katsila, Theodora, and George P Patrinos. “Whole genome sequencing in pharmacogenomics.” Frontiers in
pharmacology vol. 6 61. 26 Mar. 2015, doi:10.3389/fphar.2015.00061
The Ubiquitous Pharmacogenomics program website. Link:
Pirmohamed, M. “Pharmacogenetics and pharmacogenomics.” British journal of clinical pharmacology vol.
52,4 (2001): 345-7. doi:10.1046/j.0306-5251.2001.01498.x
Wikipedia contributors. "Bloom syndrome." Wikipedia, The Free Encyclopedia. Wikipedia, The Free
Encyclopedia, 10 Apr. 2020. Web.
Wikipedia contributors. "Disulfide." Wikipedia, The Free Encyclopedia. Wikipedia, The Free Encyclopedia,
12 Apr. 2020. Web.
Joe S. Jeffers, Frederick Sanger, Two-Time Nobel Laureate in Chemistry, 2017, Springer editions.
Wikipedia contributors. "Messenger RNA." Wikipedia, The Free Encyclopedia. Wikipedia, The Free
Encyclopedia, 18 Apr. 2020. Web.
Committee on Science, Technology, and Law, Forum on Drug Discovery, Roundtable on Translating
Genomic-Based Research for Health, National Research Council, Institute of Medicine, Direct-to-Consumer
Genetic Testing: Summary of a Workshop. 2010. National Academies Press.
Mark Henderson, Human genome sequencing: the real ethical dilemmas, 9 Sep 2013, Newspaper The
Thurston V, Wales P, Bell M, Torbeck L, Brokaw J., The current status of medical genetics instruction in U.S.
and Canadian medical schools, Academic Medicine, 2007; 82 (5):441-445
Food and Drug Administration website, FDA allows marketing of first direct-to-consumer tests that provide
genetic risk information for certain conditions, 6 avril 2017
Shendure, Jay & Balasubramanian, Shankar & Church, George & Gilbert, Walter & Rogers, Jane & Schloss,
Jeffery & Waterston, Robert. (2017). DNA sequencing at 40: Past, present and future. Nature. 550.
Title: insulin structure, with its disulfide bonds.