The Human Genome Project - Essay Example

The Human Genome Project Introduction In 1985, meetings were held at the University of California with the aim of outlining the practical task of sequencing the human genome, the scientific and new technological environment of the 1980s catalyst these meetings. During this period, DNA cloning as well as Fred Sanger’s sequencing techniques, formulated in the 1970s were being applied by scientists who believed that sequencing the human genome was probable at an experimental stage. More importantly, researchers were, simultaneously, starting to use computing applications to DNA sequencing and genetics, formulating techniques that would make viable the work of generating and managing genetic data worldwide. This ostentatious, fresh concept, a “Human Genome Project” attracted ardent supporters, who asserted that decoding the human genome would result in better understanding and advantages for human health. However, some scientists opposed the move fearing that this kind of project would lead to a product that would be of little benefit to human beings. This project thus brought heated controversy that involved politics and personalities. This paper will describe how the sequence of the human genome was determined, and try to answer this important question, has the Human Genome Project lived up to expectations?" The Human Genome Project The Human Genome Project began in 1990, with the aim of establishing the DNA sequence of the whole euchromatic human genome in a period of 15 years1. At the initial stage, the project was filled with scepticism from scientists and even ordinary people. One major issue raised was whether the enormous finances involved in the project would result in corresponding benefits. Nonetheless, the great success of the project is clear, the completion of Human Genome project brought a new age in medicine and also resulted in major advancements in the forms of technology applied in sequence DNA2. The U.S National Institute of Health (NIH) and Department of Energy funded the launch of Human Genome project in 1990.3 The labs of this project worked with international collaborators and they were able to resolve 95% of the sequence of DNA in human genome in a period of 15 years. Similarly, John Sulston in association with his colleagues at the MRC‘s laboratory of Molecular Biology in Cambridge in the UK were working at mapping the genome of the nematode worm for a number of years. From their findings, they revealed that sequencing the entire genome of the worm was feasible4. Human Genome Project in the US, it progressed well in its operations. In the United Kingdom, the MRC approached the Wellcome Trust and proposed for a new partnership to enable them get funds to facilitate the worm sequence project that John had proposed. This project was to be used as a channel for the Human Genome Project. After this proposal, things began to move quickly because the Wellcome Trust embarked its operations in the UK, since the project was promising; they appointed one of their senior administrators to analyze the viability of the sequencing initiative. In due course, John Sulston submitted a grand application of between 40 and 50 millions pounds in 1992 aimed at funding a new centre called Sanger centre. He proposed for the formation of this centre in UK to act as a British arm of the Human Genome Project to enhance sequencing efforts.5 Original principles and goals of the project From its beginning, Human Genome Project was based on two main principles, these were ADDINEN.CITE, and the project as well aimed at availing all human genome sequence information publicly within a day after it has been assembled. This initial principle guaranteed unlimited access for the scientists both in academic and industries, providing the way for quick and new discoveries by various researchers. In the course of the project, nearly 200 laboratories in America were financed by National Institutes Health or by U.S Department of Energy. More so, over 18 countries around the world contributed towards the project. The same way the Human Genome Project involved two main principles, it as well had two initial goals, one, developing genetic and physical maps of the human and mouse genomes, two, sequencing the simpler yeast and worm genomes used as test samples for sequencing the highly complex human genome.6 After being successful with the sequencing of yeast and the worm, the human genome sequencing followed. Stages of the Human Genome Project Basing on the knowledge obtained from the yeast and worm tests and studies, a two-phase method was applied by the Human Genome Project to address the human genome sequence7. The first phase referred to as shotgun phase involved dividing the human chromosomes into DNA divisions of proper size; these were later subdivided into much smaller, overlapping DNA pieces that were sequenced. As explained by DeLisi the Human Genome Project depended on the physical map of human genome developed earlier on, which acted as a foundation for generating and examining the huge volume of DNA sequence information that was generated from the phase. 8 The second phase referred as finishing phase, entailed filling in gaps as well as determining DNA sequences in unclear areas not gotten in shotgun phase. As observed in Figure 1, there is exponential raise in DNA sequence information placed in the High-Through-put Genomic Sequence (HTGS) department of GenBank at the completion of the shotgun phase. In fact, the shotgun phase resulted in 90% of the human genome sequence as draft type. The shotgun phase in this project comprised of three stages, getting a DNA clone to sequence, sequencing the obtained DNA clone and assembling sequence information from various clones to establish overlap and determine a contiguous sequence. Figure 1: Total amount of human sequence in the High Trough-put Genome Sequence (HTGS) The concept applied by the members of this project was referred to as hierarchical shotgun approach, since the team members methodically produced overlapping clones that were mapped to specific human chromosomes that were specifically sequenced basing on a shotgun method. The clones were obtained from DNA libraries developed by ligating DNA pieces created through partial restriction enzyme digestion of genomic DNA from unknown human donors to bacterial artificial chromosomes vectors that are propagated in bacteria.9 Where possible, the DNA pieces in the library vectors were then mapped to chromosomal regions through screening for sequence-tagged sites (STSs) that are DNA pieces, normally below 500 base pairs long, of identified sequence and chromosomal location, which can be amplified by use of polymerase chain reaction (PCR). At the same time, library clones were as well digested with restriction enzyme Hind III, agarose gel electrophoresis was used to determine the sizes of subsequent DNA pieces. Each one of the library clone showed a DNA piece “fingerprint,” which could later be compared to those of other library clones so as to determine overlapping clones10. Fluorescence in situ hybridization (FISH) was as well applied to map library clones to particular chromosomal regions. Together, the STS, DNA fingerprint as well as FISH data permitted the IHGSC to provide contigs, which comprised of several overlapping bacterial artificial chromosomes (BAC) library clones covering each of the 24 varied human chromosomes (22 autosomes and the X and Y chromosomes). According to Venter, after this, individual BAC clones chosen for DNA sequence examination were further divided11, and the smaller genome DNA fragments were then sub-cloned into vectors to create a BAC- develop shotgun library. Venter explains that the inserts were sequenced by primers similar to the vector sequence bordering the genomic DNA insert12, and also the overlapping shotgun clones were applied to create a DNA sequence covering the whole BAC clone. Figure 2 gives a summary of processes involved in this step. Figure 2: Levels of clone and sequence coverage (source) Munson observes that before the IHGSC had finished the initial stage of the Human Genome Project, Celera Genomics (a private biotechnology company) as well got involved in sequencing the human genome.13 Celera Genomics was led by Craig Venter, the company claimed that it had the ability to sequence the whole human genome in a period of just three year. Venter et al. notes that Celera applied two independent data groups alongside two specific computational methods to establish the sequence of the human genome.14 The first data group created by Celera comprised of 27.27 million DNA sequence reads, and each had was approximately 543 base pairs, obtained from five separate individuals, while the second data group was derived fro the publicly financed Human Genome Project and was obtained from the BAC contigs (referred to as bactigs); in this data group, Celera “sliced” the Human Genome Project DNA sequence into 550 base-pair sequence reads that represented a net total of 16.5 million sequence reads. Celera later employed entire-genome assembly approach and a regional chromosome assembly approach to sequence the human genome.15 Venter et al. explains that in the entire-genome assembly approach (also referred to as whole-genome random shotgun approach),16 Celera developed a huge shotgun library generated from its individual DNA sequence data together with the “sliced” Human Genome Project DNA sequence data, which combined accounted for 43.32 million sequence reads. The company applied computational techniques and complex algorithms to determine overlapping DNA sequences as well as to re-structure the human genome by developing a set of scaffolds. In comparison to the regional chromosomes assembly method (also known as compartmentalised shotgun assembly approach), Celera arranged its own data and that of Human Genome Project sequence into the biggest possible chromosomal segments, then, a shotgun assembly of the sequence data in each segment was done17. As pointed out by the IHGSC this method was same as the one used in hierarchical shotgun method employed by the IHGSC.18 The IHGSC adds that the initial phase of the regional assembly method entailed separating Celera reads, which was similar to Human Genome Project reads from those which were different from the public sequence data.19 Afterwards, the reads were assembled into Celera-specific and Human Genome Project-specific scaffolds; these were later combined and examined by entire-gene assembly algorithms. The subsequent bactig data were once more “sliced” to all unbiased assembly of the combined sequence data. Celera’s entire-genome and regional chromosome assembly approaches were autonomous of each other, allowing express comparison of the obtained data. Venter notes that Celera established that the regional chromosomes assembly approach was somewhat more reliable than the entire-genome assembly approach20. Employing these complementary methods, Celera developed data that was highly in agreement with results from the IHGSC.21 As indicated by IHGSC (2001) as at February 2001, the drafts of human genome sequence were then published by Celera and the Human Genome Project in different articles. Owing to technical advancement in DNA sequencing techniques and productive degree of synergy among the two groups, they both finished at the same time, well ahead of the planned schedule. An important message in DNA sequencing As earlier discussed, the IHGSC and Celera employed varied methods to establish the sequence of the human genome. Nonetheless, Hood and Galas asserts that they employed similar technique for the DNA sequencing.22 The technique they used employs DNA polymerase, a similar enzyme employed in DNA replication, to generate DNA sequence information. As indicated in Figure 3a, DNA polymerase attaches itself to a single-stranded DNA template and inserts DNA bases to the 3’ terminal of the complementary DNA strand it creates. DNA polymerase needs primer that has a free 3’ terminal to which it inserts another DNA bases in a 5’ to 3’ approach, and it progress along the template strand in a 3’ to 5’ direction23. Scientists from IHGSC and Celera put together the DNA template as they were more concerned in sequencing with the DNA polymerase, which is a single-stranded DNA primer, and free deoxynucleotide bases, and a sparse mixture of fluorescently labelled dideoxynucleoide bases (dATP), which were labelled with a specific colour and would end new DNA strand synthesis the moment integrated into the terminal of a developing DNA strand. According to Chial the mixture was initially heated to de-nature the cut-out DNA strand, it was then cooled to enable the DNA primer to anneal24. After the annealing of the primer, the polymerase created a complementary DNA strand. Lastly, the template grew in length allowing dideoxynucleotide base (ddNTP) to be integrated; the prevailing conditions allowed this integration to occur randomly along the length of the freshly created DNA strands. At the end, researchers involved in this project remained with a mixture of freshly synthesised DNA strands that had different length by one nucleotide, and were marked at their 3’ terminal with ddNTP-related colour dye molecule (as shown in figure 3b). Figure, 3: How to sequence DNA (Source, H, Chial, 2008) For the researchers to establish the sequence of the newly created colour-coded DNA strands, they required a method to separate them basing on their size, the difference was merely by a single DNA nucleotide. The researchers managed to this separation by electrophoresing the DNA through a coagulate matrix that allowed single-base variations in size to be readily differentiated. Small pieces went through the coagulate matrix more rapidly, while the larger pieces went through slowly. Through putting the whole mixture in the coagulate, a laser beam was used to check the DNA bands as they passed through the coagulate and establish heir colour, the data collected from this observation was applied to develop a sequence trace (referred to as electropherogram), which showed the colour and the signal strength of every DNA band passing through the coagulate (as shown in Figure 3d). As explained by Venter, the colour of every band represented the last 3’ base integrated at that location, and through reading from the start to the end of the coagulate, the researchers could establish the sequence of the freshly created DNA strand starting from 5’ to the 3’ terminal25. The Rough draft is transformed to the Final copy The IHGSC after completing the draft stage of the Human Genome Project started the second stage of the project; the final or finishing phase. In the final phase, Venter points out that the researchers were able to fill the gaps and resolve DNA sequences in confusing areas that had not been resolved in the shotgun phase26. According to IHGSC (2004) the final phase resulted in 99% of the human genome in their final form, it as well comprised 2.85 billion nucleotides, with an error margin of 1 occurrence for 100,000 bases sequenced.27 The IHGSC cut down the cases of gaps by 400-times, just 341 gaps from a total of 147,821 gaps remained28. IHGSC explained that the remaining gaps were due to technical difficulties in chromosomal regions. Though the initial draft publications had projected up to 40,000 protein-encoding genes, the final phase decreased this projection to about 22,000 protein-encoding genes. According to the IHGSC the future challenges at this phase entailed the identifying polymorphisms as a model for interpreting genetic relations to human disease, identifying functional elements in the genome concerned with gene control and identifying gene and protein “components” that act in agreement with each other.29 Using the digital information in molecular medicine One mainly outstanding finding in this Human Genome Project is the fact that human nucleotide sequence is almost alike (99.9%) among any two given individuals. Nonetheless, a single nucleotide variation in a one gene can end up causing a certain human disease. Owing to this aspect, our understanding of the human genome sequence has at the same time contributed extensively to our knowledge of molecular systems underlying in numerous human diseases. More so, integration of cytogenetic concepts with the human genome sequence will keep on advancing our knowledge of human diseases to a whole new understanding. Therefore, though the project was initially faced with scepticism, the Human Genome Project certainly become one of the most significant scientific happenings of modern era. Has the Human Genome Project lived up to expectations? The main aim of the Human Genome Project was to “create a high quality reference DNA sequence for the human genome’s three billion base pairs and to recognise all human genes.30” However, the project had other aims that including sequencing the genomes of sample organisms to understand the human DNA, improving the computational resources to aid future studies and commercial use, analyzing gene function by carrying out mouse-human evaluations and comparisons, examining human differences, and training up coming scientists in human genomics. The Human Genome Project is among the most ambitious scientific projects that have been undertaken. When it comes to the deadline of the project, the project surpassed its expectations by being completed two years before of its initial schedule. The Human Genome Project has provided a remarkable and unparalleled biological resource in human genome that will be used through out the century as a foundation of research, application and discovery31. Until now, the genetic information acquired by researchers during this project has presented scientists with more knowledge of the link between particular genes and certain human diseases. Doctors have as well been given an opportunity to apply the genetic information available in the prevention as well as treatment of incapacitating diseases caused genetically. More so, information regarding a person’s predisposition to certain diseases will be able to be discovered through genetic screening. Therefore, a person can be alerted and seek early medical interventions in order to get proper treatment. Individuals can as well know environmental risks that could trigger their disease. Conditions like Parkinson’s and diabetes, where some organs have failed to secrete required substance, could be eradicated by introduction of healthy genes in these organs. Another important advancement made possible through the information gotten from the Human Genome Project is pharmacogenomics, this entails manufacturing and production of custom-made medicine to treat certain group of people in accordance to their genetic composition. According to the U.S Department of Energy, in the next ten years, pharmaceutical will start relating DNA variants with responses of an individual when receiving medical treatments. Besides all these expected benefits, and possibilities that Human Genome Project was supposed to bring, it is clear that some goals of the project were not achieved. As noted by Venter the outcome for clinical medicine from the project has nonetheless been minimal32, Venter adds that it is logical and fair to assert that the Human Genome Project has failed at the moment to directly affect the healthcare of many people33. Venter on contributing to this issue also agrees that still a lot to achieve before this potential can have a major impact on healthcare and medicine.34 Lewontin also points out that the genetic connection of human diseases has turned out to be far complicated than originally projected, when the Human Genome Project goals were made35. Lewontin further adds that the Human Genome Project has not expressly resulted in new drugs; neither has it been able to accurately predict disease risk caused by genetics in a manner, maybe, initially thought would be possible36. Though, he observes that these goals may be achieved in future. Though, the early hope of hastening the breakthrough of new treatments for human diseases was not basically achieved by the Human Genome Project. However, with knowledge about the sequence of human genome, we now understand that it needs more than knowledge of how base pairs are arranged in our genome to treat diseases. Presently, researchers have focused their efforts in knowing the protein products encoded by human genes. Victor points out that the corresponding protein of a mutated gene is usually defective37. Thus, up-coming field of proteomics has aimed at understanding the manner in which protein functions as well as expressions that are changed in human disease conditions. In addition, researchers are as well turning their focus on the broad areas of human genome without usual protein-encoding genes. The world has already begun to enjoy the benefits of human genome knowledge, and as DeLisi asserts future data-mining endeavours will most definitely reveal many more thrilling and unexpected connections to human diseases. 38 Conclusion Within a period of just 13 years, mixture public and private researchers managed to successfully carryout the Human Genome Project. Though different methods and approaches were used by these researchers, they nevertheless got similar results. For example, in 1995, researchers from the Sanger Centre in association with international collaborators identified a gene called BRCA2. This gene is responsible for increasing breast cancer. On the other hand, a team from the U.S identified a gene called MSH2 that increases the risk of cancer for individuals who have this gene. Elsewhere, in Canada, researchers located five varieties on the FAD gene that together confer an almost 100 percent risk of developing a disease called Alzheimer. By doing so, these researchers answered back their critics and at the same time managed to beat the set deadline of the project by two years. Though the project did not full accomplish its goals and initial set out, the findings from the project have provided very important information concerning the human gene and the link they have on human diseases. Indeed, we are presently, enjoying the benefits of these findings in molecular and medicine fields. Maybe, even more important aspect is that, these scientists were able to inspire a continuous revolution in the fight against human diseases, and at the same time provided a fresh vision about the future of medicine, though we are yet to wholly realize the future. Bibliography Chial, H: DNA sequencing technologies key to the Human Genome Project. Nature Education, 2008, 1(1) DeLisi, C: Genomes: 15 Years Later A Perspective by Charles DeLisi, HGP Pioneer: Human Genome News 11, 2001: 3–4 Hood, L. & Galas, D. The digital code of DNA. Nature 421, 444–448 (2003) IHGSC: Finishing the euchromatic sequence of the human genome: Nature 431 (7011): 2004; 931–945. International Human Genome Sequencing Consortium. Initial sequencing and analysis of the human genome. Nature 409, 860–921 (2001) International Human Genome Sequencing Consortium: Finishing the euchromatic sequence of the human genome. Nature 431, 931–945 (2004) Lewontin, R.C. (1991). Biology as Ideology, New York: Harper Perennial. Munson, R. (1996). Intervention and Reflection: Basic Issues in Medical Ethics, 5th edition. National Human Genome Research Insititute (2008) The Human Genome Project Completion: Freuently asked Questions U.S. Department of Energy (2003): Genomics and its impact on science and society: 2003 Primer.”http://www.ornl.gov/sci/techresopurces/Human_Genome/public/primer2001/2shtml. Retrieved on 2011-09-06. Venter D: A Part of the Human Genome Sequence. Science 299 (5610): 2003, 1183–4. Venter, J. C., et al. The sequence of the human genome. Science 291.2001. 1304–1351 Victor K. M: Drawing the Map of Life: Inside the Human Genome Project.Basic Books; 2010 Read More