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CfE Higher Human Biology Unit 1 Chapter 5 - Human Genomics Human Genomics • Human genomics is the study of the human genome. It involves determining the sequence of the nucleotide base molecules all the way along the DNA and then relating this genetic information about genes to their functions. Human Genome Project • Started in 1986 (USA and UK) but officially ‘started’ in 1990, Europe and Japan joined in 1992 • Completed in 2003 – under budget and 2 years early • The sequence is not that of one person, but is a composite derived from several individuals. Therefore, it is a "representative" or generic sequence. To ensure anonymity of the DNA donors, more blood samples (nearly 100) were collected from volunteers than were used, and no names were attached to the samples that were analysed. Thus, not even the donors knew whether their samples were actually used. • Also sequenced yeast and animals used in medical research e.g. zebra fish and rats. What were the aims of the human genome project? • To identify all the approximately 20,000-25,000 genes in human DNA. • To find where each gene is located • To determine the sequences of the 3 billion chemical base pairs that make up human DNA. • Store this information in databases. • Estimated time 15 years. (started in 1980) • Estimated cost US$3 billion Human Genomics • The sequence of bases can be determined for individual genes and entire genomes • This genetic information can be used to find the function of different genes. • Entire genomes can be compared using single nucleotide polymorphisms (SNPs). (These are differences between individuals due to base substitutions. ) SNPs Sequencing DNA A portion of DNA with an unknown base sequence is chosen to be sequenced. Many copies of one of this DNA’s strands (the template) are synthesised. Then, in order to make DNA that are complementary to these template strands, all the ingredients needed for synthesis are added to the preparation. These include DNA polymerase, primer and the four types of DNA nucleotide. In addition the preparation receives a supply of modified nucleotides (ddA, ddT, ddG and ddC), each tagged with a different fluorescent dye. Every so often during the synthesis process, a molecule of modified nucleotide just happens to be taken up instead of a normal one. However, when a modified nucleotide is incorporated into the new DNA strand, it brings the synthesis of that strand to a halt because a modified nucleotide does not allow any subsequent nucleotide to become bonded to it. Provided that the process is carried out on a large enough scale, the synthesis of a complementary strand will have been stopped at every possible nucleotide position along the DNA template. • The DNA Fragments of various lengths(each with its modified nucleotide and its unique fluorescent tag) are separated using electrophoresis. The order of the nucleotides in the original DNA can then be worked out from the fluorescent dye that is used. • Usually a computer working as an automated sequence analyser, detects the four fluorescent dyes processes this information quickly and displays the sequence of bases in the DNA sample. genome sequencing animation A variation in DNA sequence that affects a single base pair in a DNA chain is called a single nucleotide polymorphism (SNP). SNPs are one of the ways in which genomes are found to differ from one individual to another. For example, the DNA of two people might differ by the SNP shown below. This difference has arisen as a result of a point mutation where one base pair has been substituted for another. Two out of every three SNPs involve the replacement of cytosine (C) and with thymine (T). SNPs can occur in coding and non-coding regions of the genome. SNP maps • Scientists have managed to catalogue more than a million SNPs. They believe that an SNP map will help them identify and understand the workings of genes associated with disease. • Some SNPs may indicate the likelihood of a person developing a particular illness. Alzheimer’s Disease • One of the genes associated with this disease is called ApoE (Apolipoprotein E) • The gene is affected by two SNPs and different combinations of these produce three different forms of the gene (Apo E2, Apo E3 and Apo E4). • Research has shown that inheritance of the ApoE4 allele increases the chance of Alzheimer’s disease whilst inheritance of the Apo E2 allele makes the person less likely to develop the condition. Bioinformatics is the name given to the fusion of molecular biology, statistical analysis and computer technology. It is an ever-advancing area that enables scientists to carry out rapid mapping and analysis of DNA sequences on a huge scale and then compare them. • Computer programs can be used to identify – Gene sequences by looking for coding sequences similar to know genes – Start sequences (there is a good chance that each of these will be followed by a coding sequence – Sequences lacking stop codons (a protein coding sequence is normally a very long chain of base triplets containing no stop codon except the one at its end Videos • BBC documentary (1 hour) https://www.youtube.com/watch?v=Fgq-XoyorWY • Ethics of genome project (5 mins) http://www.bbc.co.uk/learningzone/clips/ethics-of-thehuman-genome-project/8732.html • History of the world • https://www.youtube.com/watch?v=q13WCTOFJvo Learning Intentions • Describe what is meant by systematics • Explain the role of genomics in personalised medicine • Explain what pharmacogenetics is and how it is used Systematics Video This can be defined as the study of a group of living things with respect to their diversity, relatedness and classification. Data obtained by comparing human genome sequences are used in systematics to study the origins of modern humans and their evolutionary relationships. Unlike other primates such as orangutans, hose DNA differs among the members of the species by around 5%, the mean difference in genomic sequence among the members of the human race is only about 0.3%. This high degree of similarity indicates that all humans are more closely related to one another than other types of primate. Careful examination of the genetic differences that do exist between different human populations shows that the greatest variation occurs among populations in Africa rather than those on other continents. Furthermore, genetic evidence indicates that all human populations outside Africa possess only a small part of the total genetic diversity found among African populations. These findings support the “out of Africa” theory. This propses that humans originated in Africa and underwent early evolutionary divergence in that continent over a very long period of time (e.g. mllions of years) to form a variety of genetically different populations. Then small groups migrated out of Africa relatively recently (e.g. 100,000 yeards ago) and gave rise to all other human populations. Personal Genome Sequence Personalised Medicine A complete sequencing of a person’s DNA bases is called a personal genome sequence. The branch of genomics involved in sequencing the genomes of individuals and analysing them using bioinformatics tools is called personal genomics. As a result of advances in computer technology, the process of sequencing DNA is rapidly becoming faster and cheaper. Sequencing an individual’s DNA for medical reasons will soon become a real possibility. In years to come, a person’s entire genome may be sequenced early in life and stored as an electronic medical record available for future consultation by doctors when required. Harmful and Neutral Mutations Having located the mutant variants present in the genome, it is important to distinguish between those altered sequences that are genuinely harmful. Pharmacogenetics This can be defined as the study of the effects (therapeutic, neutral or adverse) of pharmaceutical drugs on the genetically diverse members of the human population. Already it is known that one in ten drugs (e.g. the blood thinner warfarin) varies in effect depending on differences such as SNPs in the persons DNA profile. In the future it may be possible to use genomic information and Customise medical treatment to suit an individuals exact metabolic requirements. The most suitable drug and the correct dosage would be prescribed as indicated by personal genomic sequencing. Ideally this advance would increase drug efficacy while reducing side effects and “onesize-fits-all” approach would be consigned to history. Amplification and Detection of DNA SequencesPolymerase Chain Reaction (PCR) The polymerase chain reaction (PCR) is a technique that can be used to create many copies of a piece of DNA in vitro (i.e. outside the body of an organism). This amplification of DNA involves the use of primers. In this case, each primer is a piece of single-stranded DNA complementary to a specific target sequence at the 3’ end of the DNA strand to be replicated. Amplification and Detection of DNA SequencesPolymerase Chain Reaction (PCR) The DNA is heated to break the hydrogen bonds between base pairs and separate the two strands. Cooling allows each primer to bind to its target sequence. During the next step, heat-tolerant DNA polymerase adds nucleotides to the primers at the 3’ end of the original DNA strands. Amplification and Detection of DNA SequencesPolymerase Chain Reaction (PCR) The first cycle of replication produces two identical molecules of DNA Amplification and Detection of DNA Sequences- Polymerase Chain Reaction (PCR) During the second cycle, the two DNA molecules from the first cycle split and then nucleotides are added. By the end of the second cycle, four identical molecules are produced.. This will continue and the population of DNA molecules will grow exponentially. By this means a tiny quantity of DNA can be greatly amplified and provide sufficient material for forensic and medical purposes. Amplification and Detection of DNA SequencesPolymerase Chain Reaction (PCR) By the end of the second cycle four identical DNA molecules are made. DNA Probes and Arrays One of the meanings of the word array is an orderly arrangement of many items. A DNA microarray is an orderly arrangement of thousands of different DNA probes as tiny spots attached to a glass slide. A DNA probe is a short, single stranded fragment of DNA. It is used to detect the presence of a specific sequence of nucleotide bases in a sample of DNA. The DNA under investigation is called the target DNA. A probe is able to carry out its function because its sequence of bases is complementary to the specific base sequence to be detected in the target DNA. Fluorescent labelling indicates those spots where a probe has successfully detected and combined with its complementary sequence on the target DNA. Medical and Forensic Applications of Amplified DNA- Genetic testing for Cystic Fibrosis In the UK, it is estimated that 1 in 25 people carries a recessive mutant allele for cystic fibrosis. Carriers do not suffer the disease but if two carriers produce a child, they risk a 1 in 4 chance of the child suffering cystic fibrosis. Therefore couples planning a family may decide to be tested for the presence of a mutant allele for cystic fibrosis in their genome. This can be done by genetic testing using blood cells. DNA is amplified and tested for the presence of a mutant allele using genetic probes. Genetic testing for cystic fibrosis Forensics Each human genome possesses many non-coding regions of DNA composed of a number of repetitive sequences Each repetitive sequence is unique to an individual These regions can be used to construct a person’s DNA profile, ‘genetic fingerprint’ Forensic scientists amplify DNA samples from a crime scene as well as DNA from the victim and suspects The components of the samples are compared Crime Scenes Forensic Scientists make use of the PCR reaction to amplify DNA samples at a crime scene. DNA samples are taken from the victim and the suspects are also amplified. Next components of the samples are separated using gel electrophoresis and then compared. Paternity Disputes PCR followed by gel electrophoresis can also be employed to generate genetic profiles from DNA samples and confirm genetic relationships between individuals. Each person inherits 50% of their DNA from each parent therefore every band in their DNA profile (genetic fingerprint) must match one in that of their father or their mother. The fact that each person has 50% of their bands in common with each of their parents allows paternity disputes to be settled. Evidence based on DNA amplified by PCR has also been used to identify missing people from human remains left at the site of a disaster and to secure the release of innocent people who have been wrongly imprisoned. Is he the father? Every band in a person’s ‘genetic fingerprint’ must match one in that of their father or mother! Mother Child Father Now answer the following questions 1. In PCR, what is a primer 2. Why is DNA heated during PCR process? 3. What is the purpose of cooling the DNA samples? 4. What is a genetic probe used for? 5. What feature of the human genome makes each individual unique and allows genetic profiles to be constructed? 1. In PCR, what is a primer A piece of single-stranded DNA complementary to a target sequence at the end of the DNA strand to be replicated 2. Why is DNA heated during PCR process? To break the H bonds to separate strands 3. What is the purpose of cooling the DNA samples? To allow primer to bind to its target 4. What is a genetic probe used for? To detect for the presence of a specific sequence of nucleotide bases in a sample of DNA 5. What feature of the human genome makes each individual unique and allows genetic profiles to be constructed? Each human genome possesses many non-coding regions of DNA composed of a number of repetitive sequences, each repetitive sequence is unique to an individual