The Genomics Revolution with Either the Molecular Revolution - Essay Example

Compare and contrast the genomics revolution with either the molecular revolution. Introduction Molecular Revolution Genomics Revolution Conclusion Introduction In this assignment I am going to compare some areas of molecular revolution with genomics revolution. Cyril Darlington (1903–1981) was the most famous cytologist in the world in the decades preceding the molecular revolution of the 1950s. He crossed disciplinary boundaries to create a synthesis of cytology, genetics and evolution by revealing the mechanics of chromosomal recombination and the importance of its evolution. Darlington produced a clear set of laws that described the function that chromosome mechanics has in genetic and evolutionary phenomena. The significance of meiosis, of polyploidy, of position and proportion effects, recombination and hybridity in evolution and the genetically controlled mechanisms that regulate them were all outlined. At the beginning of the 21st century, the Molecular Revolution and the Information Age merged and gave birth to the Genomics Revolution. With the rise of genomics, the life sciences have entered a new era. Maps of genomes have become the icons for a comprehensive knowledge of the organism on a previously unattained level of complexity. Changes in technology in the past decade have had such an impact on the way that molecular evolution research is done that it is difficult now to imagine working in a world without genomics or the Internet. In 1992, GenBank was less than a hundredth of its current size and was updated every three months on a huge spool of tape. Homology searches took 30 minutes and rarely found a hit. Now it is difficult to find sequences with only a few homologs to use as examples for teaching bioinformatics. For molecular evolution researchers, the genomics revolution has showered us with raw data and the information revolution has given us the wherewithal to analyze it. In broad terms, the most significant outcome from these changes has been our newfound ability to examine the evolution of genomes as a whole, enabling us to infer genome-wide evolutionary patterns and to identify subsets of genes whose evolution has been in some way a typical. Molecular evolution research has always been opportunistic. Many scientists working in the field, do little or no work at the bench and instead rely on the public DNA sequence databases to provide the grist for our research mill. This practice dates back to the earliest evolutionary analyses on the first mRNA sequences. Consequently, many discoveries in molecular evolution have been facilitated by advances in genomics technology. Frequently, data that were not originally collected for evolutionary purposes have subsequently yielded important evolutionary insights. The flip side of this opportunism is that there have been few glimpses of a 'big picture' in molecular evolution research, despite the growing data sets. Fundamental questions, such as the relative roles of neutral evolution versus darwinian selection, have not been addressed systematically but rather in a piecemeal manner, as permitted by the available data. Darwin's theory of evolution was one of society's jewels, and Darwin himself emerged as the shining star of an era that did not want God or at least felt God was a distant and remote first cause. Science alone would solve all mankind's problems. However, we now see a Western society crying out for values and a clearly defined direction. We see a society that hungers for spirituality. Could these unfulfilled desires be attributed to the propagation and advocacy of Darwinian evolution and to a rejection of God. Could these yearnings of the spirit be explained by evolution's failure to answer our most basic questions about life and our place in the cosmos. In 1944 Avery, MacLeod and McCarthy established beyond reasonable doubt that the genetic material is DNA. From a semantic viewpoint, molecular genetics therefore has its origins in their seminal discovery of the “molecule of life”. However, it was not until restriction enzymes were uncovered some 25 years later, making it possible to manipulate the molecule of life that molecular genetics emerged as a distinct subject. It is a sobering thought that the work underpinning the discovery of restriction enzymes, in an esoteric branch of bacteriophage biology, would certainly not find favour with grant-funding agencies today. It provided, completely serendipitously, a way to isolate and manipulate DNA fragments encoding specific genes in the laboratory. The repercussions of this discovery are still being felt today. It completely changed the way many geneticists did their experiments, but most importantly, the gene ceased to be a concept and became instead a physical entity, accessible to direct experimental enquiry. The years that separated the landmark discoveries of Avery, McCarthy and McLeod and Smith, Arber and Nathans were peppered with spectacular progress and startling discoveries. In 1953, Watson and Crick solved the double helical structure of the molecule of life and about a decade later Crick, Brenner, Khorana and Nirenberg deciphered the triplet nature of the genetic code . Fred Sanger devised methods for sequencing proteins as well as the molecule of life itself, for which he was awarded two Nobel Prizes in Chemistry in 1958 and 1980. It even became possible to isolate and study a few specific genes from the model organism, Escherichia coli, using derivatives of a virus called lambda bacteriophage. Molecular Revolution Restriction enzymes were an especially important development because they finally made it possible to isolate and study any gene from any organism. These enzymes recognise specific sequences of the four bases, A, G, C and T, within DNA molecules. Those used most commonly for manipulating DNA molecules recognise palindromic sequences comprising six base pairs in double-stranded DNA and cleave within those sequences. For example, the enzyme EcoRI recognises the sequence 5’-GAATTC-3’ and cleaves between the first and the second bases of the palindrome. The complementary DNA strand has the same sequence and all molecules that have been cleaved are left with compatible “sticky-ends”. Therefore they may be recombined at will in the test tube, through the agency of an enzyme called DNA ligase, to generate novel combinations of genetic material from different sources. By incorporating DNA fragments into suitable vector molecules such as plasmids or phages, it became possible to propagate them in a suitable host (usually E. coli) and make them in reagent quantities, suitable for sequencing and many other applications. Many biotechnology companies were established on the wave of euphoria that greeted the advent of recombinant DNA technology. The winners in the rush to isolate and express genes encoding high-value pharmaceutical products (such as human insulin, growth hormone and erythropoietin) are still with us today (for details see the sixth section). We soon learned about the structure and organization of genes in a multitude of different organisms and there were several surprises in store. Perhaps the most unexpected of these was the discovery by Jeffreys and Flavell, in 1977, of a fundamental difference between the genes of lower organisms and those of higher organisms. The genes of all organisms contain a “coding sequence”, which ultimately determines the precise nature and order of amino acids incorporated into the protein product, according to the rules embodied in the (essentially universal) genetic code . In addition, there are “regulatory sequences” that lie immediately upstream and downstream, which act as instructions to the complex apparatus involved in controlling gene expression. The genes of all organisms are initially copied to make a ribonucleic acid (RNA) intermediate, the existence of which was established by Brenner, Jacob and Meselson in 1961 . This RNA molecule is then decoded to form the cognate protein product. The completely unexpected discovery of Jeffreys and Flavell was that whereas genes of lower organisms have a contiguous coding sequence, those of higher organisms are generally interrupted with stretches of non-coding DNA called “introns”. Introns must be correctly spliced out of the primary RNA copy before it can be decoded to form the protein product of the gene. This unexpected complication vitiated initial attempts to obtain correct expression of genes from higher organisms in bacteria. Fortunately, in 1970, Temin and Baltimore independently discovered a viral enzyme that converts the processed RNA molecule back into a contiguous version of the interrupted DNA molecule from which it was initially copied. This development also contributed very significantly to the growth of a vibrant biotechnology industry devoted to the production of high-value pharmaceutical products for the healthcare industry (see above and the sixth section). Molecular genetics took another quantum leap forward with the development of the polymerase chain reaction (PCR). Initially, this made it much simpler to isolate specific genes from different organisms, but a multitude of additional applications has been developed. PCR has greatly simplified the manipulation of DNA molecules and spawned many new and important analytical and clinical procedures, including DNA fingerprinting in forensic science, and rapid methods for identifying microbial pathogens and determining the disease susceptibility of patients. In fact, it is no longer possible to think of any branch of biology that has not embraced PCR technology. Methods for isolating, manipulating and sequencing DNA molecules led to the emergence of molecular genetics from within the complex conceptual framework of the study of inheritance - i.e. genetics (see above). Similarly, the development of methods for generating huge quantities of DNA sequence information robotically, together with bioinformatics tools for handling and processing these data, have now made it possible to sequence and analyze the entire genetic complement (i.e. the genome) of any organism. The name given to this enterprise is genomics. Genomics Revolution Although the term genome has only come into use comparatively recently, geneticists have been studying the chromosomes that make up genomes ever since the phenomenon of linkage was first discovered. Genomics therefore has its origins in work undertaken long before the advent of molecular genetics. Genes are effectively “labelled” by mutating them and this allowed them to be organized into groups, based on the frequency of recombination between them. Mutations on different chromosomes assort randomly, whereas those that lie in close proximity on the same chromosome tend to be inherited together and are said to be linked. Countless man-years went into the laborious construction of genetic maps representing different linkage groups (chromosomes) in a multitude of different organisms. With the advent of molecular genetics, maps went physical. Restriction enzymes with complex recognition sequences were discovered and used to generate very large fragments from genomic DNA (which must be prepared with great care to avoid shearing of the delicate molecules). Smith and Cantor devised new electrophoretic procedures for separating very large DNA fragments and even entire chromosomes according to their size. Physical maps were then constructed by using combinations of these rare-cutting restriction enzymes, together with nucleic acid hybridization methods developed in the mid-1970s by Southern. Overlapping fragments could be identified and cloned genes positioned upon them. It became possible to compare maps based on physical distance with those previously constructed, based on recombination frequency. This confirmed previous work indicating that recombination frequency is not simply determined by physical distance; some DNA sequences have a greater tendency to recombine than others do. The combined genetic and physical maps provided an essential framework for arranging segments of genome sequences during several of the early bacterial genome projects. Many different strategies have been employed for genome sequencing. In some cases, libraries of large DNA fragments have been made and sequenced systematically. Specialized vectors that accept large DNA fragments are required for this. Cosmids, for example, are special plasmids that are packaged into bacteriophage lambda particles. An ordered library of overlapping cosmids was employed for sequencing the Streptomyces coelicolor genome. Bacterial artificial chromosomes (BACs) and yeast artificial chromosomes (YACs) accept even larger fragments and they were used for sequencing the genomes of Mycobacterium tuberculosis and Bacillus subtilis, respectively. BAC, YAC and cosmid libraries are currently employed for sequencing the much larger genomes of eukaryotic organisms. Microbial genomes are now being sequenced using a random “shotgun” approach, in which the primary sequence information is obtained robotically, from a random library of small DNA fragments cloned in a plasmid vector. It is claimed that the sequencing of small (bacterial) genomes can be accomplished in less than a day. Larger ones take a little longer. The sequences of many thousands of DNA fragments (usually corresponding to coverage of between 3 and 8 times the size of the bacterial genome) are then organized into contiguous segments (contigs) and this too can be handled automatically. Thereafter the whole process becomes much more tedious. Sequencing mistakes (bases that do not match in different sequence reads) have to be corrected manually. The remaining work will depend on the sequencing strategy employed. If a library of large DNA fragments is available, adjacent contigs can be identified and a rational procedure adopted using PCR primers to amplify the missing segments between them. If such a resource is not available, primers made to both extremities of all the contigs are used in all possible combinations to fill in the gaps. As the contigs coalesce, it usually becomes increasingly difficult to close the remaining gaps, some of which may represent segments of DNA that are toxic and cannot be cloned in E. coli. As genomes near completion, they are annotated. This is undertaken “gene by gene” using bioinformatics tools, which identify genes sequenced previously that are substantially similar to the one under investigation. If the similar genes have a known (or presumed) function, then a putative function may be deduced. Since all genes have been annotated in this way, the current GenBank and EMBL genomic databases do not have completely firm foundations. The great majority of assignments have been made in the absence of any experimental evidence. Moreover, the utility of these databases will be severely compromised unless they are properly curated to remove errors and update entries, as new information becomes available. We now have a huge resource of microbial genomes at our disposal, together with vast quantities of sequence information from higher organisms, including several plants and animals . At the time of writing, more than 70 microbial genomes have been completed. These, together with accumulating data from ongoing sequencing projects, can be searched with powerful algorithms to identify genes of interest. Much can be achieved at a computer terminal, using modern bioinformatics tools. For example, by undertaking genome-wide comparisons it is possible to identify genes for which counterparts exist in all sequenced genomes, from E. coli to Homo sapiens. It came as a surprise to find that several genes in this category do not have functions ascribed to them. At the other extreme, it is possible to identify, for any particular organism, a cohort of genes that are unique to that organism and its very close relatives. Some of these may represent potential targets for novel and highly specific antibacterial compounds. We are now in what has been termed the “post-genomic era”. The genomes of most important microbial pathogens have been (or are being) sequenced. Biotechnology companies have understandably seized upon genes that are essential for bacterial growth in their search for novel anti-infectives. However, this strategy is probably misguided. Bacterial gene families are abundant and some of these probably encode functionally redundant proteins that perform essential functions collectively, although individual genes within the family may be dispensable. About 30% of the genes in any bacterial genome have no function ascribed to them. One of the challenges thrown up by genome sequencing is to develop a suite of bioinformatic and experimental approaches under the umbrella of “functional genomics” to remedy this state of ignorance. For genetically tractable organisms, mutant strains can be constructed using the tools of molecular genetics and a phenotype can be sought. Indeed, the S. cerevisiae and B. subtilis genome consortia have generated banks of strains representing the entire genome, each of which is mutated in a different gene (21). However, functional analysis is no easy matter if the organism under study is difficult to manipulate genetically. In this case, it may be more appropriate to use indirect methods such as surveys of gene expression at the level of RNA (transcriptome) and protein (proteome) under different environmental conditions, as well as metabolic fingerprinting or footprinting . Conclusion The “genomics revolution”, as it has become known, amply deserves its name. In the past, geneticists made/selected functionally altered mutants and then spent years hunting for the mutated gene responsible for the phenotypic change. The problem has now been turned upon its head. We have vast numbers of genes, for which there is no known function, and we shall spend many years filling this lacuna using tools developed under the “functional genomics” umbrella. It is widely accepted that biotechnology is the science of the 21st century. What new and unexpected discoveries lie just around the corner? Both the molecular and genomics revolution would answer to this question with the passage of time. 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