Sandhills Community College Alexander the Great Re Essays - Film

Sandhills Community College Alexander the Great Re Essays - Film Sandhills Community College Alexander the Great Research Paper Shianne Buie Professor Leake HIS 111-SN 1 April sixteenth, 2017 Famous writer William Shakespeare once composed , S ome men are brought into the world extraordinary, some accomplish significance, and o thers have enormity pushed onto them . In numerous manners, Alexander III of Macedon falls into each of the three of those classes. (History Channel) During a mind-blowing course Alexander was loaded up with striking accomplishments and today is known as one of the most essential rulers ever. Because of his noteworthy extension of the Greek domain, honorable instinct as a legislator and th e l oyalty he roused in his men , Alexander the Great has earned himself the title as one of the most cultivated pioneers ever. Alexander III was conceived at some point in July of 356 BC to the leaders of Macedonia, King Phillip II and his significant other Olympias who, alongside the whole realm, had exceptionally high trusts in the k id. While the ruler would without a doubt proceed to be perceived as one of the best military prodigies ever and accumulate the biggest domain the old world had ever observed, he required some direction first. From the time he was youthful, Alexander was being prepared to one day assume control over the realm thus he spent a significant decent measure of his adolescence in the organization of mentors. He was instructed how to battle, ride like any future officer but at the same time was taught in subjects, for example, s cience, writing and theory by none of than the popular scholar Aristotle. Having this man as his instructor opened the ruler ' s brain to incalculable prospects and trusts in his future. There is in reality valid justification to accept that the savants lessons are a piece of the explanation for Alexander ' s steady want for the extension of his own domain. In spite of the fact that the matter of condition did likewise have an enormous impact in the arrangement of o ccasions that would come upon him throughout everyday life. At the point when the sovereign was only nineteen his dad, Phillip II, was killed by his guardian chief at a banquet for his little girls wedding. Alexander himself burned through no time in picking up the help he expected to guarantee the seat and once he had the sponsorship of the commanders and troops he had once battled close by, he was declared ruler. Despite the fact that still very youthful, the adolescent had just demonstrated himself to be more than fit for protecting the realm in his dads nonattendance so most had no issue tolerating him as lord. Not long after rising the seat, Alexander began on the triumph of the Persian Empire, which would get remembered perhaps the best accomplishment. Having the option to oust a domain as solid as Persia was no little feet, Alexander must be a gifted strategist, canny and have steadfast soldiers with the end goal for everything to work out. On the off chance that he had been inadequate in any of these zones, at that point the whole fight may have gone an alternate way and the world we realize today might've been totally unique. Fortunately the youthful ruler had all the devices important to prevail in this undertaking and the result ended up in support of himself. Since Persia had consistently been savage opponent of Greece, Alexander increased significantly more kindness with his kin once he declared himself the new lord in the wake of vanquishing all the contradicting powers. He would then proceed to topple Asia Minor, Egypt, India, Mesopotamia and different innumerable domains. The reality the Alexander figured out how to achieve such a great amount in just thirteen years as ruler exhibits simply was a mind blowing pioneer he was. So as to vanquish such a significant number of domains, all while guaranteeing his recently discovered subjects were content and never truly endeavored to revolt, Alexander must be a really skilled government official. Pres ently in many pieces of history once a country had been assumed control over, their way of life turned into everything except lost, their religions were prohibited and the residents were typically rewarded unfeelingly, some of the time they were even killed. Anyway this wasn't the situation with Alexander, he appeared to grasp the changed societies of each land he vanquished and even joined a few viewpoints into his own. Obviously the

Compare and contrast the 2 poems Essay Example For Students

Investigate the 2 sonnets Essay Both On my first Sonne and outcast mother and kid manage the passing of a kid. Investigate the 2 sonnets, investigating their subjects and thoughts, the artists utilization of language and your own response to them. The Poems On My First Sonne Farewell, thou offspring of my correct hand, and satisfaction; My sinne was an excess of any expectation of thee, lovd kid, Seven yeeres thowert loaned to me, and I thee pay, Exacted by thy destiny, on the only day. O, might I be able to free all dad, presently. For what reason Will man mourn the state he ought to envie? To have so soone scapd universes, and fleshes rage. What's more, if no other miserie, yet age? Rest in delicate harmony, and, askd, state here both lye BEN JONSON his best bit of poetrie. For whose purpose, subsequently forward, all his vowes be such. As what he adores may never like excessively. Ben Jonson Refugee Mother and Child No Madonna and Child could contact that image of a moms delicacy for a child she before long would need to overlook. The air was substantial with smells of looseness of the bowels of unwashed kids with cleaned out ribs and evaporated bottoms battling in toiled ventures behind blown void guts. Most moms there had since quite a while ago stopped to mind yet not this one; she held a phantom grin between her teeth and in her eyes the apparition of a moms pride as she brushed the rust-shaded hair left on his skull and afterward singing in her eyes started cautiously to part it In another life this probably been a little every day demonstration of no result before his morning meal and school; presently she did it like putting blossoms on a minuscule grave. Chinua Achebe Background on Ben Jonson Ben Jonson was conceived on the eleventh of June, 1572. He was instructed at Westminster School by William Camden and worked his stepfathers exchange, bricklaying. Jonson joined the military, disappointed with his activity, serving in Flanders. He came back to England in 1592 and wedded Anne Lewis on November fourteenth, 1595. Ben Jonson joined the showy organization of Philip Henslow as an on-screen character and a dramatist at around 1595. In 1597 Jonson was captured for his association in a parody entitled The Isle of Dogs. This was announced subversive by the specialists. After a year Jonson slaughtered a kindred entertainer. His name was Gabriel Spencer. Jonson murdered him in a duel in the Fields at Shoreditch. Foundation on Chinua Achebe Chinua Achebe was conceived in eastern Nigeria. He began his training at a congregation preacher society school. His optional and college trainings were likewise in Nigeria. He moved on from University College, Ibadan, in 1953. At that point, he worked for more than ten years for the Nigerian Broadcasting Company. He left the N. B. C. in 1966 and set out toward a profession recorded as a hard copy and instructing. Achebe assumed a critical job in the advancement of the Heinemann African Writers Series, an arrangement which has given numerous Africans a voice in the western world and which, outside of Africa, distributes progressively African (and Caribbean) scholars than some other distributing house.

The disscussion of the conceptual framework Essay

The disscussion of the applied system - Essay Example The IASB’s primary job is to help global clients to structure a worldwide system for the readiness and introduction of budgetary report, a job that has throughout the years confronted significant analysis because of the shortcomings in the destinations and the ideas that it depends on. The Framework is created to address the central purpose behind the nearness of the budget summaries, which is to give valuable money related data about the revealing substance to all the related gatherings (financial specialists, loan specialists, different leasers) for settling on choices about giving assets to the element. The substance of the Framework are focal around the five components from which budget summaries are built. This paper targets distinguishing and examining the execution of the IASB’s calculated structure dependent on its hands-on destinations for the most part that of giving a premise to managerial standard-setting and settling bookkeeping discussions in thought layou t to the bookkeeping ideas, standards and core interest. The paper is composed into three areas; the principal segment clarifies the ideas, goals and focal point of the IASB’s applied system, the subsequent segment portrays the shortcomings and the premise of the analysis confronting the IASB structure while the third segment is a decision, rundown and a conversation of the analysis confronting the current IASB’s conversation paper. As expressed, the IASB’s structure sets ideas that shape how fiscal summaries are arranged and introduced for outer clients. The IASB system needs bookkeeping standard status in contrast with the announcement of standards got from the United Kingdom Accounting Standards Board (ASB). The IASB structure contains seven fragments that spread territories, for example, targets of money related detailing, the basic suppositions, Qualitative highlights of monetary data, the major guidelines of budgetary

Essay on The Business Environment at Mcdonalds - 2230 Words

Contents Page: Table of Contents Contents Page: 2 Introduction 3 Choosing an Organization 3 Brief introduction to McDonalds 4 Aims of the Report 4 SAFE Framework 5 Strategic option 1 6 Frozen Foods 6 Strategic Option 2 11 McDonalds in a Supermarket 11 Strategic Option 3 15 McCafe 15 Conclusion 19 References 20 Appendices 21 Introduction Choosing an Organization McDonalds was chosen to be the organization as a group decision. The reason being that it was an organization recognized all over the world, having a high profile and a strong iconic brand. Given the size of the organization, it was decided that there are a lot of strategic options available to McDonalds. Brief introduction to McDonalds McDonalds was†¦show more content†¦These 3 strategic options selected by the group have been individually examined and evaluated using the framework SAFE, developed by Johnson Scholes (2008). SAFE would be used to assess whether the strategies are Suitable, Acceptable and Feasible. Looking at the evaluation of the 3 strategies using SAFE would allow identifying and determining the chosen strategic option for McDonalds. The 3 strategies are shown in figure 1: (Figure 1) SAFE Framework To evaluate the 3 strategic options the SAFE framework would be used, below explains how the SAFE framework works and what each of the stages mean: Johnson, Scholes and Whittington argue that for a strategy to be successful it must satisfy three criteria shown in figure 2 (Kfknowledgebank.kaplan.co.uk, 2014): †¢ (Figure 2) (Kfknowledgebank.kaplan.co.uk, 2014) Strategic option 1 Frozen Foods The first option available to McDonalds is frozen foods. This option would allow McDonalds to enter a new market, thus leading to potentially increasing their profit margins. Since McDonalds are successfully established, consumers may not hesitate to buy the McDonalds product frozen at their local supermarket, as this would give them a choice and time period of when they can cook it. An article from the Guardian mentions, ‘sales of frozen food have increased and more upmarket options have become available’ (Hardwick, 2014). This statement shows how McDonalds canShow MoreRelatedMcdonald Business Environment906 Words   |  4 PagesIntroduction McDonald Corporation is the world’s largest chain of hamburger fast food restaurant. There are over 30,000 McDonald’s restaurants in more than 100 countries and served an average of 50 million customer daily. A McDonald restaurant is operated by either a franchisee or by the corporation itself. The very first McDonald was open in Singapore in 1979 and today there are over 120 McDonald restaurants across the island which served 1.2 million customers weekly. McDonald employs around 9Read MoreMcdonald s The Best Food Retail Brand Of The World Essay1222 Words   |  5 PagesSumanpreet Introduction McDonald is one of the best food retail brand in the world. McDonald have almost 35000 locations worldwide and caters approximately 70 million consumers. Most of the McDonald outlets are independently owned by local business persons. McDonald has been providing sustainable supply chain to ensure benefit to its supplier as well as distributors to maximise profit.in addition to this MacDonald continuously working with charity foundations and provide sponsorships for events toRead MoreMcdonald s Strength And Weakness1280 Words   |  6 PagesAbstract McDonalds is considered to be the world’s largest and most globally known fast food restaurant, which serves approximately 69 million consumers daily. This report will evaluate McDonald’s strength and weakness in terms of their ethical claims. The main aim of this report is to consider the reasons why McDonalds has been criticized by most of its consumers in terms of its pollution to its environment and the destruction natural rain forest, furthermore this report will discuss the nutritionRead MoreCorporate Social Responsibility : Mcdonald s The World s Largest Chain Of Hamburger Fast Food Restaurants1014 Words   |  5 Pagesthis statement. CSR (Corporate Social Responsibility) is a form of self-regulation integrated into a business model whereby companies and enterprises consider the interest of society by taking responsibility of many objects such as employees, shareholders, customers, even communities and environment. President CEO of McDonald s corporation Mr.Don Thompson has stated : â€Å"We realise that our business and responsibilities to society are inextricably linked.† In fact, this firm has worked diligentlyRead MorePestle Analysis Of Mcdonalds1628 Words   |  7 PagesPESTLE analysis PESTLE is the analysis of the factors that could affect a business on a macro level. McDonalds franchise strategizes issues identified in this PESTLE analysis. The PESTLE analysis model determines the different external factors that offers opportunities or threats to the business based on its macro-environment. In the global fast food restaurant market, McDonalds is focused on economic and sociocultural factors. Nonetheless, the company’s success is indicative of its effective strategiesRead MoreHuman Resource Planning and Development Leads to Success Essay examples1396 Words   |  6 Pagesconcept can be attributed to the establishment of apprenticeship programs in the 18th Century (Werner DeSimone, 2011, p.5). The significance of human resource development in today’s business environment is attributed to its role in organizational profitability. Human resource planning involves the use of business activities to forecast how management strategy changes will impact human resource needs in the future. On the contrary, human resource development activities help an organization to ensureRead MoreMcdonalds Porters Five Forces Model Essay1184 Words   |  5 PagesAnalysing McDonalds (fast food outlets) using Porters 5 Forces model – sometimes called the Competitive Forces model. Introduction McDonalds Canada opened in 1967, thirteen years after McDonalds had taken the United States by storm. This was the first restaurant to be opened outside of the United States. It was in 1965 that McDonalds went public and offered shares on Wall Street. Since then it has been important for McDonalds to continually monitor its performance, to make sure it is competitiveRead MoreEssay about Organizational Structure- Mgt/2301127 Words   |  5 Pages The central thesis of this paper examines the organizational structures of McDonalds, Burger King, and Wendy’s food restaurants. It will examine the comparison and contrast of the organizational structure of McDonalds with Burger King, and Wendy’s Corporations. What functions influence McDonalds, and explains how the organizational design helps determine the structure that best suits McDonalds needs, as a business. Organizational Structure Burgers are one of the most favored junk foodsRead MoreThe Marketing Environment, Consumer Markets, The Worlds Largest Fast Food Chain And His Presence1214 Words   |  5 Pagesorganization as McDonalds. I will make particular reference of the marketing environment, consumer markets, market segmentation and the importance of the role of the marketing mix. In addition, I will focus on the concepts of marketing in terms of supporting their brand standards. McDonalds is known as the world s largest fast food chain and his presence is global. McDonald s has over 25,000 restaurants in the world., The organization, founded in 1940 by Richard and Maurice McDonald, has now operatingRead MoreMcdonald s Of New Zealand Essay1648 Words   |  7 PagesCompany Background And History Here I choose a McDonald s of New Zealand for my assignment. I choose this organization because it is very up growing organization of New Zealand. The biggest reason for choosing this organization is that because I am working here and I know much more about that. It all began in the USA in 1954 with a milkshake machine salesman named Ray Kroc. Ray received an order from the McDonald brothers hamburger outlet in California. He was fascinated by their operation - the

Types of Death Penalty - 1564 Words

â€Å"The death penalty for heinous crime is as justifiable, if society deem it necessary for its well-being, as is the demand upon the citizen-warrior to meet death upon the battle-field, or upon the doctor to remain steadfast at his duty in the plague-stricken city. The good of society is the prime reason for the punishment of criminals and their reformation is justifiable only when it conduces to this end† (Hall, 1902, p. 390). In recent years, death penalty has become a much talked about and controversial issue regardless of nation, culture or race. One of the earliest and known death penalty sentenced would be the crucifixion of Jesus Christ. This proves that death penalty has existed since a long time ago. Globally, death penalty is†¦show more content†¦This form of hanging is not to break the neck of the prisoners; rather letting them die of strangulation. Last of all, the most humane form would be the long drop hanging. The prisoners would be allowed to fall a pre-determined height, then jerked upwards. The prisoners’ neck break easily and immediately as the heads are still attached to the noose; thus, lessening the pain and suffering of the prisoners. Death Penalty Information Center (n.d.) has concluded the process of general hanging. The process is started when the prisoner is weighed a day before the execution and the prison officers rehearse using a bag of sand weighing the same as the prisoner. By rehearsing, the prison officers could determine the length of the rope so as not to prolong the torture and suffering of the prisoner. Before the rope can be used, it has to be treated. Firstly, the 3/4inch to 1  ¼ inch rope is â€Å"boiled and stretched to eliminate spring or coiling. The knot should be lubricated with wax or soap† (â€Å"Hanged by the Neck†, n.d.) â€Å"to ensure a smooth sliding action† (US Army Manual, 1969). Now, the prisoner is blindfolded, hands and legs bounded, and noose is placed on his neck. Then, the trapdoor underneath the prisoner is opened, allowing the prisoner to fall through. Should everything is properly in place, a rapid fracture dislocation of the neck will occur. However, Weisberg (1991) says â€Å"instantaneous death rarely occurs†.Show MoreRelatedTypes Of Punishments That Comes With The Death Penalty918 Words   |  4 Pagesare five different types of punishments that comes with the death penalty. The five methods of the death penalties are Lethal Injection, Gas, electric chair . One thing that I always think of is what each methods has upon the humans body. In this research paper you ll learn what these execution methods does to a human and also the steps these inmates go through when they are sentenced to execution. The first type of execution is the gas chamber . In the gas chamber the type of gas which is dispersesRead MoreShould The Death Penalty Be A Type Of Corporal Punishment?844 Words   |  4 Pagesa serious crime has no severe punishment, the nation should adopt a type of corporal punishment. As adults, we have grown out of spankings from our parents, but that also means that the responsibility for actions has been placed upon our shoulders. Whether it be positive or negative, there is a consequence for every action taken. Therefore, with a negative action a negative consequence should be given. While the death penalty may appear faintly harsh and inhuman, so is the killing of other humansRead MoreThe Death Penalty Is The Most Appropriate Type Of Punishment843 Words   |  4 Pagescommitting an immoral act is the most appropriate type of punishment that should be given to a convicted criminal. The name of this capital punishment is mostly referred to as, The Death Penalty, and it can be presented in different ways. This consists of being vitally shot at by a firing squad, poisoned in a gas chamber, given a lethal injection, and so on. Today in the United States, there are thirty one states that approve of the death penalty, while nineteen states believe it is unethical andRead MoreThe Death Penalty : A Multi Level Analysis Of Public Opinion1179 Words   |  5 PagesIn society, the death penalty is used to punish those who had committed certain types of crimes. Over the years, the percentage of US citizens supporting the death penalty has decrease, however Proximally 64%-70% of the population still support it. The majority of the polls created to evaluate the opinion of citizens on the death penalty use simple questions to address the issue. Those questions separate people into those against it and those supporting it. The death penalty opinion is more complexRead MoreThe Debate Over Death Penalty1618 Words   |  7 PagesReiner Writing 39C 7/20/16 Debates over Death Penalty in the United States The issue of death penalty today is a popular topic for numerous public and scholarly discussions. The death penalty has a long and distinguished history in the United States, as it has been around in some form—either official or otherwise—since the beginning of American society. America originally adopted the British justice system, with hundreds of crimes being punishable by death. Slowly but surely, states began to eliminateRead MoreDeath Penalty Is The Last Legal Resource Of Justice1472 Words   |  6 Pagesrevenge instead of retribution as the main motivating factor. This essay will explore the most credible and valid arguments that justify death penalty as the last legal resource of justice. When I write about something, I am really writing about my point of view. Then, I need to start this essay writing about who criminal really deserve death penalty. For me, death penalty is the last resource of justice, just applicable when criminals: - don’t have regrets - have the tendency to repeat the crime - theRead More Its Time to Abolish the Death Penalty Essay1187 Words   |  5 Pages The death penalty has been around since the beginning of time as a means of punishing criminals, undisputed until the last century or so in terms of whether or not it is an ethical practice. The proponents for the death penalty offer up its ability to deter crime as their main reason for supporting it, their view supported by a functionalist sociological view in that using the death penalty, enough fear will be generated that people will refrain from committing the types of crime that theRead MoreShould The Us Stop Using The Death Penalty?1599 Words   |  7 Pagesusing the death penalty? Capital punishment is a legal authorization to kill someone for a crime they have committed. The death penalty has been the highest form of criminal punishment in the American judicial system since the 13 colonies. It has taken the form of hanging, stoning, drowning, burning, beheading, gassing, electrocution, and injection. The taking of a man s life as penance for criminal behavior is wrong. The moral injustice of murder, the cruelty of execution, and the death of innocentRead MoreBanning the Death Penalty1620 Words   |  6 PagesShould the death penalty be banned internationally as a type of punishment? This form of punishment has been quite a controversial issue worldwide for quite a few years. The death penalty for hundreds of thousands of years has been a punishment for criminals throughout the world; in the past ranging from what we would now consider small crimes to huge ones, to the present where most if not all those punished with death penalty are for fairly large crime s. Actual laws involving death penalty is knownRead MoreThe Debate On The Death Penalty1597 Words   |  7 PagesThe debate on the death penalty has been the ongoing question for generations. There are plenty of different sides to take on this issue. You have the side where some people think that putting prisoners to death is inhumane and that no matter what they did putting a human to death is wrong. While others may think that if the crime is extreme enough that the prisoner who committed this crime should pay for it with their own life. Throughout history there have been several instances where prisoners

George Orwells Style of Writing Essay example - 1038 Words

Just by a simple glance at George Orwells style of writing one can grasp a lot. He first starts the essay by discussing how the narrator was poorly treated in Lower Burma when serving as a police officer. More specifically the narrator states, I was hated by large numbers of people,, and anti-European feeling was very bitter. From there one could interpret that although he was a British soldier with much power he was baited whenever it seemed safe to do so. He then goes into much detail and paints scenes into the readers mind such as how a European woman could not cross the market without having someone spit betel juice over her dress, to create a sense of remorse for the roughly treated Europeans. Furthermore, Orwell creates†¦show more content†¦The narrator then elaborates on his struggle of not making his hatred towards his job obvious; and says I hated it more bitterly than I can perhaps make clear. He even claims that it consisted of the dirty work of Empire at close q uarters. Furthermore, the narrator claims that seeing the convicts and prisoners oppressed him with suffering guilt; which in my opinion makes the reader sympathize for him being part of the British establishment; which restricts him from following his sympathy towards the Burmese. Orwells broad style of writing sways the reader into feeling how oppressed the narrator was to be part of the British establishment in Burma. Moreover, one could sense the narrators guilt of being part of the establishment and on the other hand, how he is forced to stay serene about which clearly symbolizes his lack of freedom. Orwells concise style of writing makes it clear that there was no freedom for the British in Burma whether they held a high position or no position at all. Later on, as the narrator receives a call about the elephant wrecking the bazaar, he arms himself with a Winchester rifle. I believe that a weapon here is used as a sign of the British power; since, they have the weapons unlike the Burmese who had no weapons and were quite helpless against it. As the phone rings, the narrator is put in a complex situationShow MoreRelatedBig Brother Is Watching You Essay880 Words   |  4 Pagesrealizes the true price of freedom as he sacrifices his life to attain it. In 1984 (New American Library, 1950), George Orwell, the author, proficiently integrates short, direct sentences and bleak, elementary vocabulary into his uniquely passive style of writing to thoroughly convey his appalling vision of a totalitarian regime to the reader. George Orwell’s passive style of writing incorporates short, direct sentences that vividly portray the bleakness of Oceania. Orwell conveys the appallingRead MorePolitics and the English Language: George Orwell955 Words   |  4 Pages George Orwell’s article â€Å"Politics and the English Language† gives six rules that authors should follow by to be good writers. After reading the article, one notices how often these rules are broken by authors all the time. George Orwell’s article makes readers question whether or not an author knows how to dictate their own thoughts. By breaking Orwell’s rules, an author loses attachment with a reader, and therefore will be unable to convey their message successfully. George Orwell’s first ruleRead MoreAnalysis Of George Orwell s Dystopia 881 Words   |  4 PagesOne of the most prominent examples of the hot topic today, â€Å"Dystopia†, was a novel written and published in 1949 by Secker and Warburg. Its name being â€Å"1984† by George Orwell. â€Å"Big brother is always watching,† the language the author utilizes drops subtle hints from time to time about what could possibly happen in the real world in near future. 1984 still remains one of the most intense and powerful warning signals about the peril of total government control. TheRead MoreAnimal Farm Literary Analysis710 Words   |  3 PagesGeorge Orwell, a writer of many novels and other literature, one of his most known is Animal Farm.This book is where Orwell’s political style as well as other techniques he used in his writing were used most. Animal Farm is about farm animals who are being neglected by their owner, and they decides to overthrow him and take control of their farm. However, that is only the the outer layer of the story, looking under the surface, this is an allegory detailing the Russian Revolution. The author wroteRead MoreAnalysis Of George Orwell s The Voice Of A Generation 884 Words   |  4 PagesWhat would make a story the voice of a generation? George Orwell’s pieces of writing were the voice of an age due to his style of writing. Orwell follows the principles of imagery, tone and ethos, meanwhile creating his own rules. Orwell weaves these principles together to write two famous stories that are pack ed with ethos and told in great detail. Why give a common man more credibility than an emperor? Orwell’s writing style gives him immense credibility because of the sheer detail, vivid colorRead MorePolitics And Language In Animal Farm, By George Orwell720 Words   |  3 PagesGeorge Orwell was a political writer who made it his life’s goal to expose the injustices he saw in the world. He does this through the many novels and essays he writes. 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Construction, Design and Analysis of a Single

Construction, Design and Analysis of a Single-lesi Essay on Containing Shuttle Vector for Use in Studies of Transcription-coupled DNA Repair This summer, I have had the incredible opportunity of working with ___ under the direct mentorship of ___ on designing, constructing and analyzing a single-lesion containing shuttle vector to be used for studies of transcription-coupled DNA repair (TCR). This experience has provided me with insight into the logic behind genetic engineering and experiment design and given me greater aptitude in various laboratory procedures including RT-PCR (reverse transcriptase polymerase chain reaction), plasmid transfection and recovery, plasmid transformation and amplification, and RNA isolation. It was the creativity of the design-associated aspects of our project that I found most satiating. In fact, in order to keep up with the evolution of the construction design, I learneda good deal of interesting biology that I now find the most valuable aspect of my internship. Enthusiastic about the knowledge I have gained while spending my days at the Cooper lab, I feel compelled to write this web-page with a focus that emphasizes the progression of my understanding about TCR and single-lesion containing shuttle vectors while it describes the experiments and protocol optimizations I performed. AbstractOur goal is to design a shuttle vector that contains a unique, site-specific lesion in order to study transcription-coupled repair of human DNA. In our system, the lesion is introduced by insertion of a synthesized 8-oxoguanine-containing oligomer into a pS189-derived plasmid at either of two locations: within the t-antigen (Tag) intron 400 bases beyond the ATG translation start codon, or at the end of the Tag, after the polyadenylation signal. The pS189 shuttle vector was modified to increase the transcription frequency of the Tag, prevent plasmid replication, and distinguish between Tag derived from SV40-transformed cells and that from the shuttle vector. Initial studies were undertaken to optimize the transfection conditions and also to verify the various plasmid alterations. Preliminary RT-PCR of mRNA harvested 24 hours after plasmid transfection has demonstrated that use of primers tuned to the Tag modifications do successfully distinguish plasmid from cellular RNA. Replication assays using methylation-sensitive endonucleases have verified the competence of engineered mutations in the SV40 ori in achieving preclusion of plasmid replication. RT-PCR has also shown low amplification near the Bgl II site, suggesting its removal during the processing of mature mRNA. It will therefore be necessary to construct a new site for lesion insertion before the poly-adenylation signal. In conclusion, with the competency of the pS189-derived plasmids confirmed by RT-PCR, both the shuttle vector and the transfection protocol have been optimized for TCR studies, and we are ready to insert the 8-oxoG-containing oligomer. IntroductionToward maintenance of genomic integrity and to combat the mutations and genetic degradation associated with aging and cancer, several intricate and interconnected pathways exist for DNA repair. These repair pathways include nucleotide excision repair (NER) and base excision repair (BER). Excision and repair of lesions on the transcribed strand (TS) occurs at a higher frequency than on the non-transcribed strand (NTS). This phenomenon called transcription-coupled repair (TCR) seems to be initiated by RNA polymerase II stalled at lesions or lesion-protein complexes. Lesions introduced by reactive oxygen species generated during cellular metabolism, including 8-oxoguanine (8-oxoG), are subject to TCR, even though they do not impede DNA polymerase during replication. If left unrepaired, 8-oxoG lesions can mispair with adenine and, upon replication, cause a guanine to thymine transversion with 50% frequency. Impedance of TCR results in a degenerative disease, Cockayne Syndrome (CS), characterized by postnatal developmental failure, neurological degeneration and early death. Another disease, Xeroderma Pigmentosum (XP), involves the global malfunction of nucleotide-excision repair (NER) due to mutations in the XPA-XPG proteins. Previously, investigators introduced oxidative lesions in a random, stochastic manner, making the detailed kinetics of TCR difficult to assay. .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 , .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .postImageUrl , .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .centered-text-area { min-height: 80px; position: relative; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 , .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:hover , .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:visited , .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:active { border:0!important; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .clearfix:after { content: ""; display: table; clear: both; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 { display: block; transition: background-color 250ms; webkit-transition: background-color 250ms; width: 100%; opacity: 1; transition: opacity 250ms; webkit-transition: opacity 250ms; background-color: #95A5A6; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:active , .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:hover { opacity: 1; transition: opacity 250ms; webkit-transition: opacity 250ms; background-color: #2C3E50; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .centered-text-area { width: 100%; position: relative ; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .ctaText { border-bottom: 0 solid #fff; color: #2980B9; font-size: 16px; font-weight: bold; margin: 0; padding: 0; text-decoration: underline; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .postTitle { color: #FFFFFF; font-size: 16px; font-weight: 600; margin: 0; padding: 0; width: 100%; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .ctaButton { background-color: #7F8C8D!important; color: #2980B9; border: none; border-radius: 3px; box-shadow: none; font-size: 14px; font-weight: bold; line-height: 26px; moz-border-radius: 3px; text-align: center; text-decoration: none; text-shadow: none; width: 80px; min-height: 80px; background: url(https://artscolumbia.org/wp-content/plugins/intelly-related-posts/assets/images/simple-arrow.png)no-repeat; position: absolute; right: 0; top: 0; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:hover .ctaButton { background-color: #34495E!important; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .centered-text { display: table; height: 80px; padding-left : 18px; top: 0; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5 .u4ec2d6ecb734c6d9b46b4d2362d6dfd5-content { display: table-cell; margin: 0; padding: 0; padding-right: 108px; position: relative; vertical-align: middle; width: 100%; } .u4ec2d6ecb734c6d9b46b4d2362d6dfd5:after { content: ""; display: block; clear: both; } READ: Frees: Black Success through Hard Work or Af Essay These limitations can be surpassed with the constructing of site-specific single-lesion containing shuttle vectors. Our Research GoalsTo study transcription-coupled repair of oxidative damage to DNA in various cell lines from Cockayne Syndrome (CS) and Xeroderma Pigmentosum (XP) patients, using a single-lesion containing shuttle vector. To further characterize both the requirements for XPD/XPB/XPG in TCR, and the behavior of RNA Polymerase near oxidative lesions. To further assess the degree to which lesions caused by oxidative damage (specifically