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An Antibiotic Resistance Gene Using Recombinant DNA

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An Antibiotic Resistance Gene Using Recombinant DNA. Introduction Recombinant DNA is the joining together of DNA molecules from two different molecules that are inserted into a host organism to make a new genetic combination. They are used in science, medicine, agriculture, and industry. Recombinant DNA was first brought out in 1980, it has helped a lot in the advancement of science. DNA recombination involves the process of cutting and joining DNA of different species through restriction enzymes and ligations (2). The goal of this method is to have an antibiotic resistance gene using recombinant DNA. When it’s inserted into a host cell, it allows the bacteria to grow. The antibiotic gene is formulated from two relaxed plasmid reagents, pKAN and pUC19, each of which contains a single antibiotic resistance gene. These plasmids are digested in a pre-constructed hybrid restriction enzyme mixtures of BamHI and HindIII (3). Each plasmid has a sequence where each enzyme cuts, making two restriction fragments which are produced and analyzed through a process called electrophoresis. Electrophoresis is a laboratory method used to separate mixtures of DNA, RNA, or proteins according to molecular size through the passing of electric current (1). The cutting ability of hybrid restriction enzymes is confirmed and enzyme fragments are separated from the DNA fragments through electrophoresis. On the other hand, ligation is a method that requires the presence of ATP and the ends of the fragments are fused by ligation (1). These ligations occur by hydrogen and phosphodiester bonds, which is what forms the recombinant DNA molecule. This recombinant DNA then goes through transformation with E. coli. Transformation is done in order to be able to select the antibiotic resistant gene. The colonies on the plates show how successful the transformation was and if the resistance was accomplished. X-Gal is used in the transformation process for identification purposes. It is a blue/white screening technique that allows for indication of whether a cell expresses a functional B-galactosidase enzyme in blue/white screening. The phenotype is then identified by conducting a purification procedure (4). Materials and Methods Materials pUC19 (0.20 ug/ul) pKAN (0.20 ug/ul) pUC19 (0.1 ug/ul) pKAN (0.1ug/ul) agarose gel BamHI/HindIII Ethidium bromide Loading dye Restriction buffer 1x TBE Beakers Electrophoresis supplies PPE equipment Micropipettor plus tips Tubes Water bath Recombination of antibiotic resistance genes Two plastic white centrifuge tubes were obtained and labeled; one pKAN and the other pUC19. Tubes were prepared according to table below. Sample Mini prep DNA 10x Buffer RNase BamHI/HindIII H2O pKAN 8uL 2uL 2uL 4uL 8uL pUC19 8uL 2uL 2uL 4uL 8uL After preparation, the tubes were incubated for 30 minutes at 37C. Once digestion was completed, the tubes were removed and spun down. 5ul of sample was removed from tube and placed in a fresh tube and 1ul of loading dye was added, and the tube was then placed in the freezer. This was done for both of the tubes. The digestion tube was put back into incubator for an additional 15 minutes. While waiting for that the gel was run and the 0.1ug/ul uncut plasmid was provided. When the 15 minutes digestion was complete, the tube was put into the digestion tub in 80C for 20 minutes to start the ligation process. Once the 20 minutes was complete, it was taken out and kept at room temperature for five minutes, then it was prepared according to the chart below. Ligation: Sample dH2O 10X ligation buffer Digested pKAN Digested pUC19 DNA ligase ligation 10uL 3uL 3uL 3uL 2uL The tube was incubated at room temperature for one hour then 4C for overnight incubation. Then the samples were stores at 0C until next class. For ligations, ATP is used because the energy given off of ATP is required. A general molecule of DNA is stable itself and doesn’t tend to bond with other DNA molecules: the 3′ end ends with a -OH group, while the 5′ end ends with a phosphate group. The DNA ligase then removes the OH– and binds the two segments together. Materials CaCl2 (50mM) Mid-log MM294 cells (two 10ml cultures) Beakers for ice Bunsen burner Centrifuge Micropipettor Pipettes Water bath Shaking incubator Sterile loop Transformation: While performing this exercise it was important to make sure everything was on ice throughout the experiment. Three plastic tubes were labeled the following way; lig, pUC19, and pKAN. The cells were added to each of the tubes, then the pDNA was added to the tubes and placed on ice. All tubes should stayed on ice for 15 minutes. The cells were carried in a beaker of ice to the 42C where cells were heat shocked for 90 seconds. Following that, they were returned to ice for one minute. Next, 500uL of LB broth was added to the tubes with the p1000 pipettes. And then the tubes were ready for recovery, which would be in the shaking incubator. The tubes were put into the shaking incubator at 37C for about 60 minutes. In the meantime, three plates of each type were obtained and labelled: LB/Amp/IPTG/XGal, LB/Kan/IPTG/XGal, and LB/Amp Kan/IPTG/XGal.. Set “A” plates were for ligation and should be labeled with the letter set “B” were for pUC19 and set “C” is for pKAN. Set A was labelled with “L”, B with “A” and C with “K.” Once the 60 minutes were done, it was time to plate. 100ul of each tube were plated on each plate. One plate was done at a time and the hockey stick was flamed between each spread to avoid contamination. The tubes were then incubated for 24 hours. Materials Agarose gel supplies BamHI/HindIII Ethidium bromide Lambda DNA Loading dye Miniprep DNA/TE pUC19 (0.1 ug/ul) pUC19/pKAN (0.1 ug/ul) restriction buffer/RNase 1x TBE Beakers Pipettors Tubes Water baths Electrophoresis supplies Isolation of plasmid DNA: Two cultures were obtained, both of which consisted of E.coli MM294 containing a plasmid. Two microcentrifuge tubes were labeled with identification. A 1.5mL of cells was spun down in each of the microcentrifuge tubes for two minutes. Once the two minutes were up, the supernatant was poured off into a waste beaker containing bleach and then the process was again with the remainder of the sample. Plasmid preparation: 100uL of cold GTE was added to the sample. The pellets were resuspended using the vortex mixer or a pipette. After, resuspension, add 200uL of SDS/NaOH buffer was added, and rapidly mixed by inversion. Once mixed, the tube was set aside for 5 minutes on ice. The suspension was clear at this point. Next, 150uL of cold KOAc was added and mixed by inversion, which caused the protein and cell membranes to precipitate. After mixing, the tubes were placed on ice for 5 minutes again then spun to remove the precipitate. Next, 400uL of the suspension was transferred to a new tube. The suspension was mixed rapidly with an equal volume of isopropanol, then it was left at room temperature for 5 minutes. After 5 minutes, the tubes were spun down for an additional 5 minutes to pellet the pDNA. Next, the supernatant was taken out. The procedure was then repeated with 200uL of 100% ethanol to clean the DNA of any residual salt. Once DNA was cleaned, the tube was dried with direct air from a heat gun. After all ethanol evaporated and the pellet was completely dry, the pellet was resuspended in 15ul of TE. Restriction Digest: The chart shown below was used to set up the second digest. Each tube had the following samples in it. Tubes M1 M2 pUC19 pKAN PUC19/PKAN mix Buffer BamHI/HindIII RNase H20 M1- 5uL 1uL 1uL 3uL M2- 5uL 1uL 1uL 3uL A- 5uL 1uL 1uL 3uL K- 5uL 1uL 1uL 3uL M1 M2 AK 5uL 1uL 2uL 1uL 3ul Once the tubes were prepared, they were incubated at 37C for 45 minutes. In the meantime a well agarose gel was prepared. Once incubation was completed, the gel was loading as follows: M1-, M2-, A-, K-, lambda ladder, M1 , M2 , AK Results Figure 1: pKAN and pUC19 Digestion Figure 1 shows the first digestion of pKAN and pUC19. Table 1: Transformation Plate Ligated DNA L pUC19 control A pKAN control K LB/amp/IPTG/Xgal Low amounts Low amounts TNTC LB/kan/IPTG/Xgal Low amounts TNTC TNTC LB/amp kan/IPTG/Xgal TNTC TNTC TNTC Table 1 shows the results from the plates and how much grew on each plate. The results weren’t as expected. Figure 2: Miniprep digestion Figure 2 shows the miniprep digestion. Lane 1 has miniprep 1 uncut, Lane 2 has miniprep 2 uncut, Lane 3 has A uncut, Lane 4 has K uncut, Lane 5 has lambda ladder, Lane 6 has miniprep 1 cut, Lane 7 has miniprep 2 cut and lane 8 has AK cut. Transformation Efficiency DNA in cell sample = Concentration of plasmid X volume of plasmid (0.2 ug/ul) x (24 ul) = 4.8 ug DNA on plate = DNA in cell sample X volume of cells spread on plate Total volume of cell sample (4.8 ug) x (100 ul) / Transformation Efficiency = number of colonies DNA on plate Discussion We successfully completed all the digests and ligations in the laboratory but we could not achieve the formation of a recombinant DNA molecule. The first digest worked and cut the enzymes as we hoped it would. Although our ligation was done properly, the transformation didn’t work. The plates had no growth which means it was unsuccessful. This could have happened because of a few reasons; the digest of pKAN and pUC19 didn’t yield enough DNA to finish the ligation, the tubes had to be in the heat shock for longer than 90 seconds, or maybe the ligation had to be run for longer. Whichever caused the transformation to be unsuccessful inhibited the DNA from ligating, however, there are future precautions that can be taken to make sure this can be avoided. Properly and accurately digesting DNA is important. It is easy to make a mistake when making the samples and following the little steps so paying close attention is a good start. Also, it wouldn’t be harmful to extend the time for the heat shock and the ligation time by just a little. Even though our results didn’t turn out as expected, we can still analyze what was supposed to have happened if the experiment was successful. Some results include; the A LB/amp plate should have had more growth than the L LB/amp plate due to the mass of the ligated DNA. The A LB/kan plate shouldn’t have had any growth because of the resistance gene present in it. The L LB/kan plate should have had growth compared to the A LB/kan plate because of the presence of kanamycin. There should be growth on both, the A LB/amp and K LB/amp plates, but the A LB/amp should have a little bit more because of the ampicillin. The L LB/amp should have more growth than the L LB/kan because of the kanamycin. The L LB/amp should have more growth when compared to the L LB/amp kan. And lastly, the due to the antibiotic present in both of these, L LB/kan and L LB/amp kan had the same amount of growth. During our second digest we completed the experiment according to what was given. When we ran the gel after the digest, the uncuts showed but not the cuts. This could have happened for a few different reasons. It is possible that the digest wasn’t run properly, something that needed to be in the tubes could have been missed. Also, it’s possible not enough sample was loaded into the gel was caused it to not show. It is also possible that not enough loading dye was added to the samples before loading the gel. It is always important to carefully read each step before preforming the exercise as it is easy to miss the little steps, however, future precautions can be taken to avoid this from happening again. Simply by reading everything thoroughly, a lot of small and simple errors can be eliminated. Making sure you are adding the right amounts of all the samples is also very important. You can also have the digest run for a little longer to make sure everything does cut. Literature cited Freeman WH. Gel Electrophoresis. Nature News. 2014. https://www.nature.com/scitable/definition/gel-electrophoresis-286 “Genomic Determination of the CR1 (CD35) Density Polymorphism on Erythrocytes Using Polymerase Chain Reaction Amplification and HindIII Restriction Enzyme Digestion.”Journal of Immunological Methods, Elsevier, 12 Nov. 2002, www.sciencedirect.com/science/article/pii/0022175991900062 Griffiths AJF. Recombinant DNA technology. 2018 Nov . https://www.britannica.com/science/recombinant-DNA-technology Micklos, D. A, Freyer, G. A, et al. DNA Science a First Course. 2nd ed., Cold Spring Harbor Laboratory Press, 2003. An Antibiotic Resistance Gene Using Recombinant DNA
George Wallace Speech Discussion.

Here are the instructions: Locate at least three key claims in the speech. What is Wallace arguing? What sort of reaction is he seeking from his audience? (6-7 sentences)What features of demagoguery do you see exemplified in this speech? Find at least two quotations and describe, in detail, how closely they demonstrate Roberts-Miller’s notions of demagoguery. (7-9 sentences)What fallacies do you see in this speech? Locate at least one, cite, and explain. (5-6 sentences)What are some other striking characteristics of this address? For example, observe Wallace’s diction (word choice), use of imagery, metaphor, and analogy. What is particularly revealing about it? (6-7 sentences)the file below is the speech to answer these questions. some extra historical context is: 1957: More than 1,000 paratroopers from the 101st Airborne Division and a federalized Arkansas National Guardprotect nine black students integrating Central High School in Little Rock, Ark.1962: A federal appeals court orders the University of Mississippi to admit James Meredith, an African American student. Upon his arrival, a mob of more than 2,000 white people riots.1963: http://www.infoplease.com/spot/civilrightstimeline1.html (Links to an external site.) Martin Luther King is arrested and jailed during anti-segregation protests in Birmingham, Ala.; he writes his seminal “Letter from Birmingham Jail (Links to an external site.),” arguing that individuals have the moral duty to disobey unjust laws.1963: Two African American students, Vivian Malone and James A. Hood, successfully register at the University of Alabama despite George Wallace’s “stand in the schoolhouse door” — but only after President Kennedy federalizes the Alabama National Guard.
George Wallace Speech Discussion

Strayer University Discrimination in Pricing Discussion.

ContextThis week’s discussion will provide you with an opportunity to apply Froeb’s analytic method.Read the example in the discussion instructions while keeping in mind the following questions:Who made the bad decision?What information did they have? And was it good, bad, or unclear?What was their incentive?InstructionsRead the following and then respond to the discussion prompt.Intel made large loyalty payments to HP in exchange for HP buying most of their chips from Intel instead of rival AMD. AMD sued Intel under the antitrust laws, and Intel settled the case by paying $1.25 billion to AMD.Address the following in your discussion post:What incentive conflict was being controlled by these loyalty payments?What advice did Intel ignore when they adopted this practice (consider how the Robinson-Patman Act applies to their practice) and speculate why Intel ignored the advice.
Strayer University Discrimination in Pricing Discussion

Beta Lactam Antibiotics: Examples and Uses. The beta-lactam antibiotics for their extensive scale of actions are preferred most among antimicrobial factors. The penicillins and cephalosporins are the two categories of this β – lactam antibodies that are extraordinarily less toxic to organisms.(1) At present ,the β-lactam groups of antibiotics are the highest frequently used universal antibiotics .(2) Cellular membrane of most bacteria enclosed by a cell wall but an extra outermost layer seen on some of them. The periplasmic space in gram negative bacteria is the cavity in the middle of the cell membrane and the cell wall. Periplasm instead of a clearly defined periplasmic space is retained by most gram positive bacteria .(3) But peptidoglycan is the greatest significant element of the cell wall that linked as a new cell by way of the metabolic absorption in periplasm is a polymer made of N-acetyl muramic acid alternating with N-acetyl glucosamine.Arises of the bacterial cell that is actually a process of peptidoglycan synthesis where accumulation of 5 amino acids to N-acetyl muramic acid is one of the leading phases. A precursor of peptidoglycan that conducted by a cell wall acceptor crossway the cell membrane in the periplasm and developed by linking N-acetyl glucosamine to the N-acetyl muramic acid . Generous crosslinking occurs for two key enzymes (trans peptidase and D-alanyl carboxypeptidase) and for the capacity to bind penicillins and cephalosporins, they are recognized as the penicillin binding proteins. B4 Development of cell wall by cross linking of a number of films of peptidoglycan grounds numerous layers and a much denser cell wall in gram positive bacteria than gram negative bacteria. Beta-lactam ring attach enzymes to cross-link peptidoglycans, that is a chemical structure which is available in the beta-lactam antibiotics consist of all penicillins and cephalosporins. Synthesis of bacterial cell wall is prevented by the affect of beta-lactam when transpeptidase and D-alanyl carboxypeptidase enzymes are attaching there by means of cross-linking and cause deterioration of bacterial cell wall.b5 As a bactericidal agents the antibiotic-penicillin binding protein complex of beta-lactam antibiotics excites autolysin discharge that have the capability of digest cell wall that left after bursting a cell. Generally, excessive inner osmotic pressure possessed by gram positive bacteria and in a low osmotic pressure enclosed atmosphere , cells those are lack of a usual and rigid cell wall are burst out.b6 There are many different types of methods of that bacteria became reistance to beta-lactam antibiotics. Transformation is one of the most important mechanisms among them and in the mean time of this process transfer of chromosomal genes between bacterium happens. Due to the death of a a resistance gene in a bacterium releasing of naked DNA in surrounding environment happens. a process known as homologous transformation and by this method the resistance gene in the host bacteria transferred from the naked DNA to the chromosome. the segment of the host DNA have been remodelled by resistance genes results altered penicillin binding proteins production by coding for cross-linking enzymes. But still cross linking of the peptidoglycan layers of the cell wall happens due to these altered penicillin binding proteins and reduces affinity for beta-lactam antibiotics and the bacterium became resistance. In penicillin-resistant S. pneumonia, this process caused the acquirement of genes from other naturally arising penicillin-resistant Streptococcus species. Bacteria grow into resistant to beta-lactam antibiotics by one more significant system is by the construction of enzymes capable of deactivating or altering the drug formerly it has a chance to apply its outcome on the bacteria. peni The first human gammaretrovirus that is Xenotropic murine leukemia virus-related virus (XMRV) and responsible for chronic fatigue syndrome and prostate cancer (PC) have been revealed in recent times. Gammaretroviruses family is famous for their capability to activate cancer in the infested hosts. Analyzing study showed that vaccine-induced XMRV Env -specific binding and neutralizing antibodies (NAb) titers had limited span but highly changeable. in antibody levels, the different incidence stated for XMRV in a number of prostate cancer and chronic fatigue syndrome cohorts can be explained by the reasonably fast diminution . (2) Monoclonal antibodies (mAbs) have exceptional therapeutic applications in ophthalmology and can be used therapeutically by binding to molecular objects with high specificity. Tumour necrosis factor (TNF), epithelial growth factor receptor, vascular endothelial growth factor (VEGF) receptor, basic fibroblast growth factor receptor, platelet-derived growth factor, and cluster of differentiation antigens repressed by a number of single-agent therapies. Existing and future mAbs in contrast to different cytokines were evaluated for ocular disease treatment and two anti-VEGF mAbs( bevacizumab and ranibizumab), and three anti-TNF agents (infliximab, etanercept, and adalimumab), instruct ocular neovascularization and intraocular inflammation. Other mAbs showed positive results for ocular lymphoma or ocular inflammation but Ranibizumab is the only FDA-approved therapy. Intravenous application of mAbs has established satisfactory toxicity profiles, while intraocular injection may decrease the chances of systemic complications . To develop the excellence and extent of responses is the challenge for the future by merging biologic therapies while lessening side effects. 2i Leading causes of death in the world for coronary syndromes, stroke and other ischaemic arterial diseases . Therapy involves with medical actions correlating thrombolysis, antiplatelet drugs, and the re-opening of the coronary artery by angioplasty. In ischaemic cardiovascular diseases, platelet initiation is a acute phase . Chimeric Fab, c7E3 or abciximab is the only one recombinant antithrombotic antibody currently used in therapy and obstructs the ultimate phase of platelet aggregation. Subendothelium matrix activation by other platelet receptors have been recognized as prospective targets for the improvement of antithrombotic antibodies .2ii In drug development, insulin-like growth factor receptor I (IGF-IR) is becoming an attractive target. IGF-IR owed confined homology to insulin receptor and its specificity permits to distinguish between the two receptors. Recently there are some ongoing on IGF-IR and ongoing clinical trials on anti-IGF-IR monoclonal antibodies and combined treatments. 2iii Beta Lactam Antibiotics: Examples and Uses

English Language and Literature homework help

English Language and Literature homework help. This is a paper that is focusing on a foreign policy brief to understand America’s reaction to other countries. The paper also provides additional information to use in writing this assignment paper clearly.,A foreign policy brief to understand America’s reaction,– Foreign Policy Brief 2.5-3 pages,To understand the way America reacts to another country, we must first understand the other country. Many of the policies seem odd or upsetting to some, while those same policies are praised by others. In this assignment, you will select one of the listed countries, prepare a brief on that country, decide if you think America should have trade with that country, and then research the current American position to see if it aligns with your recommendation.,Lesson Outcomes • Analyze and evaluate significant political actions of another country (CLO 2) • Communicate a political policy position in written form (CLO 4),Steps to Complete the Task,Step 1: Choose a Country. There is a wide array of countries you may choose from, depending on your interests.,The following selections were narrowed to countries normally not viewed significantly in the American media. This project is to help you understand a new culture, as well as, understand policy., The list is separated into parts of the world. Determine what part of the world interests you and then select one of the countries listed., Europe: Spain, Portugal, Italy, France, Germany, Greece, Finland, Sweden, Eastern Europe: Kazakstan, Ukraine, Bulgaria, Romania, Poland, Hungary, Turkey, Middle East/North Africa: Oman, UAE, Yemen, Pakistan, Morocco, Algeria, Kuwait, Sub Saharan Africa: Sudan, Angola, Kenya, Nigeria, Cameroon, Chad, Central African Republic, Asia: South Korea, Indonesia, Thailand, Malaysia, Brunei, Cambodia,Step 2: Secondly, research the Country You need to become very familiar with your countries background, geography, economics, population, and industries., Here are some interesting and informative sites to help you with this:, World Fact Book ,https://www.cia.gov/library/publications/the-world-factbook/, Infoplease ,https://www.infoplease.com/world/countries, US 2019 Trade Policy  ,https://ustr.gov/sites/default/files/2019_Trade_Policy_Agenda_and_2018_Annual_Report.pdf, Writing a Policy Brief ,https://www.idrc.ca/sites/default/files/idrcpolicybrieftoolkit.pdf,Step 3,Step 3: Thirdly, write the Brief A brief is simply a summary of important facts about a country. Your brief should be 2.5-3 pages in length. Be sure to review the Writing a Policy Brief information., A policy brief includes the following information:, 1. Title: Name of the Country you are analyzing., 2. Executive Summary: This section is often one to two paragraphs long; It will include an overview of the people, government structure (i.e. Democracy or authoritarian), historical view of the country (are they stable or prone to political violence), 3. Context or Scope of Problem: This section will be one to two paragraphs outlining what the country has that America might want to trade or find desirable.,4. Policy Recommendations: Based on your view of the countries structure, do you recommend trade with this country? What will be the advantages? Do you see any disadvantages with trading with them (i.e. making other countries mad at us or replacing something we are getting from another country), 5. US Current Policy: Now do a quick survey in the supplied web pages or on your own and state the current US trade policy towards your country. Is it the same are you recommend?, 6. Sources: These should be reliable sources that you have used throughout your brief to guide your policy discussion and also recommendations. You may use APA or MLA style (APA is preferred).,A foreign policy brief to understand America’s reaction,Criteria for Success To be successful, your paper should:, 1. Firstly, be accurate, 2. Secondly, include an introduction and background of your chosen country, 3. Thirdly, address and explain the advantages or disadvantages to US trade, 4. Fourthly, give a concise and pointed recommendation, 5. Be written in your “own voice” with minimal errors in grammar, punctuation, and spelling, 6. Lastly, use academic English (no slang or texting language). (Hint: read your responses out loud before submitting.) 7. Be properly formatted and 2.5-3 pages in length.,Attachments,Click Here To Download,English Language and Literature homework help

Mount Saint Vincent University Tourism in Shanghai Discussion

write my term paper Mount Saint Vincent University Tourism in Shanghai Discussion.

The paper should be approximately 800 words I would like to see you do some significant research on your chosen destination and you should have a minimum of 3 references in your list of resources. You must use the APA system to provide citations in the body of your paper. Failure to properly use APA will result in a significant loss of marks. Destination paper should be visually appealing and presented in report format. it include: the geographies of identity and the destination.The nature of inbound and domestic tourists in the destinationFuture issues with the potential to impact tourism at this destination and the solutionThe city is china shanghai
Mount Saint Vincent University Tourism in Shanghai Discussion

Pizza Huts Service Quality Marketing Essay

Main objectives to conduct this research were to find out the current Pizza Hut’s service quality and its claim regarding the exceeding customers’ expectations. This research also got the point of views of the management that what sort of problems they face while delivering the service quality and how they deal all these challenges to improve their service quality in competitive market. The results collected using the questionnaires assist in finding the current service quality of Pizza Hut and on basis of those collected results management answered the questions regarding the problems they do face while delivering the service quality and the Pizza Hut’s claim about exceeding customers’ expectations. This research also focuses that whether the Pizza Hut is successful in gaining a retaining their customers and what is level of their satisfaction regarding the present service quality. In literature various results can be found regarding the service quality which emphasis a lot regarding the importance of service quality in any firm. Using literature review, analysis of exceeding customers’ expectation of Pizza Hut, is more clear and significant on its current service quality as well. Whereas it is clear that every organization tries its level best to deliver the best service quality to its customers but is very difficult to maintain the standard in delivering the competitive service quality. According to Management point of views, it is crystal clear that there still need to put more attention for company to deliver the good service quality so the customers’ satisfaction level could be increased. There always been a solid association between service quality and customer satisfaction, while a word of mouth has great importance in building the customers’ expectation as well. According to Parasuraman et al (1988), an ability to meet customers’ expectation is known as service quality. Its mean when company meets with the expectations of customers they automatically get satisfied and come again. Gap Model is very supporting in finding the major dilemmas and gaps in current service quality of company. First gap deals with perception of management regarding customers’ expectations, and what they receive. Mostly it happens that companies are failed in delivering the promising service quality to its customers, hence customers can not be satisfied. In second gap, company still did not take any particular measures so the customers could be satisfied, while third gap deals with difference in specified service quality and current service quality. Here in gap four, service quality and overall performance is not fulfilling the promising service, while gap five discuses the difference in customer expectation and what they receive in real. Mix mode research conducted for this research, questionnaires from customers and on the basis of those results management was interviewed. This research gives many findings regarding the customers’ expectations and service quality currently being delivered by the Pizza Hut. 5.1 Analysis of Pizza Hut customer satisfaction Data collected by the customers using the questionnaires gave results in negative and over all score regarding the service quality was -2.87. This service quality overall score of Pizza Hut explains that still Pizza Hut which is know world’s biggest fast food restaurant is not meeting the service quality requirement and customers are still not satisfied by the service. This result is totally in opposite direction of company’s claim that Pizza Hut exceeding customers’ expectations, whereas company even does not meet customers’ expectations and customer are not satisfied as well. 5.1.1 Analysis of SERVQUAL dimensions During the study, all the dimensions got negative score especially in reliability the performance was found very poor. In the table 4.3 customers showed the main gap and regarding time preferences results were found really poor. Regarding the time, orders were not being delivered well in time and customers sometimes wait a lot in getting their orders. In tangibility results are bit better rather than all other dimensions. Sitting and eating environment in some level are nearly match the standards because customers expressed satisfactory views regarding the food and hygiene standards but still gap is present. Complaints are not handled according to the customers’ will. It takes mostly long time in delivery, while takeaway and Dine-ins are considered better rather than delivery. Company is still fail in delivering the orders well in time. While looking in to the responsiveness, in the results it is on the top place so far in the research. Receptionists’ responses towards the customers need professionalism and still demands to make it perfect and up to the level. Pizza Hut is still unsuccessful in assuring its customers that they get the best services. Staff behavior with customers is not friendly and customers are not satisfied while performing any transaction. In empathy the staff is fail to give the attention to customers and their needs. Every customer needs much care and attention of the staff member. If the staff does not show deep concerns the customer would never visit again. 5.2 in the bottom Major gaps in Pizza Hut’s customer service 5.3 Factors involve in performance of service quality In this research using the questionnaires and interviews with the management, many gaps have identified. There are many factors for these gaps which are lying in details as under. 5.3.1 Communication gap between departments Communication and understanding between all the departments is very important for a successful business. There is much lack of communication in between operations, marketing and management departments. Customers’ expectations are obviously very high and when the company does not meet with expectations then customer can not be satisfied. For instance, marketing department gives offers to its customers while management provide less hours and due to shortage of staff it is impossible to deliver satisfactory service quality. Non-promising service Customers do have very high expectation and it is tough to meet the expectations. Pizza Hut always claims for exceeding customers’ expectations and good service quality, but still fails in doing so. Like, Pizza Hut promises for delivering the orders well-in-time, but mostly fails. Customers feel not good when they expect more and get less when they visit the store which also damages the good-will of the company and call it poor service quality. 5.3.3 Improper implementations of policies Pizza Hut has many set standards and policies which all the outlets are directed to implement. Unfortunately due to lack of professionalism there are still deficiencies in implementations of these policies. Like CHAMPS stands for, Cleaning, Hospitality, Accuracy, Maintenance, Product Quality and Speed which are not accurately followed by the stores. Many standards like food standards, communications standards, well in time delivery and environment standards are part of theory. Practically things are not done properly in stores, for this reason company fails in meeting the exceeding customer expectation. 5.3.4 Impact on service quality of employees’ dissatisfaction According to Kristen A. Riscinto-Kozub (2008), Chang-Hsing Chang (2007), Bitner et al. (1990), in their research there is strong relationship between employees and customer satisfaction. Management and other employees were major part of this research and low-level staff and management was interviewed to find out the fact and figures. This research found that the Pizza Hut staff was not much happy and satisfied with the management. Some employees said that their hourly pay rate is less than other competitor companies. They said there is no any reward system on performance basis and very less chances for promotion are available. Some of them respond that mostly students work round about 20 hours in Pizza Hut and they do not put more attentions towards the tasks which affect the Pizza Hut’s overall service quality as well. In response to a question one employee said, Pizza Hut should spend on employees more so they could perform better to prove best service quality for the customers’ expectations. Note below: Checks: confirm the table 4.3 5.2 Major gaps in Pizza Hut’s customer service This study has evolved and found out that there are three gaps present in Pizza Hut-UK’s customer service quality. From the perspective of Gaps model, gap 3, 4 and 5 are present in the company. ‘Gap 3’ is about consistently meeting of standards. Basically it is the difference between service quality specifications and actual service delivery to the customers. Company has designed very good service quality specifications and standards which are aims to meet and exceed customer expectation, but company fails to deliver such level of service quality. From this research there are various reasons which came forth and are going to be discussed later in this chapter. The second major gap is ‘Gap 4′, which is in the service delivery being communicated externally. When managers were interviewed they responded that there is lack of communication in between various departments, like marketing department and operation department and policy makers’ level. This gap is about consistently fulfilment of promises made by external communications. Marketing department is the one which communicates externally. As it is known that expectations of customers build up on the basis of past experiences and communications, so marketing department makes customers expect more but company fails to deliver that. Third major gap is ‘Gap 5′, which is alarming for Pizza Hut-UK. It means a difference between what customers expect of a service and what they actually receive from the company. In other works it is a gap between expected service quality and perceived service quality. In questionnaires, customers’ response depicted a different picture of the company with major negative gaps in all service quality dimensions. Research concludes that customer never gets what he expects from Pizza Hut, which is really crucial for the company and is totally against company’s claims.

Importance of International Business Management for Running a Successful Company Research Paper

Importance of International Business Management for Running a Successful Company Research Paper. Introduction Well established modern business organizations have increasingly been seeking to expand their operations beyond the boundaries of their mother countries. Consequently, international trade has been stirred up and accelerated thanks to advancement made by man in Information Communication Technology (ICT) and other areas during 19th and 20th Centuries. Business organizations particularly from world’s major economies and the emerging Asian economies offering goods and services in different commercial sectors or industries are striving to establish a high profile presence in the international markets in an attempt to enhance their profitability and remain competitive in an era of accelerated globalization. Some of them are part and parcel of the business organizations known as multinational corporations (MNCs) or multinational enterprises (MNEs) which have operational branches in virtually all stable nations world over. There are various reasons that encourage business organizations desire to expand in to the international or foreign markets. Ireland, Hoskisson and Hitt (2008) have identified four main reasons that usually motivates companies, or business organizations seek to expand in to international markets namely the desire to use current resources and gain access to new resources, seeking to expand or develop new markets, competitive rivalry and controlling core competences and learning. These authors point out that companies search for economies of scale in the use of their existing resources by expanding in to new foreign markets. At times they enter international markets in order to reach specific valuable resources like raw materials, specialized knowledge, or cheap labour. As companies become well established in the domestic markets, they begin searching for international markets in order to increase their revenue and gain more profits, enhance their competition with major rivals and influence further development of their core competencies (Ireland, Hoskisson and Hitt, 2008; Ajami and Goddard,2006). Background Increased and accelerated international business activities have brought up the need for special managerial knowledge and skills on how to manage profitably and successfully companies that seeks to expand their operations in to new global markets. The need for this knowledge has been partly due to the fact that irrespective of the many benefits that a business organization accrues because of expanding into new foreign markets, they encounter various problems and challenges in their preferred international markets DewanImportance of International Business Management for Running a Successful Company Research Paper