《基因工程》习题及参考答案

《基因工程》习题及参考答案一、习题：1. What are biotechnology and genetic engineering?2. What is a gene?3. What are genetically engineered medicines?4. What do genome research and human genetics deal with?5. What potentials are held out by genetic diagnosis?6. What options are given by gene therapy?7. What is an embryo - and what is a fetus?8 What is a genetic fingerprint?9 What does the term "therapeutic cloning" mean?10 What are stem cells?11 What is a transgenic organism?12 What does xenotransplantation mean?13 How will genetic engineering be used in agriculture?14 How are genetically modified organisms assessed?15 What does the German Embryo Protection Act regulate?16. What is a genome?17. Is there a risk of bioterrorism?18. How does genetic engineering affect the environment?19. Are genetically engineered crops good for farmers?20.What is the difference between restriction digestion and restriction mapping?21.Can you combine two different restriction enzymes in the same reaction tubes todigest the DNA molecules?22.Why should we need to generate restriction mapping data?23.How many restriction enzymes available now on the market?24.Why do you consider mutagenesis in vitro as one of the most critical techniquesfor us to understand in genetic engineering class?25.How do we choose the methods for DNA modification?26.How do we choose a gene expression system?27.How can we express eukaryotic gene in E.coli?28.What should we consider before we start the recombinant protein expressionexperiment?29.What is the advantage of yeast expression system?30.What is the advantage of insect expression system?31.Why there are so many different types of vectors available for cloning?32.What is the difference between cloning vector and expression vector?33.What is a genetic fingerprint?34. 基因具体分成多少种类？35. 什么叫印记基因？36．什么叫遗传漂变？37．人类基因组图谱和初步分析结果是在哪一年公布的？38．人类基因组共有多少基因?39. 克隆羊成功的技术关键是什么？40. 有人计划将两个不同物种的动植物体细胞进行融合，然后将融合体的核移植到其中一种生物的未受精卵细胞中，进行体细胞克隆。

此计划可行吗？为什么？41.切口移位标记探针的主要步骤有哪些？42.用EcoRⅠ和Hind Ⅲ分别切割同一来源的染色体DNA，并进行克隆，在前者的克隆中筛选到A基因，但在后者的克隆中未筛选到A基因，是什么原因？43.在基因工程中，为了在细菌细胞中表达真核细胞的基因产物，为什么通常要用cDNA而不用基因组DNA？为什么要在cDNA前加上细菌的启动子？44.当两种限制性内切酶的作用条件不同时，若要进行双酶切，应采取什么措施？为什么？45.有些噬菌体和质粒常常编码一些抗限制性酶的蛋白以中和宿主的限制系统。

除此之外噬菌体和质粒还有哪些可能的方式避免宿主的限制系统？46.为什么大多数内切酶被称为“限制”酶？47.何谓同裂酶？48.什么是限制性物理图谱？49.为什么反转录酶在聚合反应中会出错？50.RNase A和.RNase H在催化活性和应用上有何不同？51.切口移位标记DNA前，用DNaseⅠ处理DNA时，应注意什么？52.DNA连接酶对DNA的复制是很重要的，但RNA的合成一般却不需要连接酶，为什么？53.RNA聚合酶Ⅲ特异性地转录小分子RNA，但为什么不转录5.8SrRNA?54.用EcoRⅠ切割外源DNA片段时出现了星号活性，可能是什么原因？二、参考答案：1 What are biotechnology and genetic engineering?Biotechnology is a field of bioscience. In very general terms it can be described as the theory of the use of biological systems in research and applications. Biotechnology applications are a rapidly growing sector of the economy. The number of biotech companies in Germany has increased considerably over the past seven years. Genetic engineering includes all biotechnological processes aimed at effecting specific changes in the genetic material of cells. The principle is, in general, the following: sections of DNA (= deoxyribonucleic acid) from outside sources are introduced into a cell in order to bring about defined changes. What is involved in most cases is the synthesis of molecules, the information for which is contained in the introduced DNA. This process has been used to produce new medicines (see genetically engineered medicines).2 What is a gene?A gene is the smallest unit of genetic material. It describes a specific section of DNA which, as a result of its nucleotide (= DNA component) sequence, determines specific functions, characteristics, features, or structures of a cell. The way genes are expressed in each case is dependent on their interaction with other genes and with the environment.The genome is the entirety of genes contained in an organism. The term is synonymous with genetic material.3 What are genetically engineered medicines?In the genetic engineering of medicines genes containing the codes of therapeutically valuable substances are transferred to easily cultivated cells. In most cases bacteria are used, more rarely also yeast cells and the cells of mammals. Medicines produced in this way are called recombinant medicines.The human insulin introduced (for diabetics) in 1982 was the first recombinant medicine. Since the mid-1990s persons with hemophilia A (the most common form of this blood disease) have been treated to an increasing extent with recombinant factor VIII. Prior to that the only treatment available was factor VIII produced from human plasma.4What do genome research and human genetics deal with?Genome research pursues the objective of identifying the structures and functions of the individual genes in the genome. The object of genome research is the study of the genetic material of all organisms, meaning that the genomes of certain plants, animals, and microorganisms are just as much a subject of study as the human genome.It is an important task of human genome research to identify what genes are involved and in what ways they are involved in causing diseases. This research is expected to bring new approaches in the treatment of cardiovascular diseases, cancer, infectious diseases, and diseases of the central nervous system such as Parkinson's disease, multiple sclerosis, or Alzheimer's disease.Scientists presented the largely decoded human genome on June 26, 2000. Both scientists and government leaders throughout the world noted this as an epoch-making event and an important milestone in the history of mankind. At the same time this has created new challenges for science, government, and ethics. The field of ethics is called upon to show whether and how this knowledge can be used responsibly in such widely diverse fields as medicine and agriculture.Human genetics is a medical discipline that deals with human heredity, the causes and diagnosis of hereditary diseases, and the search for possible therapies. Diagnostic procedures today include state-of-the-art genetic diagnoses in addition to traditional analyses of hereditary mechanisms.5 What potentials are held out by genetic diagnosis?Modern genetic diagnosis methods make it possible to analyze individual structures of the genome and, in doing so, to identify gene defects associated with hereditary diseases such as cystic fibrosis. This would be beneficial to persons who carry the gene and would run the risk of passing it on to their children. It is conceivable that awareness of an inherited predisposition for a disease that emerges later in life would cause the persons in question to take special precautions such as going in for preventive checkups or pursuing a lifestyle aimed at preventing an outbreak of the disease.Gene tests can be used to confirm the diagnosis of suspected diseases. More than that, modern methods (such as gene chips) can be used to help a wide range of people, including those who are not sick, to identify genetic characteristics such as predispositions for certain diseases. Here, entirely new ethical and legal questions are arising for society. In the case of predictive gene tests, i.e. tests used to make forecasts as to risks of hereditary illness, it will be a separate task to determine whether these tests will be of value to individuals and to society as a whole.We need to distinguish between preimplantation genetic diagnosis (PGD), prenata l genetic diagnosis in the context of prenatal care, and postnatal genetic diagnosis. Preimplantation genetic diagnosis (PGD) is a controversial area of genetic diagnosis. After in vitro fertilization PGD can be used to carry out a gene test on an embryo before it is implanted in the mother's uterus. By this means embryos showing a specific genetic defect can be sorted out.6What options are given by gene therapy?The objective of gene therapy is to treat or prevent diseases, making use of genetic engineering principles. DNA can, for instance, be introduced into blood or liver cells by means of vectors in order to eliminate genetic defects present there. Or cells that have been genetically modified outside of the body may be implanted in the patient. A further approach is the injection of so-called naked DNA as a kind of vaccine. Naked DNA makes it possible to synthesize proteins against which the vaccinated person builds up an immune reaction as in the case of a conventional vaccination, except that here a much more specifically targeted effect on the immune system is expected. Somatic gene therapy must be distinguished from germ line gene therapy. After germ line gene therapy the effected change in genetic information would be passed on to the offspring of the individual in question. The prerequisite for this would be manipulation of the genetic material contained in an egg cell or a sperm cell or of the genetic material contained in an embryo in the early stages of development. The German Embryo Protection Act imposes a comprehensive ban on carrying out germ line gene therapy in humans.In the case of somatic gene therapy (= therapy of or involving body cells) only body cells (= somatic cells) are the object of change. Since this type of treatment is directed exclusively at somatic cells, it will not have an effect on the genetic material passed on to the patient's offspring. The predisposition for a given disease will be passed on to the patient's children even if gene therapy is successful. It should be noted that at the present time gene therapy is used mostly to treat malignant cancers, and in Germany exclusively in such cases. The cause of the disease is usually not associated with one single gene but rather with a combination of external factors involving several genes. For part of the diseases treated there is no evidence of a genetic predisposition and in the underlying therapeutic approach the question of a genetic predisposition is without significance.7 What is an embryo - and what is a fetus?In human medicine an embryo is usually defined as a fertilized egg cell in the early stages of development up until the point in time when the organs have been fully formed (eighth week of pregnancy).After that (from the ninth to the thirty-eighth weeks of pregnancy) we speak of a fetus. Sometimes these terms are defined differently from this under the law.8 What is a genetic fingerprint?Genetic fingerprinting (also known as DNA fingerprinting) is a procedure provided by molecular biology with which the unique genetic patterns of any individual can be determined. Except in the case of identical twins no two genetic fingerprints are the same.In paternity suits or in criminal investigations comparisons of DNA fingerprints make it easier or, indeed, may be the only way to identify the guilty party.9 What does the term "therapeutic cloning" mean?Cloning is generally understood to be the artificial production of a genetically identical copy of a living organism. The concept refers to complex organisms as well as to individual cells. All the bacteria in a specific colony are genetically identical. In humans this situation is given only in the case of identical twins.Reproductive cloning is the process of creating genetically identical organisms. For this purpose embryos created artificially, i.e. by means of a nucleus transfer, are implanted in the uterus of the prospective mother where they are to develop into "normal" organisms. The most widely known case of reproductive cloning is Dolly, the sheep that was presented to the public by British scientists in 1997.The process of cloning consists in removing the (haploid) nucleus from an egg cell and replacing it with the (diploid) nucleus of any given cell of any given origin. This creates the situation in which an embryo and, in the end, a fully developed organism can grow from the egg cell, as in the case of a fertilized egg.In the case of therapeutic cloning the diploid nucleus comes from the patient in whom destroyed tissue is to be replaced, e.g. following a heart attack or a spinal injury. Incontrast to reproductive cloning, the embryo that has been created artificially by means of a nucleus transfer is not implanted in the uterus of a prospective mother. Instead, in the so-called blastocyst stage of development (about the sixth day) the inner cells are removed from the embryo, destroying it in the process. The stem cells taken from the embryo are used as a raw material for the development of the cell/tissue types needed. The cells or tissues grown from them will not be rejected later when they are transferred to the patient, since they are genetically identical with those already in the patient's body. In contrast to the transplantation of cells and tissues customary today, no medication is needed to suppress defensive reactions of the immune system and the long-term survival of implants becomes feasible. Therapeutic and reproductive cloning in humans is prohibited in Germany under the provisions of the Embryo Protection Act. In the United Kingdom of Great Britain and Northern Ireland therapeutic cloning in humans is now possible under strict regulations, but reproductive cloning continues to be prohibited. It is assumed that many years of research are needed yet before cells or tissues can be transferred successfully in humans on the basis of therapeutic cloning. There has not been enough research carried out thus far on the extent to which the growth of transplanted cells and tissues can be controlled and whether tumors or malignant tissue growth could occur. The cloning of complex organs such as hearts or kidneys is currently beyond reach.10 What are stem cells?Stem cells are the predecessors of more differentiated and, as such, more specialized cells. Bonn marrow, for instance, contains stem cells for the various cells that occur in blood such as lymphocytes, granulocytes, or thrombocytes. Stem cells show different degrees of differentiation. They may have the ability to develop into a complete organism, into a variety of different cells, or into specific cells only. These stages are reflections of embryonic development.In the first days after fertilization embryonic stem cells have what is known as "totipotence", i.e. the ability to develop into a complete organism. In the furthercourse of development they have varying degrees of "pluripotence", i.e. the ability to develop into specific tissues with specific functions such as liver, kidney, or brain tissue. Small numbers of stem cells are found in these types of tissue in adults or in the blood of the umbilical cords of newborn babies. The latter could possibly serve as an alternative for embryonic stem cells. The extent to which this is the case is currently a subject of research.Members of the medical community attach considerable hope to stem cell research. They see potential for being able to replace damaged tissues such as myocardial tissue after a heart attack or nerve tissue in the brain in connection with the treatment of Parkinson's disease or in the spinal cord after back injuries. Adult cells cannot be used for this, since they, in contrast to stem cells, are no longer able to integrate themselves into the tissue of an organ and take over the necessary functions.It is of fundamental importance for implementation in clinical practice to be able to grow unlimited numbers of stem cells at specific levels of differentiation potential and, at the same time, to be able exert precise control over their growth into certain types of cells as needed. Similar to the situation given in connection with therapeutic cloning, questions as to the ability to control tissue growth and potential for the formation of tumors and malignant tissue are still unanswered. There is a need for extensive research here.11 What is a transgenic organism?A transgenic organism is an organism, into the genetic material of which exogenous genes have been introduced with the help of genetic engineering methods. This process is also called "genetically engineered change", "gene manipulation", or "germ line modification" in reference to animals. Transgenic organisms pass on the new genetic information to their offspring. Manipulation of human germ lines is prohibited in Germany.Transgenic animals are being used to a growing extent in medical research, e.g. transgenic mice for research on the Creutzfeldt-Jakob disease (CJD).Transgenic plants are being developed mostly for use in agriculture. Transgenic microorganisms have been used primarily in the production of medicines and enzymes. Other uses of genetically modified microorganisms include food processing and the production of biodegradable plastics. In some cases transgenic animals, plants, and microorganisms have been patented.12 What does xenotransplantation mean?Xenotransplantation means the transplantation of animal organs, tissues, or cells into humans as well as exposure to animal organs or cells through the bloodstream. Although primates were originally considered to be suitable donors for xenotransplantation, pigs are now considered to be a better choice for a number of reasons.The transplantation of entire organs from pigs is prevented by uncontrollable hyperacute reactions of the human immune system. The blood vessels of the transplanted organ clog up within a few minutes and the organ dies. An attempt is being made to create donor animals genetically engineered to reduce or entirely eliminate the rejection of transplants by the immune system. At present it is not yet possible to say whether xenotransplantation will become a clinical reality.There are fundamental reservations with regard to xenotransplantation. They derive from the fear that new types of infectious pathogens could develop from animal cells or organs. Infections of this kind might not be limited to the individuals concerned and spread throughout the population. The probable need to suppress immune reactions to the transplant that would be stronger than in the case of an allotransplantation (i.e. human to human) would tend to favor the occurrence of infections.13How will genetic engineering be used in agriculture?When genetically modified organisms are developed for use in agriculture we speak of "green biotech". Their use usually means a large-scale presence in the environment and needs to be approached carefully for this reason. Consumer acceptance of greenbiotech has been low thus far and the use of genetic engineering in animal husbandry raises ethical questions.Genetic engineering is being used in plant breeding to create disease and pest-resistant plants and, in this way, reduce the use of pesticides. There is a need, in particular, to monitor the long-term effects of genetically altered organisms on the environment, since this cannot be fully determined in advance on the basis of experiments subjected to limitations with regard to time and geography. There is also a need for long-term monitoring after admission of genetically modified organisms to the market. It is only in this way that it will be possible to identify possible undesirable long-term effects of their use on a large scale and to obtain reliable data for evaluation.Objectives being pursued with the help of genetic engineering in the field of agriculture:Studies are being carried out on the use of genetic engineering* to produce pest and disease resistance, i.e. fungus resistance, bacteria resistance, virus resistance and insect resistance in plants* for biological pest control making use of genetically engineered viruses* to produce herbicide resistance in plants (worldwide the most common genetic change being made in crop plants)* to enhance the quality of plant products, e.g. improve the shelf life, storage life, digestibility, nutritional value, and taste of foods* to produce feeds* to produce regenerative raw materials. Genetic engineering has the potential to broaden the range of regenerative resources produced for the chemicals, cosmetics, and pharmaceuticals industries, to broaden the range of raw materials available for energy consumption, or to make applications of this kind possible, and, as such, to supplement the petroleum resource base.* to produce developments in the breeding and raising of livestock aimed at reducing costs by means of higher fattening performance, lower material consumption, improved profitability, the development of new products, or the development of disease resistance.A process of rethinking the foundations and conditions of food production has begun, triggered by the BSE crisis. The German government takes the view that it is only through a systematic, consumer-oriented reorientation of agricultural policy that it will be possible to restore the confidence of the public in food production.The objective pursued in connection with the reorientation of agricultural policy will be to give a convincing response to the increased requirements of society with regard to health and environmental protection with a consumer-oriented agricultural policy and, in doing so, to include the conscionable potentials of green biotech.14 How are genetically modified organisms assessed?Approval regulations and assessment standards exist for genetically modified organisms under the Genetic Engineering Act. Under the provisions of the Genetic Engineering Act characteristics of genetically modified organisms must be studied and their interactions with the environment must be predicted and evaluated before they can be intentionally released into the environment. On this basis it is decided whether approval can be given for test releases or for their marketing (commercial trade). When genetically modified organisms are handled in experimental facilities (e.g. labs, fermenters) for research or production purposes, they, along with the research work or production processes being carried out, are assessed with regard to their safety, and the experimental facility in question is subject to licensing approval. The EU member countries and the European Parliament adopted a fundamental amendment to Council Directive 90/220/EEC on the deliberate release into the environment of genetically modified organisms. The new directive entered into force at the end of February 2001. Under the new directive it will be possible to combine the approval of genetically modified organisms for market consumption with binding regulations on monitoring the GMO in question. In addition, general monitoring of genetically modified organisms will be introduced. Both the monitoring of specific GMOs released into the environment as well as general monitoring are aimed at determining and assessing environmental changes and possible effects on human health. In particular, knowledge gaps on environmental effects are to be closed.15 What does the German Embryo Protection Act regulate?The Embryo Protection Act entered into force on January 1, 1991.It contains the following elements enforceable under criminal law:* a ban on the illegal application of procedures used in reproductive medicine,* the protection of human embryos against illegal use,* a ban on artificial fertilization of egg cells with the sperm of persons who are dead, * a ban on gender selection,* protection against arbitrary fertilization and transfer of embryos,* a ban on gene transfers in germ cells,* a ban on cloning, and* a ban on the creation of chimeras and hybrids.Cartagena Protocol on BiosafetyThe Second Conference of the Parties to the Convention on Biological Diversity held in Jakarta from 6 to 17 November 1995 adopted a decision to formulate a supplementary protocol to the convention which has come to be known as the Cartagena Protocol on Biosafety. After long and difficult negotiations this legally binding protocol was adopted in Montreal in January 2000 and since then has been signed by about 80 countries. The protocol contains provisions on the safe handling and transfer of living modified organisms or LMOs (synonymous with genetically modified organisms or GMOs). Particularly important in this context are the provisions on transboundary movement of LMOs. Exports for experimental release and transfer of LMOs are to take place only if and when the receiving state has been informed in advance and in detail of the risks involved, in particular of risks for biological diversity, and has given its approval. There is also a provision requiring that biotech accidents be reported if transboundary effects are to be reckoned with.In the framework of the Convention on Biological Diversity agreements were concluded on the handling of biotechnology and the distribution of its benefits. On the basis of these agreements a Clearing House Mechanism (CHM) was established which serves as a liaison office for information on biological diversity. It also collectsinformation on the biological safety regulations that exist in the countries who are parties to the Convention on Biological Diversity, detailed reports and articles on biosafety-related questions and issues, as well as the latest news items and scheduled meetings.16. What is a genome?The standard definition of a genome is that it is the whole collection of the genes of an organism. However, a genome is much more. It is not only the set of all genes present in the nucleus of the cells of the organism, but also the way these genes are organised next to each other. If you consider a recipe for your favourite dish, you can easily understand that having only the list of ingredients would not be enough to allow you to prepare the dish! A genome is the whole text of the DNA present in every cell of a living organism. It therefore comprises regions corresponding to CDSs, as well as all the regions in between ("intergenic regions").The genome organisation is so important that in fact one can often tell something about the order of the genes in the genome, genes with related functions being often in some sort of proximity.17. Is there a risk of bioterrorism?This text was written at the creation of the Centre, well before the terrorists attacks using anthrax, demonstrating, alas, that our world is not as good as it should be.... Unfortunately, human beings are often enflamed by wrong or even plainly destructive ideas. This gives rise to aggressive behaviour. Biological warfare existed for a long time. The Spanish invadors of America used it, and, more recently, there have been quite a few attempts in China by Japanese militaries. One cannot, therefore, dispell the idea as far-fetched. However, it must be understood that biological warfare, in contrast to all other means used in wars, is difficult, if not impossible, to control once it has been launched. In particular, agents such as Bacillus anthracis, which have been used for that purpose, remain in the environment for decades or even centuries. A test bomb, exploded by the British in Gruinard Island, and full of B. anthracis spores,resulted in contamination of the island for 50 years, and one had to scrape the surface of its soil and spread strong bactericides to get rid of most of the spores! This does not mean of course that this type of warfare will not be used, but it is clearly a very mad way to proceed. Please note here the symptoms of anthrax. The Institut Pasteur in Paris provides a large sum of information on the topic (in French).Can we trust the sanity of human beings? To counteract this possibility the best is certainly to control as much as possible information on the matter, and refrain to spread it. Clearly, freedom does not mean freedom to kill. Misinformation is another possibility, but it is difficult to control.18. How does genetic engineering affect the environment?Unlike previous pollution genetically engineered crops are living organisms, they can reproduce and spread. This poses an ongoing threat to the environment. If something goes wrong this GE pollution cannot simply be recalled or cleaned up.Ecologists believe that the release of these unpredictable organisms could have far reaching consequences in much the same way that non-native species such as the cane toad have become pests. Such effects may only emerge after a few generations. Persistent GE crops may cross with related weeds to become superweeds? Particularly vigorous weeds that out-compete other plants and destroy natural plant ecosystems. GE crops produce seed and pollen which can contaminate other crops and also soil. Soil is extremely complex and there is already some evidence that GE crops have affected the fertility of soil.One type of GE crops designed to produce their own insecticide have been shown to be harmful to the larvae of butterflies, to ladybirds and other important insects. Beekeepers are concerned that such crops may also be harmful to bees who ingest pollen. The real impact of GE crops on the many types of insect and wildlife found in the environment has not yet been checked.So far most of the GE crops that are grown are designed to be used with powerful weedkillers. When the Roundup Ready canola? It is sprayed with roundup weedkiller。