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1、Stem Cells & Regenerative MedicinenTransgenic AnimalsnCloningnStem Cells & Regenerative MedicineTransgenic AnimalsTransgenic AnimalsOutlinenWhy develop transgenic animals?nHow are heterologous genes introduced and expressed in animal systems?nCan animals be cloned?nWhat is the current technology in
2、animal transgenesis?nWhat are the applications of transgenic animal technology?Transgenic AnimalsWhy?nSelective breeding has been used over many generations to genetically improve animals for desirable traitsnBreeding pure lines is time-consuming and costlynIntroduction of a new trait into a pure br
3、eeding line is difficultnLimited by naturally occurring traitsTransgenic Animal ApplicationsnGenetic engineering of desirable traitsnRapid creation of pure breeding lines and identical offspring (clones)nAnimal model systems for research in human diseasesnAnimal systems as bioreactors for production
4、 of heterologous biological molecules (“pharming”)How to Introduce Foreign GenesBasic StrategynInject a cloned gene into the nucleus of a fertilized eggnImplant the genetically engineered egg into a receptive femalenScreen for animals with the gene integrated into their germ line cellsnSelective bre
5、eding to establish new genetic linesMouse TrangenesisMethodologynRetroviral vectors that infect cells of early stage embryos followed by implantation and selective breedingnMicroinjection of DNA into the nucleus of a fertilized egg followed by implantation and selective breedingnIntroduction of gene
6、tically engineered embryonic stem cells into early stage embryos followed by implantation and selective breedingMouse TransgenesisRetroviral Vector MethodMouse TransgenesisRetroviral Vector MethodnEfficient mechanism of integrating the transgene into the recipient host cellnCan transfer only small f
7、ragments of genetic information ( 8 Kb)nRandom insertion of transgene in host genomenPotential for retroviral contamination in transgenic animalMouse TransgenesisMicroinjection MethodTransgenic animals“A little bit of this, and a little bit of that?”Mouse TransgenesisMicroinjection MethodnLow effici
8、ency of transgenesis ( 5%)nInjected transgene integrates at random sites within the host genomenVariable levels of expressionnGenes for selection not requiredEfficiency of MicroinjectionMouse TransgenesisEmbryonic Stem Cell MethodMouse TransgenesisEmbryonic Stem Cell MethodnEfficient transfection in
9、 culturenPositive-negative selection and enrichment of ES cells in culturenTargeted insertion in host cell genomenTargeted gene disruption and development of gene-knockout strainsNegative SelectionPositive SelectionGene KnockoutTransgenic Mice: ApplicationsnModel systems for human diseases and feasi
10、bility studies for production of therapeutic agentsnMouse models for human diseases such as Alzheimer disease, arthritis, muscular dystrophy, tumorigenesis, hypertension, neurodegenerative disorders, and coronary diseaseAnimal Cloning:To Clone, or not to CloneDollyGeorgia Agriculture Education Curri
11、culum OfficeJuly 2003Pros:nCure human diseasesnUsing animal organsnCreate animals that are disease resistantnMore consistent food productsnSave endangered speciesCons:nPublic perceptionnUse technology to clone humansnExpensivenNot efficientnCloned products cant be marketedCloningDefinition: The proc
12、ess of making identical genomic copies of an original animal.Encyclopedia Britannica: An individual organism that was grown from a single body cell of its parent and that is genetically identical to it.Brief History of Cloningn1902: Walter Sutton proves chromosomes hold genetic information.n1902: Ge
13、rman scientist Hans Spemann divides a salamander embryo.nSpemann proposes a “fantastical experiment”Brief History of Cloningn1952: Briggs and King clone tadpoles.n1953: Watson and Crick find the structure of DNA.n1962: John Gurdon clones frogs from differentiated cells.n1963: J.B.S. Haldane coins th
14、e term clone.Brief History of Cloningn1977: Karl Illmensee creates mice with only one parent,n1984: Twinning- create genetic copies from embryonic cells.n1996: First animal cloned from adult cells is born.The Cloning Processn1978: Splitting embryosn1986: Embryo Cloningn1994: Embryonic cell line clon
15、ingn1996: Adult or Somatic cell cloningCreating DollyCloning by Nuclear TransferStage 1Cell collected from a sheeps udder.Stage 2Nucleus is removed from unfertilized egg of second sheep.Stage 3Udder cell is inserted into egg with no nucleus.Stage 5Electrical charge is supplied.Stage 4Insertion is su
16、ccessful.Stage 6Cells begin to divide.Stages 7 & 8Cloning FactsnPlant cloning has been around for thousands of yearsnFarm animal cloning has been around for over 20 yearsnCloning is a form of asexual reproductionnClones arent exact copiesnCloned animals are safe to raise and eatCloning FallaciesnGen
17、etic make-up is alterednMutants are creatednClones are unhealthynWill eventually lead to cloning humansnPossible to recreate people such as HitlerHouse Bill 2505Human Cloning Prohibition Actn Prohibition on human cloningnCriminal Penalty: Up to 10 years imprisonmentnCivil penalty: Minimum 1 million
18、dollar fineFinal ThoughtsnCloning has been around for a long timenCloned products are safenUseful in medical and pharmacological fieldsnWill not replace traditional animal agriculturenNeed to better educate publicnClose regulationStem Cells & Regenerative MedicineScience of Stem Cells?nStem cell res
19、earch has potential to contribute to new therapies and improved quality of life.nLate 1998 a group of scientists, led by James Thomson, at UW Madison were the first to isolate and culture human embryonic stem cells.nRevolution in medicine & basic biology.Why are Stem cells important?nAbility to deve
20、lop into virtually any other cell made by human body.nIn theory, if stem cells are grown they could be of significant medical importance to grow bone marrow, neural tissue or muscle to treat Disease.nParkinsons*, Alzheimers, Diabetes, Heart Disease, Stroke, Down syndrome, Spinal Cord injuries and Bu
21、rns.Two Kinds of Stem Cells Embryonic (also called “pluripotent”) stem cells are capable of developing into all the cell types of the body. Adult stem cells are less versatile and more difficult to identify, isolate, and purify.What are Embryonic Stem Cells?nUndifferentiated cells with the ability t
22、o form any adult cell. They can proliferate indefinitely in culture & provide unlimited source of adult cells like bones, muscle, liver or blood cells.nOriginally produced to treat infertility and surplus embryos were discarded. nEmbryos created using IVF then tested using DNA analysis when only 3 d
23、ays old and only a few cells in size. Embryonic Stem Cells:Researchers extract stem cells from a 5-7 days oldResearchers extract stem cells from a 5-7 days old blastocystblastocyst. .Stem cells can divide in culture to form more of their own kind, Stem cells can divide in culture to form more of the
24、ir own kind, thereby creating a stem cell line.thereby creating a stem cell line.The research aims to induce these cells to generate healthy tissue needed by patients.Two Sources of Embryonic Stem Cells1. Excess fertilized eggs from IVF (in-vitro fertilization) clinics2. Therapeutic cloning (somatic
25、 cell nuclear transfer)Tens of thousands of frozen embryos are routinely destroyed when couples finish their treatment.These surplus embryos can be used to produce stem cells. Regenerative medical research aims to develop these cells into new, healthy tissue to heal severe illnesses.Somatic Cell Nuc
26、lear Somatic Cell Nuclear TransferTransfer The nucleus of a donated egg is removed and replaced with the nucleus of a mature, somatic cell (a skin cell, for example). No sperm is involved in this process, and no embryo is created to be implanted in a womans womb.The resulting stem cells can potentia
27、lly develop into specialized cells that are useful for treating severe illnesses.International LegislationnEmbryonic Stem cell research is highly controversial not only in the United States but worldwide. nIn the past two years, many nations have begun to tolerate, if not to support, the research.nI
28、n the fall of 2004, the United Nations will consider enacting a global ban on both therapeutic and reproductive cloning.Stem Cell Research WorldwideUnited NationsnIn November 2003, a committee of the General Assembly defeated by only one vote a proposal to ban therapeutic cloning research. The Unite
29、d States delegation strongly supported this proposal.nIn September 2004, the General Assembly will again consider the proposal to ban the research.2004。4。22胚胎生物技术目标：中国是该技术的创造国和输出国而不是被动的引进和模仿交大药学院韩伟干细胞生物技术的定义转基因转基因克隆克隆胚胎胚胎转基因动物转基因动物克隆动物克隆动物胚胎干细胞胚胎干细胞胚胎生物技术的意义转基因动物转基因动物克隆动物克隆动物胚胎干细胞胚胎干细胞1. 基础研究基础研究2.
30、生产蛋白生产蛋白 抗体药物抗体药物3. 抗疾病动物抗疾病动物4. 异种器官移植异种器官移植1. 基础研究基础研究2. 保持优良基因保持优良基因3. 农业上引种农业上引种4. 生产转基因动物生产转基因动物5. 挽救濒危动物挽救濒危动物1. 提供可移植的提供可移植的 器官和组织，器官和组织， 避免免疫排斥反应，避免免疫排斥反应， 解决器官组织不足解决器官组织不足 问题。问题。2. 获取药物筛选获取药物筛选 所需细胞类型所需细胞类型生农医药生农医药生农生农医药医药干细胞生物技术相关公司举例转基因动物转基因动物克隆动物克隆动物胚胎干细胞胚胎干细胞1.PPL, 英国英国2. 3.2. GTC, 美国美国
31、4. 5.3. Nexia, 加拿大加拿大6. 7.4. Revivicor, 美国美国8. R1.Cyagra (美国）美国）2. 2.Genetic Savings 3. and Clone (美国）美国） 3.Clone International 4. (澳大利亚）澳大利亚） .au4. Infigen (美国）美国） 1.ACT(美国）美国）2. 2.Geron (美国）美国）3. 3.BresaGen (澳大利亚）澳大利亚）4. .au4.Nucleotech (美国）美国）5. 干细胞生物技术在西方的困惑和限制人胚胎干细胞治疗性克隆技术在西方国家阻力非常大： “非自然”，“违反上
32、帝意志”，“谋杀”德国禁止治疗性克隆的研究。美国治疗性克隆不能得到联邦政府资助。在2001年8月前建立的人胚胎干细胞系方可用美国政 府资助研究。转基因技术生产的蛋白药物审批手续冗长（15-20年）。干细胞生物技术在美国的现状美国前10名大学美元（万）哈佛大学10000斯坦福大学1200加州大学500康州大学1000华盛顿大学500普林斯顿大学600哥伦比亚大学800耶鲁大学威斯康辛大学宾西法尼亚大学芝加哥大学干细胞研究中心干细胞研究中心干细胞研究所干细胞研究所干细胞兴趣小组干细胞兴趣小组干细胞沙龙干细胞沙龙中国在干细胞生物技术领域的优势宽松的社会环境，不同的伦理观念。政府的支持，反对生殖性克隆
33、人，允许治疗性克隆研究。中国在胚胎生物技术研究方面有比较好的开始，克隆羊、 牛、大鼠。曾溢滔、盛慧珍、陈大元、赵春华、韩忠朝、 曹谊林、陆道培，李凌松，裴雪涛等。清华、北大、复旦、上海生科院、上二医、中国医学科学 院、军事医学科学院等相继成立了专门干细胞研究机构。中国在干细胞生物技术领域的优势国家干细胞工程技术研究中心成立投入使用。国家干细胞工程产品产业化基地建成。筹建中的北京国家生物研究所。中科院干细胞生物学重点实验室。北京大学干细胞研究中心。清华大学组织工程和干细胞研究所。复旦大学干细胞和组织工程研究中心。浙江大学干细胞研究所。发展和壮大的必需条件发展和壮大的必需条件科研工作的合作精神科研
34、工作的合作精神干细胞生物技术在中国的干细胞生物技术的特点跨学科：转基因转基因克隆克隆胚胎胚胎转基因动物转基因动物克隆动物克隆动物胚胎干细胞胚胎干细胞生物学、农学、医学、药学生物学、农学、医学、药学干细胞生物技术的特点跨专业：转基因转基因克隆克隆胚胎胚胎转基因动物转基因动物克隆动物克隆动物治疗性克隆研究治疗性克隆研究胚胎生物学胚胎生物学干细胞生物学干细胞生物学分子生物学分子生物学发育生物学发育生物学组织工程学组织工程学遗传学遗传学免疫学免疫学器官移植等器官移植等干细胞生物技术交大的优势：生农医药蓄势待发。上海交通大学胚胎生物技术中心？上海交通大学胚胎生物技术中心？盛慧珍盛慧珍赵春华赵春华科研合作
35、科研合作杨向中杨向中Geron, ACT 美国公司合作美国公司合作Malcolm A.S. Moore国内国内国外国外组织形式目标驱动的目标驱动的个人组织个人组织如：如：973 盛慧珍盛慧珍“干细胞的基础研究与临床应用干细胞的基础研究与临床应用”1。陈大元，中科院动物所。陈大元，中科院动物所2。裴雪涛，军事医学科学院。裴雪涛，军事医学科学院3。盛慧珍，中科院。盛慧珍，中科院4。魏于全，四川大学。魏于全，四川大学5。杨雄里，复旦大学。杨雄里，复旦大学围绕新目标围绕新目标建立具有建立具有自我自我完善机制完善机制的的组织实体，适应带动组织实体，适应带动传统学科发展的传统学科发展的时代要求时代要求 虚拟组织虚拟组织实体组织实体组织目标目标组织组织行动行动
