分子诊断学English课件

分子生物学进展分子生物学进展MD.,Ph.D.MD.,Ph.D.,200920091 1Introduction to Molecular Diagnostics2 2Presentation Outline Concept of Molecular Diagnostics History of Molecular Diagnostics Impact on Human Diseases Basis for Molecular Assay Management of the course3 31.Molecular DiagnosisMolecular diagnosis of human disorders is referred to as the detection of the various pathogenic mutations in DNA and/or RNA samples in order to facilitate detection,diagnosis,sub-classification,prognosis,and monitoring response to therapy.4 41.Molecular DiagnosticsThe use of molecular biology techniques to expand scientific knowledge of the natural history of diseases,identify people who are at risk for acquiring specific diseases,and diagnose human diseases at the nucleic acid level.5 51.Molecular DiagnosticsMolecular diagnostics combines laboratory medicine with the knowledge and technology of molecular genetics and has been enormously revolutionized over the last decades,benefiting from the discoveries in the field of molecular biology.6 6The The information revolutioninformation revolution in molecular in molecular biology is permeating every aspect of biology is permeating every aspect of medical practicemedical practiceThe rate of disease gene discovery is The rate of disease gene discovery is increasing exponentially,which facilitates increasing exponentially,which facilitates the understanding diseases at molecular the understanding diseases at molecular levellevelMolecular understanding of disease is Molecular understanding of disease is translated into diagnostic testing,translated into diagnostic testing,therapeutics,and eventually preventive therapeutics,and eventually preventive therapiestherapies1.Molecular Diagnostics:EmergenceEmergence7 71.Molecular Diagnostics:SignificanceSignificanceTo face the new century,the medical practitioner not only understand molecular biology,but must also embrace the use of this rapidly expanding body of information in his medical practice,whether practicing family medicine,oncology,obstetrics and gynecology,pathology,or any other medical specialty.8 8To introduce essential concepts in molecular diagnostics that impact on the identification of novel markers of human diseasesTo develop and apply useful molecular assays to monitor disease,determine appropriate treatment strategies,and predict disease outcomes.1.Molecular Diagnostics:Goal9 92.History of Molecular Diagnostics1865Gregor Mendel,Law of Heredity1866Johann Miescher,Purification of DNA19531970Recombinant DNA Technology1977DNA sequencing 1985In Vitro Amplification of DNA(PCR)2001The Human Genome ProjectWatson and Crick,Structure of DNAThe Molecular Biology TimelineSickle Cell Anemia Mutation 194910102.History of Molecular DiagnosticsPauling introduced the term molecular disease in the medical vocabulary,based on their discovery that a single amino acid change leads to a sickle cell anemia.In principle,their findings have set the foundations of molecular diagnostics.Sickle cell anemia is a genetic disease which is caused by a single nucleotide change in the 6th aa of the-chain of hemoglobin.Sickle cell anemia1111J.D.Watson and F.H.C.Crick(1953)Discovery of DNA Structure1212J.D.Watson and F.H.C.Crick(1953)A structure for deoxyribose nucleic acid.Nature 171:737“We wish to suggest a structure for the salt of deoxyribose nucleic acid(D.N.A.).This structure has novel features which are of considerable biological interest.”Discovery of DNA StructureOne of the most important biological discovery in the 20th century 1313Discovery of DNA StructureRosalind E.Franklin19201958The structure of DNA was determined using X-ray diffraction techniques.Much of the original X-ray diffraction data was generated by Rosalind E.Franklin.14142.History of Molecular DiagnosticsThe PCR Revolution Kary Mullis Kary Mullis 1985 41y Invention of PCR 1993 49y Received the Noble Prize 15152.History of Molecular DiagnosticsThe PCR Revolution PCR has greatly facilitated and revolutionized molecular diagnostics.Its most powerful feature-large amount of copies of the target sequence generated by its exponential amplification,-allows the identification of a known mutation within a single day.markedly decreased need for radioactivity,allowed molecular diagnostics to enter the clinical laboratory.1616 U.S.Government project coordinated by the Dept.of U.S.Government project coordinated by the Dept.of Energy and NIH Energy and NIH Goals of the Human Genome Project Goals of the Human Genome Project(19902006)(19902006)To identify all of the genes in human DNA;To identify all of the genes in human DNA;To determine the sequences of the 3 billion bases To determine the sequences of the 3 billion bases that make up human DNA;that make up human DNA;To create databases;To create databases;To develop tools for data analysis;and To develop tools for data analysis;and To address the ethical,legal,and social issues To address the ethical,legal,and social issues that arise from genome researchthat arise from genome research2.History of Molecular DiagnosticsHuman Genome Project1717Discovery of potential novel molecular markers of human diseases Identification of novel molecular markers of human diseases Utility of molecular markers to develop useful molecular assays for detection,diagnosis,and prediction of disease outcomes3.Impact on Human Diseases18183.Impact on Human Diseases:AdvantageAdvantageMonitor diseases more accurately Allows for early treatment and better patient careDetermine most appropriate treatment Reduces or eliminates unnecessary treatment Reduces or eliminates inadequate treatment Yields greater cost effectivenessReduce patient morbidity and mortality1919 Diagnostic-Identity of a disease Prognostic-Outcome of a disease Predictive-Possibility of a disease Therapeutic-Response of a disease to treatment3.Impact on Human Diseases:Practical application2020HEMATOLOGYINFECTIOUS DISEASE IDENTITYTESTINGGENETICDISEASE SOLIDTUMORSMolecularPathology3.Impact on Human Diseases21214.Basis for Technology:Fundamental(1)Fundamental(1)Advance in the understanding of the structure and chemistry of nucleic acids have facilitated the development of technologies that can be employed effectively in molecular diagnostics.22224.Basis for Technology:PlatformPlatformAmplification Techniques PCR polymerase chain reaction多聚酶链反应 LCR ligase chain reaction 连接酶链反应 NASBA nucleic-acid sequence-based amplification 核酸序列依赖的扩增Molecular Technologies in the Clinical LaboratoryDNA Sequencing 23234.Basis for Technology:PlatformPlatformElectrophoretic Methods SSCP(single-strand conformation polymorphism)单链构象多态性 DGGE(denaturing gradient gel electrophoresis)变性梯度凝胶电泳法Molecular Technologies in the Clinical LaboratoryHybridization Techniques Southern hybridization Blot Northern hybridization Blot24244.Basis for Technology:PlatformPlatformBiochip Technology DNA micro-array Protein micro-arrayMolecular Technologies in the Clinical LaboratoryRecombinant DNA Technology25254.Basis for Technology:Target specialtyTarget specialty Genetically-based diseases can be diagnosed Specificity can be controlled Single base changes can be detected Expression of gene product is not required Targets can be amplified 105Nucleic acids are targeted by molecular assays 26264.Basis for Molecular Assay:Pathogenesis(1)Pathogenesis(1)Diagnostic Distinguishing variants of human disease based on presence of specific molecular markers(chromosome translocations in Burkitts lymphoma:c-myc)Understanding molecular pathogenesis of human disease enables effective utilization of molecular assays27274.Basis for Molecular Assay:Pathogenesis(1)Pathogenesis(1)Prognostic Prediction of likely patient outcomes based on presence of specific molecular markers(gene mutations predicting clinical course in cancer)Understanding molecular pathogenesis of human disease enables effective utilization of molecular assays28284.Basis for Molecular Assay:Pathogenesis(2)Pathogenesis(2)Understanding molecular pathogenesis of human disease enables effective utilization of molecular assaysTherapeutic Prediction of response to specific therapies based on presence of specific molecular markers(gene mutations predicting poor drug sensitivity in lung cancer:p53,k-ras)2929 结束语当你尽了自己的最大努力时，失败也是伟大的，所以不要放弃，坚持就是正确的。When You Do Your Best,Failure Is Great,So DonT Give Up,Stick To The End谢谢大家荣幸这一路，与你同行ItS An Honor To Walk With You All The Way演讲人：XXXXXX 时 间：XX年XX月XX日