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1、,为什么要用分子生物学技术诊断人体疾病? (Why do we need molecular biology techniques for clinical diagnosis?),不同层次检验分析,分子水平 (molecular) 生化水平 (biochemistry) 细胞水平 (cell) 组织水平 (tissue) 个体水平 (individual) 群体水平 (population),致病有机体,人体内部系统自身紊乱,（微生物寄生虫等）,（如：malaria）,原核,真核,胞膜、胞浆,胞核（染色体等）,蛋白、糖、脂核酸（DNA、RNA等）,分子水平,组织,胞膜、胞浆（包括线粒体等），
2、胞核（染色体等）,蛋白、糖、脂核酸（DNA、RNA等-）,化学物、空气等,机体水平,(组织）细胞水平,亚细胞结构,细胞,人 体 疾 病 Human Disease,microRNA, siRNA,外伤 Trauma,什么是分子生物学？ What does molecular biology mean?,Molecular biology is the study of biology at a molecular level. The field overlaps with other areas of biology and chemistry, particularly genetics
3、and biochemistry. Molecular biology chiefly concerns itself with understanding the interactions between DNA, RNA and protein biosythesis and learning how these interactions are regulated,Expression cloning Polymerase chain reaction (PCR) Gel electrophoresis Southern blotting Northern blotting Wester
4、n blotting Sequencing Linkage anlysis Arrays Allele specific oligonucleotide Abandoned technology,什么是分子生物学技术？ What are techniques of molecular biology?,分子生物学技术字在临床检验中的应用,1.感染性疾病中应用 2.人类疾病相关基因的检查 3.肿瘤的检测(differentiate)、预防和治疗（包括疗效观察） 4.pharmacogenomics 5.组织配型 HLA 2（DR、Q） 6.亲子鉴定 7.法医鉴定,Cleveland Clinic
5、al Foundation,ARUP,遗传性疾病的类型,1. Chromsomal disease 2. Single-gene disease, monogenic disease Autosomal dominant inheritance, AD Autosomal recessive inheritance, AR X-linked dominant inheritance, XD X-linked recessive inheritance, XR Y-linked inheritance,3. Polygenic disease, Multifactorial disease (1
6、) A. Major gene +other genes+other factors B. Many minor gene+other factors (2) Very important, including a lot of common diseases (3) Research methods Population genetics, STR linkage analysis, SNPs 4. Mitochondrial genic disease, mtDNA 5. Somatic cell genetics disease,遗传性疾病的类型 （续）,DNA 分子结构改变-基因突变,
7、1. Chromosome aberration 2. Gene mutation Point mutation Deletion Insertion Tandem repeat,基因突变的类型,1.transition or transversion same sense mutation missense mutation non-sense mutation termination codon mutation 2. Frameshift mutationa (including some splicing mutations) 3. Condon mutation (Condon in
8、sertion or insertion) 4. Mispaired synapsis and unequal crossing-over -Fusion gene 5. Dynamic mutation,3. Condon mutation (Condon insertion or insertion) 4. Mispaired synapsis and unequal crossing-over -Fusion gene 5. Dynamic mutation,基因突变的类型 (续）,疾病相关基因的检查,细胞遗传学（Cytogenetics） 分子遗传学（Molecular Genetic
9、s）,细胞遗传学（Cytogenetics）,是遗传学的一个分支 主要研究染色体和细胞分裂( Chromosomes and cell division) 常用的分析技术包括G带染色(G-banded)等和分子细胞遗传学技术(Molecular Cyrogenetics) 如FISH (Fluorescent in situ hybridization) 和CGH(Comparative genomic hybridization),Human male karyotype,A metaphase cell positive for the bcr/abl rearrangement usin
10、g FISH,Interphase cells positive for a t(9;22) rearrangement,分子遗传学（Cytogenetics）,Molecular genetics is the field of biology which studies the structure and function of genes at a molecular level. The field studies how the genes are transferred from generation to generation. Molecular genetics employ
11、s the methods of genetics and molecular biology. It is so-called to differentiate it from other sub fields of genetics such as ecological genetics and population genetics.,An important area within molecular genetics is the use of molecular information to determine the patterns if descent, and theref
12、ore the correct scientific classification of organisms: this is called molecular systematics. Along with determining the pattern of descendants, molecular genetics helps in understanding genetics mutations that can cause certain types of disease. Through utilizing the methods of genetics discovers t
13、he reseasons why traits are carried on and how and why some may mutate.,分子遗传学（Cytogenetics）(续）,分子遗传学技术 （ Techniques of molecular biology）,Forward genetics Reverse genetics Gene Therapy Techniques in molecular genetics Amplification: PCR and cloning DNA in bacteria Separation and detection: cell cult
14、ure, DNA Isolation, mRNA isolation The molecular genetics project 基因表达定量,检查突变的方法,SSCP DGGE DHPLC Digestion Prime 3- ASO Direct Sequencing Genotyping (STR) Snapshot,Pharmacogenomics简介,疾病相关基因检查的质量控制,1.检查灵敏度 2.检查特异性 3.准确性 4.质量控制,疾病相关基因检查的未来展望,遗传基因几乎与所有人类疾病相关,医学科学的发展使感染性疾病和营养性因素所致疾病的发病率越来越低，并被逐渐控制。医学工作者的注意力已转向遗传性疾病的防治。随着人类遗传学和医学遗传学的发展，人们已认识到遗传因素存在于除外伤以外几乎所有的疾病之中。遗传学也已渗透到医学各学科领域, 其中包括疾病基因诊断的临床应用。,我国单基因疾病基因的检查现状与未来,方法学 需要大量的疾病基因鉴定工作 需要大量的病例的基因突变分析分析（Mutation screening) 建立完善的疾病基因及其突变数据库 国家FDA公认的检查方法,涉及的病人多，更为复杂 基础研究刚刚起步，还有大量的工作要做 最终建立一个疾病的多基因检查模型,我国多基因疾病基因的检查现状与未来,