基因组学课件：Understanding The Complexity ofGenomes

1、Understanding The Complexity of Genomes,Jun YU BIG, CAS,The Power of Mind and Knowledge: 哪一个图是正确的?,“Judgment is a utensil proper for all subjects and will have an oar in everything.” Michel Eyquem de Montaigne (French, 28 February 1533 13 September 1592),2020/8/27,基因组学入门,3,判断力是一个万能的工具，放之四海而皆准。,科学的目的

2、是解释,“To explain a phenomenon is to give an account of its causal history.” PETER LIPTON, INFERENCE TO THE BEST EXPLANATION 30 (2nd ed., 2004). “. . . it is the aim of science to find satisfactory explanations or whatever strikes us as being in need of explanation.” KARL POPPER, OBJECTIVE KNOWLEDGE:

3、AN EVOLUTIONARY APPROACH 191 (rev. ed., 1981).,2020/8/27,基因组学入门,4,Evidence, Model, and Explanation,2020/8/27,基因组学入门,5,基于证据的解释 基于模型和理论的解释 基于证据、概念、模型、理论的解释,历史常常由胜利者来书写。,Integrity: 整实性,Integrityis a concept ofconsistencyof actions, values, methods, measures, principles, expectations, and outcomes. Inet

4、hics, integrity is regarded as the honesty and truthfulnessoraccuracyof ones actions. Integrity can be regarded as the opposite ofhypocrisy,1in that integrity regards internal consistency as a virtue, and suggests that parties holding apparently conflicting values should account for the discrepancy

5、or alter their beliefs. The word integrity stems from the Latin adjectiveinteger(whole, complete).2In this context, integrity is the inner sense of wholeness deriving from qualities such ashonestyand consistency ofcharacter. As such, one may judge that others have integrity to the extent that they a

6、ct according to the values, beliefs and principles they claim to hold.,2020/8/27,基因组学入门,6,Evolutionary Timeline,Lamarck (evolution),Darwin (evolution, natural selection),2020/8/27,基因组学入门,7,2020/8/27,基因组学入门,8,Darwins Theory: 适者生存,Individual organisms in nature differ from one another. Some of this va

7、riation is inherited Organisms in nature produce more offspring than can survive, and many of these offspring do not reproduce Because more organisms are produced than can survive, members of each species must compete for limited resources Because each organism is unique, each has different advantag

8、es SBL, sequence-by-ligation; SBP, sequence-by-pyrosequencing; SM, single molecule; FD, fixed DNA; FE, fixed enzyme; PoC, proof-of-concept; ?, expected technology,2020/8/27,21,基因组学入门,多代共存：个体化基因组研究、测定所有物种、metagenomics、pangenomics、多层次数据的获取、niche market等决定 多原理共存：缺乏“完美”技术、专利的限制和新原理的不断出现 高低通量共存：科研需求、应用需求

9、、市场需求 长读长与短读长共存：读长、质量、通量和价格 等无法同时满足 常规应用和新应用共存：新的应用还在不断被发现,DNA测序技术发展的切入点,2020/8/27,基因组学入门,22,Zero Mode Waveguide,Each SMRT chip contains thousands of ZMWs,Each ZMW is a cylindrical hole tens of nanometers in diameter, perforating a thin metal (e.g. aluminum) film supported by a transparent substrate

10、,2020/8/27,23,基因组学入门,When the ZMW is illuminated through the transparent substrate by laser light, the wavelength of the light is too large to pass through the waveguides aperture. Attenuated evanescent light from the excitation beam penetrates the lower 20-30nm of each waveguide, creating a detecti

11、on volume of only 20 zeptoliters (10-21 liters). This dramatic reduction in the detection volume provides the needed 1000-fold improvement in rejection of background fluorescence.,2020/8/27,24,基因组学入门,Immobilized Polymerase,Selective immobilization of polymerase to the fused silica floor of the ZMW w

12、as achieved by passivation of the metal cladding surface using polyphosphonate chemistry, producing enzyme density contrasts of glass over aluminum in excess of 400:1. Yields of single-molecule occupancies of approximately 30% were obtained for a range of ZMW diameters.,2020/8/27,25,基因组学入门,2020/8/27

13、,26,基因组学入门,2020/8/27,27,基因组学入门,2020/8/27,28,基因组学入门,First Ready for Market in 2010,Pacific Biosciences predicts that by 2013, its machines will be able to sequence a persons genome in 15 minutes, for less than $1,000. Technology Review January/February 2009， Interpreting the Genome By Emily Singer,20

14、20/8/27,29,基因组学入门,PacBios Technology,Current version at least 1,500 bp read length at launch next year short run time: 15 minutes the current DNA polymerase, which runs at 3 to 5 bases per second ZMW fill-in rate currently one-third $60 in reagents per run machine cost: as much as current high-throu

15、ghput sequencers Future version 100 gigabases of sequence per hour grow to between 50,000 and 100,000 bp in length at 15 bases per second 80 percent to 90 percent fill-in rate a million ZMW,2020/8/27,30,基因组学入门,IonTorrent and Proton II,2020/8/27,基因组学入门,31,New Promises,2020/8/27,基因组学入门,32,生物纳米孔 (a) -溶

16、血素蛋白,Oxford Nanopore Technologies said that it plans to commercialize two DNA strand sequencing instruments this year: a higher-throughput version that uses its GridIon platform and a disposable MinIon system that will be the size of a USB memory stick and cost less than $900. Running 20 GridIon ins

17、truments, or “nodes,” in parallel will enable users to sequence a human genome at 15-fold coverage in 15 minutes for less than $10 per gigabase.,The recent promises,HiSeq2000 to 2500 IonTorrent to Proton II Goals: $1000 and $100 per human genome within a day,2020/8/27,33,基因组学入门,2500, January 11, 201

18、2,Illumina announced the introduction of its HiSeq 2500 and upgrades to its MiSeq platform yesterday, according to a company press release. The HiSeq 2500 is expected to be able to sequence a genome in about a day, and, as our sister publication In Sequence reports, it is priced at $740,000 or $50,0

19、00 for an upgrade from the HiSeq 2000. In addition, the MiSeq will have increased throughput, longer reads, and a faster run time, Illumina says. Illuminas announcement came the same day as one from Life Technologies, setting up a sequencing battle, as Matthew Herper at Forbes puts it. The differenc

20、e between the machines, he adds, will be data quality and cost. Costs: $1000 per genome in a day? Machine cost?,2020/8/27,34,基因组学入门,3nd-generation sequencers should offer us: Supreme contiguity (tens of kb) Detection at single-molecule level and of covalent modifications Single-cell capacity in samp

21、le processing Measurement in real-time Quantitative and no amplifications,Why New Sequencers?,2020/8/27,基因组学入门,Bocklandt S, Lin W, Sehl ME, Snchez FJ, Sinsheimer JS, et al. (2011) Epigenetic Predictor of Age. PLoS ONE 6(6): e14821. doi:10.1371/journal.pone.0014821,35,Dreams and Promises,Throughput (

22、low cost), contiguity (read length), and quality (fidelity) $1000 per genome $100 per genome Less than $100 per genome High-throughput but short reads (2ndG) High-throughput and long reads (2ndG ) High quality, high-throughput, and long reads (3ndG ),2020/8/27,36,基因组学入门,From HGP to New Paradigm,The

23、HGP Era (1984-2003): “One human genome” done, and we ignored the differences among individuals The Post HGP Era: the HapMap Projects and hundreds of genomes done, and we realized that the differences among people are vast in numbers The New Paradigm: tens of thousands of genomes done, and we realize

24、d that we can map all sequence variations and tailor them to human health and disease status,37,2020/8/27,基因组学入门,New Parameters,Dale Yuzuki blog: /the-ion-torrent-proton-compared-to-the-illumina-hiseq-2500/,2020/8/27,38,基因组学入门,人类基因计划带动经济发展的因素,“人类基因组计划”投入38亿美金，而截至2010年它的影响已经是带来796

25、0亿美金的经济价值，以及31万个就业机会。,2020/8/27,39,基因组学入门,2020/8/27,Improving Healthcare,Science of Medicine,Biology of Disease,Biology of Genome,Structure of Genome,基因组学发展不同阶段需要强调不同研究对象和研究目标,40,基因组学入门,Churchs PGP,2020/8/27,41,基因组学入门,Personal Genome withIdentity Revealed,2020/8/27,42,Photo: Reuters,Photo: Reuters,L

26、evy, S. et al. PLoS Biol. 5, e254e286 (2007). Wheeler, D. A. et al. Nature 452, 872876 (2008).,$100M,$1M,基因组学入门,$100K,$50K,Japanese,2020/8/27,43,基因组学入门,Yes, we are now able to sequence individual genomes ($10,000-$5,000; seems not going down any time soon; $7,000 now offered in USA ) Yes, we can seq

27、uence only the exome (protein-coding sequences, 60Mb; 100 x or 200 x or $7,000 per exome); may not cheaper but easy for analysis Yes, we can sequence some genes individually even cheaper We can also sequence individual variations (sites),The “Personal Era” of Genomics Is Here!,2020/8/27,44,基因组学入门,20

28、20/8/27,For the First time: Whole-Genome Sequencing Delivers New Treatment Options for Twins with Rare Genetic Movement Disorder,45,基因组学入门,Yes, it is here the era of personalized medicinewhen physicians use whole-genome sequence to improve medical care,2020/8/27,Whole-Genome Sequencing for Optimized

29、 Patient Management. Bainbridge et al. 2011. Sci Transl. Med 3:87re3.,46,基因组学入门,50-Hour Whole Genome Sequencing Provides Rapid Diagnosis for Children With Genetic Disorders,Sauders et al., Sci Transl Med 3 October 2012 4:154ra135. DOI:10.1126/scitranslmed.3004041,2020/8/27,47,基因组学入门,Babys DNA in mot

30、hers blood,2020/8/27,48,基因组学入门,Fifty-Hour Whole Genome Sequencing Provides Rapid Diagnosis for Children With Genetic Disorders,2020/8/27,49,基因组学入门,C4RCD has four major components clinical evaluation and genomic diagnosis; counseling and optimization of conventional therapy; novel therapy development

31、; and community outreach. Each child being seen at the center will receive a clinical evaluation, including whole-genome sequencing.,Different from clinically integrated centers and research labs,2020/8/27,50,基因组学入门,Vision:To provide innovative genomic medicine for children with genetic diseases,htt

32、p://,Mission: To be a global leader and referral center for diagnosis, molecular understanding and treatment of inherited childhood diseases. We do this through: Timely molecular diagnosis Novel disease diagnosis Multidisciplinary team-based approaches to patient management,Tea

33、m members: experienced in genome analysis Board-certified Clinical Molecular Geneticists Board-certified Clinical Geneticists and Genetic Counselors Pediatricians representing every sub-specialty in the hospital Bioinformatics Experts Software Developers,2020/8/27,51,基因组学入门,2020/8/27,52,基因组学入门,2020/

34、8/27,53,基因组学入门,2020/8/27,54,The 600 Children Project,基因组学入门,2020/8/27,55,基因组学入门,2020/8/27,56,基因组学入门,2020/8/27,57,基因组学入门,What cheaper sequencing can do?,Genome-centric goals: genome sequences and genomic variomes for population genetics and evolution Cell-centric goals: transcriptomes and proteomes f

35、or cell biology, physiology, and pathology Epigenome-centric goals: methylomes and nucleosome marks/fringerprints for gene regulation and phenomes Ribogenome-centric goals: RNA-mediated mechanisms other than mRNAs for cellular mechanisms and cellular processes,2020/8/27,基因组学入门,58,Integrative Persona

36、l Omics Profiling,2020/8/27,基因组学入门,59,iPOP,2020/8/27,基因组学入门,60,2020/8/27,基因组学入门,61,2020/8/27,基因组学入门,62,GENETIC TESTS may be done for diverse purposes pertaining to clinical genetics, including the diagnosis of genetic diseases in children and adults; the identification of future disease risks; the p

37、rediction of drug responses; and the detection of risks of disease to future children.,Pre-implantation diagnosis Prenatal diagnosis Newborn screening Carrier testing Diagnostic testing Pre-symptomatic testing Predictive testing,2020/8/27,63,基因组学入门,DIAGNOSIS stage at diagnosis),National Cancer Insti

38、tute Cancer Trends Progress Report 2005 Update ,晚期癌症的新发病例,前列腺癌,结肠癌,乳腺癌（女）,肛肠癌 阴道癌,2020/8/27,64,基因组学入门,2020/8/27,基因组学入门,65,2020/8/27,基因组学入门,66,Three-Tiered Research of Genome Biology,Genomics Dynamics in composition, gene organization, and genome organization Genome bio

39、logy Multi-tracks: infT, opeT, comT, homT, and plaT Mechanisms and processes Lineage-specific genome biology Vertebrates and arthropods,2020/8/27,基因组学入门,67,Reconciling Mendel with Darwin,2020/8/27,基因组学入门,68,interacting molecules,systems biology,the genomic synthesis in biology,2020/8/27,基因组学入门,69,Le

40、ading to New Synthesis and a Paradigm Shift: Start from the Central Dogma,基因组学入门,Reverse transcriptase was discovered by Howard Temin at the University of WisconsinMadison, and independently by David Baltimore in 1970 at MIT. The two shared the 1975 Nobel Prize in Physiology or Medicine with Renato

41、Dulbecco for their discovery.,2020/8/27,70,The RNA World Is Early but Not Small!,基因组学入门,The RNA World (Gilbert) Ribosyme (Cech and Altman); In 1989, Thomas R. Cech and Sidney Altman won the Nobel Prize in chemistry for their discovery of catalytic properties of RNA.” In 2006, Craig Mello and Andrew

42、won Nobel Prize on the discovery of RNA interference (RNAi).,2020/8/27,71,Central Dogma vs Integrative Biology,基因组学入门,72,Central Dogma,Modified Central Dogma,Ribogenome-CD,Ribo-Epigenome-CD,2020/8/27,DNA-centric View of at Four Dimensions,First dimensional or compositional dynamics (NUCLEOTIDES AND

43、GENOTYPES): Sequences and simple variations (SNP, indels, CNVs, satellites, CpG islands, other structural elements, etc.) Chemical modifications (5-methylcytosine, 5-hydroxymethylcytosine, A methylation in bacteria and J base in kinetoplatids) Second dimensional or structural dynamics (GENE AND GENO

44、ME): Protein-coding genes and other transcripts (TEs, ncRNAs, small RNAs, etc.) Genome duplications (whole genome, segmental, and tandem duplication, etc.) Genome and gene organization (clustering, sub-telomeric and centromeric regions, etc.),2020/8/27,73,基因组学入门,Third dimensional or organizational d

45、ynamics (CHROMOSOMES) DNA modifications and their structural roles Histone modifications and nucleosome positioning Chromosome replication Chromosomes folding and conformation in a context of cell physiology Fourth dimensional or functional dynamics (NUCLEUS AND PHENOTYPES) Chromosome anchorage on t

46、he nuclear envelope Active foci and transcription activities Distribution, movement, and segregations Conformation dynamics in a context of cellular differentiation,2020/8/27,基因组学入门,74,Three-Tiered Research of Genome Biology,Genomics Dynamics in composition, gene organization, and genome organizatio

47、n Genome biology Multi-tracks: infT, opeT, comT, homT, and plaT Mechanisms and processes Lineage-specific genome biology Vertebrates and arthropods,2020/8/27,基因组学入门,75,基因组生物学“五行”说,信息流：遗传学与变演论The Informational Track: Genetics and Evolution 操作流：细胞生物学与分子生物学 操作流之一：表观组学The Operational Track-I: Epigenomic

48、s 操作流之二：核酸组学The Operational Track-II: Ribogenomics 操作流之三：蛋白质组学The Operational Track-III: Proteomics 平衡流：生物化学、生理学与药理学 平衡流之一：能源流The Homeostatic Track-I: Energy 平衡流之二：物质流The Homeostatic Track-II: Matter 分室流：生命的起源、系统发育和发生The Compartmental Track: Origin and Ontology of Life 可塑流：表型可塑性与神经生物学 可塑流之一：表型可塑性The

49、 Plasticity Track-I: Phenotypic Plasticity 可塑流之二：行为与认知的可塑性The Plasticity Track-II: The Plasticity of Affect, Behavior, and Cognition,2020/8/27,基因组学入门,76,The 5-Track Genome Biology,Informational Track (genetics, genomics, evolution) DNA sequence and its variations Operational Track (molecular biology

50、, cell biology, physiology) Epigenomics (DNA-centric) Ribogenomics (RNA-centric) Proteomics (protein-centric) Homeostatic Track (pharmacology, biochemistry) Energy-centric Material-centric Compartmental Track (development, anatomy, histology, and origin of life) Origin of life Unicellular to multice

51、llular life Development and differentiation Plasticity Track (psychiatry, psychology, and neurobiology) Phenotypic plasticity Plasticity of affect, behavior and cognition,2020/8/27,77,基因组学入门,2020/8/27,基因组学入门,78,2020/8/27,Jun Yu. (2012). Life on Two Tracks. Genomics Proteomics Bioinf, online June 22,

52、 2012.,基因组学入门,79,Genotype,Phenotype,Genome,Transcriptome,Proteome,Metabolome,Epigenome,Ribogenome,Nucleosome-space occupancy,Histone Marks,Gene Clustering,mRNA Splicing,miRNAs,lncRNAs,Other Small RNAs,DNA modifications,IT-Major,OT-Epigenome,OT-Ribogenome,OT-Major,Chromatin Dynamics,Chromosome Foldin

53、g,RNA Degradation,CpG density,2020/8/27,Physiome,Exposome,RNA Editing,RNA Routing,RNA Modification,基因组学入门,80,Influences from Both EpiG and RiboG,Genotype,RiboG,EpiG,2020/8/27,基因组学入门,81,Influences from Both EpiG and RiboG,Genotype,RiboG,EpiG,2020/8/27,基因组学入门,A ribogenomic flip,A Epigenomic flpp,82,To

54、tal Mass per Cell865 ug/109cells Protein per Cell450 ug/109cells (52%) DNA per Cell20 ug/109cells (2.31%) RNA per Cell200 ug/109cells (23%) Fractions of each RNA group: rRNA0.84 tRNA0.14 mRNA0.02 72 X 103ribosomes/cell 669 X 103tRNAmolecules/cell,Mass parameters of E. coli,83,2020/8/27,基因组学入门,Transc

55、ription Load,2020/8/27,基因组学入门,84,Are all epigenetic factors environmental?,2020/8/27,fudan Lecture,85,Epigenetic,Non-genetic,2020/8/27,BIG-CAS,86,Phenotype VariabilityPenetrance and Expressivity,A specific example of incomplete penetrance is the human bone disease osteogenesis imperfecta (OI). The m

56、ajority of people with this disease have a dominant mutation in one of the two genes that produce type 1 collagen, COL1A1 or COL1A2. Collagen is a tissue that strengthens bones and muscles and multiple body tissues. People with OI have weak bones, bluish color in the whites of their eyes, and a vari

57、ety of afflictions that cause weakness in their joints and teeth. However, this disease doesnt affect everyone who has COLIA1 and COLIA2 mutations in the same way. In fact, some people can carry the mutation but have no symptoms. Thus, families can unknowingly transmit the mutation from one generati

58、on to the next through someone who carries the mutation but does not express the OI phenotype. Miko, I. (2008) Phenotype variability: penetrance and expressivity. Nature Education 1(1),2020/8/27,BIG-CAS,87,Penetrance,Interestingly, some scientists have actually tried to do this by observing how the

59、genetic mutations that cause OI affect mice. These investigators inserted a mutated form of COL1A1 into mice and bred them so that they all contained this mutation. The mice were affected in similar ways to those with human OI: Many had severe bone weakness and multiple bone fractures, even at birth. In fact, when the researchers examined the mouse bones closely, they found that 70% of mice with the mutated COL1A1 gene showed evidence of OI (bone fractures); however, the remaining 30% appeared completely normal. In these m