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HistoryandBackgroundofHuman

Genome

ProjectHaroldRiethman,Ph.D.

[email protected]


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I. Background&OverviewofTheHuman

GenomeProject

II.ChromosomeStructureReview


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PhysicalBasisFor

GeneticInformation

Replication

Finite

Watsonand

Crick,

Nature

1953


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Image from NHGRI Web Site

Human male karyotype


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MendelsLawsofInheritance Mendel,1866

Genetic

Linkage

Maps Sturtevant,

1913

PhysicalMaps(Cytogenetic) Painter,1933

Structureof

DNA Watson

&

Crick,

1953

MolecularCloning CohenandBoyer,1972

DNASequencing Sanger,

Maxam,

and

Gilbert,Midseventies

HumanRFLPMapping Botsteinetal.,1980

GenomicsPre1982:


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Genomics 19821988:

Pulsedfield gel electrophoresis Schwartz and Cantor, 1983

Polymerase Chain Reaction Mullis 1983

Multipoint Linkage Mapping Lathrop et al., 1984.

Global Physical Clone Maps Olson et al., 1985

Sulston et al., 1985

First proposals to sequence DeLisi, DOE, mid80s

human genome

Yeast Artificial Chromosome (YAC) Burke et al., 1987

Automated DNA Sequencers Hood, midlate 80s

Mapping and Sequencing the National Research Council, 1988

Human Genome


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1. PulsedfieldGel

Electrophoresis

2. Maplarge

chromosomeregions

3. 2.Largeinsert

Cloning

FromRiethmanetal.,GenomeAnalysisVol 1,1997


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PolymeraseChainReaction

Invitro

synthesis

of

DNA

Markerstorageanddistribution

bycomputer

Sensitivescreeningand

detection

FromFanningandGibbs,GenomeAnalysisVol 1,1997


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MappingandSequencingtheHumanGenome

NationalResearch

Council,

1988

Aspecialeffortshouldbeorganizedandfundedtomap,sequence,andincreaseunderstandingofthe

HumanGenome.

Technologydevelopmentessentialforeveryphaseoftheproject.

Earlygoalsshouldbeahighresolutiongeneticmap,acollectionoforderedclones,andaseriesof

complementaryphysicalmapsofincreasingresolution.Thenucleotidesequence,theultimategoal,will

requiremajoradvancesinDNAhandlingandsequencingtechnologies.

Acomparativegeneticapproachisessentialforinterpretingtheinformationinthehumangenome.

Competing,peerreviewedprogramsemphasizingtechnologydevelopment.

Componentsubprojectsshouldhavethepotentialtoimproveby5to10foldincrementsthescaleor

efficiencyofmapping,sequencing,analyzing,orinterpretingtheinformationinthehumangenome.

Establishcentralizedfacilitiesforstoringanddistributingclones,andadatacenterforthecomputer

basedcollectionanddistributionoflargeamountsofDNAsequenceinformation.


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Maps

Landmarks Fragment Ends

DNA probes Restriction Enzyme

STS Meiosis

RadiationClone ends


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SequenceTaggedSite(STS)1989

ReplacesclonedDNAprobemappinglandmarkswithPCRassays.

EachSTSisuniquelydescribedbyapairofoligonucleotides,aproductsize,and

PCRreactionconditions. Canbestoredanddistributedelectronically.

Enables mergingofmappingdataobtainedfrommanylabsusingmanydifferentmethods

intoasingleconsensusmapoflandmarksalongachromosome.

Eliminatestheneedforhugecollectionsofclonedprobesegmentsuponwhich

priormaps

depended.

UniversalLandmark


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MeioticBreaks GeneticLinkageMaps


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RadiationinducedBreaks RadiationHybrid(RH)Maps


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Cloneends

Clonebased

PhysicalMap


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Thefirst5yearplan(19911995)

GeneticMap:

fullyconnected,2to5centimorgan resolution,eachmarkeridentifiedbyanSTS

PhysicalMap:

100kbresolutionSTSmap

2Mbcontigs formostofgenome

Sequencing:

Improve/developtechnologytoreducecostto0.50/base

Generatetotalof10Mbofhumansequenceinlargestretchesduringtechnologydevelopment

ModelOrganisms:

Geneticmap

of

mouse

Generatetotalof20Mbofsequencefrommodelorganisms,focusoncontiguousstretchesof1Mb,during

technologydevelopment

Informatics:

Developsoftware

and

database

designs

to

support

large

scale

mapping

&

sequencing

Createdatabasetoolsthatprovideeasyaccessandpermitcomparisonsofuptodatedatasets

Developalgorithmsandanalyticaltoolstointerpretinformation

ELSI Ethical,Legal,andSocialImplications

Training

TechnologyDevelopment

Support

innovative

&

high

risk

projects

TechnologyTransfer toindustryandthemedicalcommunity


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2nd 5yrplan

19931998

HGP

FrancisCollinsandDavidGalas

OriginallypublishedinScience

262:4346(1993)


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Figure 2 Worldwide human genome sequencing progress is shown (measured as base pairs

of fin ished sequence deposited with GenBank).

F S Coll ins et al. Science 1998;282:682-689

Published by AAAS HGP3rd 5yrplan19982003


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CapillarySequencers,Automation,19971998


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Green,1997 WeberandMeyers,1997


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InternationalHumanGenomeSequencingConsortium

DraftHuman

Genome

Sequence:

Nature

Feb.15,

2001


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CeleraGenomicsDraftHumanGenomeSequence

ScienceFeb.16,

2001


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InternationalHumanGenomeSequencingConsortium

FinishedHumanGenomeSequence:Oct.21,2004


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AVisionfortheFutureofGenomicsResearch, Nature,April24,

2003.Collins,Green,Guttmacher &Guyer,NHGRI


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Genomics

to

Biology

Comprehensivelyidentifythestructuralandfunctionalcomponents

encodedinthehumangenome.Encode.

Elucidatetheorganizationofgeneticnetworksandproteinpathwaysand

establishhowtheycontributetocellularandorganismalphenotypes.

Develop

a

detailed

understanding

of

the

heritable

variation

in

the

human

genome.HapMap

Understandevolutionaryvariationacrossspeciesandthemechanisms

underlying

it.

Developpolicyoptionsthatfacilitatethewidespreaduseofgenome

informationinbothresearchandclinicalsettings


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Technologydevelopment

DNAsequencing

Geneticvariation

Thegenome

'parts

list

Proteomics

Pathwaysandnetworks

Geneticcontributionstohealth,diseaseanddrugresponse

Molecularprobesforexploringbasicbiologyanddisease

Computationalresourcesforstorage,analysis,andintegrationof

hugedatasets


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Theabilitytodetermineagenotypeatverylowcost,allowinganassociation

studyinwhich2,000individualscouldbescreenedwithabout400,000genetic

markersfor$10,000orless.

Theability

to

sequence

DNA

at

costs

that

are

lower

by

four

to

five

orders

of

magnitudethanthecurrentcost,allowingahumangenometobesequenced

for$1,000orless.

The

ability

to

synthesize

long

DNA

molecules

at

high

accuracy

for

$0.01

per

base,allowingthesynthesisofgenesizedpiecesofDNAofanysequencefor

between$10and$10,000.

TheabilitytodeterminethemethylationstatusofalltheDNAinasinglecell.

Theabilitytomonitorthestateofallproteinsinasinglecellinasingle

experiment.

QuantumLeaps


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ChromosomeStructure


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Imagefrom

NHGRI

Web

Site

Humanmalekaryotype


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Gene

GeneticConcept Functionalpieceof achromosome

PhysicalConcept Stringofnucleotidesthatencodea

discretefunction

Proteinencodinggene

NoncodingRNAs


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Heterochromatin

Highly condensed chromosome regions

Large heterochromatic tracts are usually repetitive

sequence

Can be very difficult to clone and assemble sequences

Often gene-poor, low transcription levels

Enriched near centromeres and telomeres


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Heterochromatic Centromere Satellites

Heterochromatin


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Euchromatin

Less condensed chromosome regions

Very wide range of DNA sequence types andorganization

Contains most genes, but very unevenly distributed

Mostly accessible to cloning and assembly


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Euchromatic Region

FromUCSC

Web

Browser


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SegmentalDuplicatons

LargesegmentsofrecentlyduplicatedDNA

>1kb,

>90

%

Highlyenrichedinpericentromeresandsubtelomeres

Associated

with

large

polymorphisms

and

diseaseassociateddeletions


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Hillieretal.

Nature2003


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ChromosomeFunction


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Replication


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DNAReplication


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Centromeresand

Telomeres


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Centromeres

ensure

proper

segregation

of

chromosomes

Sequence Organization of Human Centromere and Pericentromere


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SequenceOrganizationofHumanCentromereandPericentromere

She et al. Nature 2004


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Telomeresarerequiredforchromosomereplicationandstability


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http://www.biologyreference.com/

Telomerase

replicates

telomeres


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Subtelomeres Rapidlyevolving

Highlyvariable

Duplicatedsequences


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Pryde etal,1997.

Curr Opin GenetDev


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Cell

cycle,

Meiosis,

Mitosis


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I=interphase

M=mitosis

S=DNAsynthesis

CellCycle


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MarstonAL,

Amon

A.

2004.

Nat

Rev

Mol

Cell

Biol.

MitosisandMeiosis

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