The nucleosome: the fundamental unit of The nucleosome: the fundamental unit of chromosomal packaging of DNA with histoneschromosomal packaging of DNA with histones

DNA does not coil DNA does not coil smoothlysmoothly

Base sequences dictate Base sequences dictate preferred nucleosome preferred nucleosome positions along DNApositions along DNA

Spacing and structure Spacing and structure affect gene functionaffect gene function

Fig. 12.3 b

Models of higher level compaction seek to explain Models of higher level compaction seek to explain extreme compaction of chromosomes at mitosisextreme compaction of chromosomes at mitosis

Formation of 300 A Formation of 300 A fiber through fiber through supercoilingsupercoiling

Fig. 12.4 a

Models of higher level compaction seek to Models of higher level compaction seek to explain extreme compaction of chromosomes explain extreme compaction of chromosomes

at mitosisat mitosis Radial loop-scaffold Radial loop-scaffold

model for higher model for higher levels of compactionlevels of compaction Each loop contains Each loop contains

60-100 kb of DNA 60-100 kb of DNA tethered by tethered by nonhistone scaffold nonhistone scaffold proteinsproteins

Fig. 12.4 b

Radial loop-scaffold model Radial loop-scaffold model continuedcontinued

Fig. 12.4 c

A closer look at karyotypes: fully compacted A closer look at karyotypes: fully compacted metaphase chromosomes have unique, metaphase chromosomes have unique,

reproducible banding patternsreproducible banding patterns

Banding Banding patterns are patterns are highly highly reproduciblereproducible

Not known Not known what they what they representrepresent

Fig. 12.6 a

A closer look at karyotypesA closer look at karyotypes

Banding Banding patterns help patterns help locate geneslocate genes

Fig. 12.6 b

Polytene chromosomes are an Polytene chromosomes are an invaluable tool for geneticistsinvaluable tool for geneticists

in situin situ hybridization hybridization of white gene to a of white gene to a single band (3C2) single band (3C2) near the tip of the near the tip of the Drosophila X Drosophila X chromosomechromosome

Fig. 12.15 c

Polytene chromosomes are an Polytene chromosomes are an invaluable tool for geneticistsinvaluable tool for geneticists

Banding patterns can Banding patterns can be used to analyze be used to analyze chromosomal chromosomal differences between differences between speciesspecies

Can also be used to Can also be used to reveal cause of reveal cause of genetic diseasegenetic disease e.g., Downs e.g., Downs

syndrome – 3 copies syndrome – 3 copies of chromosome 21of chromosome 21

Fig. 12.6 c

A closer look at karyotypesA closer look at karyotypes

Specialized chromosomal elements ensure accurate Specialized chromosomal elements ensure accurate replication and segregation of chromosomesreplication and segregation of chromosomes

There are many origins of replicationThere are many origins of replication Replication occurs in about 8 hours during S phase in Replication occurs in about 8 hours during S phase in

actively dividing human cellsactively dividing human cells DNA polymerase can assemble new DNA at a rate of DNA polymerase can assemble new DNA at a rate of

about 50 nucleotides per secondabout 50 nucleotides per second Many origins of replication are required to complete Many origins of replication are required to complete

the task of copying the DNA in a genomethe task of copying the DNA in a genome In mammals, there are 10,000 origins of replicationIn mammals, there are 10,000 origins of replication Origins of replication are scattered throughout the Origins of replication are scattered throughout the

chromatin, 30 – 300 kb apartchromatin, 30 – 300 kb apart

Structure of yeast origin of Structure of yeast origin of replicationreplication

Autonomously replicating sequences (ARSs) in Autonomously replicating sequences (ARSs) in yeast consist of an A – T rich regionyeast consist of an A – T rich region

ARSs permit replication of plasmids in yeast cellsARSs permit replication of plasmids in yeast cells

Fig. 12.11 b

Telomeres preserve the integrity of Telomeres preserve the integrity of linear chromosomeslinear chromosomes

Telomeres are Telomeres are protective caps on protective caps on eukaryotic eukaryotic chromosomeschromosomes Prevent fusion with Prevent fusion with

other chromosomesother chromosomes Protect tips from Protect tips from

degradationdegradation Solve the end-Solve the end-

replication problemreplication problemFig. 12.8

DNA DNA polymerase polymerase cannot cannot reconstruct 5’ reconstruct 5’ end of a DNA end of a DNA strandstrand

Fig. 12.9

Binding of Binding of telomerase to telomerase to TTAGGG and TTAGGG and addition of addition of RNA extends RNA extends the endsthe ends

Fig. 12.10

Segregation of condensed chromosomes Segregation of condensed chromosomes depends on centromeresdepends on centromeres

Centromeres appear as constrictions on Centromeres appear as constrictions on chromosomeschromosomes Contained within blocks of repetitive, noncoding Contained within blocks of repetitive, noncoding

sequences called satellite DNAsequences called satellite DNA Satellite DNA consists of short sequences 5-300 bases in Satellite DNA consists of short sequences 5-300 bases in

lengthlength Centromeres have two functionsCentromeres have two functions

Hold sister chromatids togetherHold sister chromatids together Kinetochore – structure composed of DNA and protein Kinetochore – structure composed of DNA and protein

that help power chromosome movementthat help power chromosome movement

Centromere structure and functionCentromere structure and function

Fig. 12.11 a

Structure of yeast centromereStructure of yeast centromere

Fig. 12.11 b

Studies using DNase identify Studies using DNase identify decompacted regionsdecompacted regions

Fig. 12.12 a

Position effect variegation in Drosophila: moving a Position effect variegation in Drosophila: moving a gene near heterochromatin prevents it expressiongene near heterochromatin prevents it expression

Facultative Facultative heterochromatinheterochromatin Moving a gene Moving a gene

near near heterochromatin heterochromatin silences its silences its activity in some activity in some cells and not cells and not othersothers

Fig. 12.14 a

Comparing the mouse and Comparing the mouse and human genomeshuman genomes

The loss or gain of one or more The loss or gain of one or more chromosomes results in aneuploidychromosomes results in aneuploidy

Autosomal aneuploidy is harmful to the organismAutosomal aneuploidy is harmful to the organism

Monosomy usually lethalMonosomy usually lethal Trisomies – highly deleteriousTrisomies – highly deleterious

Trisomy 18 – Edwards syndromeTrisomy 18 – Edwards syndrome Trisomy 13 – Patau syndromeTrisomy 13 – Patau syndrome Trisomy 21 – Down syndromeTrisomy 21 – Down syndrome

Humans tolerate X chromosome aneuploidy Humans tolerate X chromosome aneuploidy because X inactivation compensates for dosagebecause X inactivation compensates for dosage

Fig. 13.27

Meiotic nondisjunctionMeiotic nondisjunction Failure of two sister chromatids to separate during meiotic Failure of two sister chromatids to separate during meiotic

anaphaseanaphase Generates reciprocal trisomic and monosomic daughter cellsGenerates reciprocal trisomic and monosomic daughter cells

Chromosome lossChromosome loss Produces one monosomic and one diploid daughter cellProduces one monosomic and one diploid daughter cell

Fig. 13.28 a

Mosaics – aneuploid and normal tissues that lie Mosaics – aneuploid and normal tissues that lie side-by-sideside-by-side Aneuploids give rise to aneuploid clonesAneuploids give rise to aneuploid clones

Fig. 13.28 b

Gynandromorph in Drosophila results from female Gynandromorph in Drosophila results from female losing one X chromosome during first mitotic losing one X chromosome during first mitotic

division after fertilizationdivision after fertilization

Fig. 13.29

Euploid individuals contain only Euploid individuals contain only complete sets of chromosomescomplete sets of chromosomes

Monoploid organisms contain a single copy of Monoploid organisms contain a single copy of each chromosome and are usually infertileeach chromosome and are usually infertile

Monoploid plants have many usesMonoploid plants have many uses Visualize recessive traits directlyVisualize recessive traits directly Introduction of mutations into individual cellsIntroduction of mutations into individual cells Select for desirable phenotpyes (herbicide Select for desirable phenotpyes (herbicide

resistance)resistance) Hormone treatment to grow selected cellsHormone treatment to grow selected cells

Fig. 13.30

Treatment with colchicine converts back to diploidTreatment with colchicine converts back to diploid

plants that express desired phenotypesplants that express desired phenotypes

Fig. 13.30 c

Polyploidy has accompanied the Polyploidy has accompanied the evolution of many cultivated plantsevolution of many cultivated plants

1 out of 3 flowering plants are polyploid1 out of 3 flowering plants are polyploid Polyploid often increases size and vigorPolyploid often increases size and vigor Often selected for agricultural cultivationOften selected for agricultural cultivation

Tetraploids - alfalfa, coffee, peanutsTetraploids - alfalfa, coffee, peanuts Octaploid - strawberriesOctaploid - strawberries

Fig. 13.31

Triploids are almost Triploids are almost always sterilealways sterile

Result from union of Result from union of monoploid and monoploid and diploid gametesdiploid gametes

Meiosis produces Meiosis produces unbalanced gametesunbalanced gametes

Fig. 13.32

Tetraploids are often source of new speciesTetraploids are often source of new species Failure of chromosomes to separate into two Failure of chromosomes to separate into two

daughter cells during mitosis in diploiddaughter cells during mitosis in diploid Cross between tetraploid and diploid creates Cross between tetraploid and diploid creates

triploids – new species, autopolyploidstriploids – new species, autopolyploids

13.33 a

Maintenance of tetraploid Maintenance of tetraploid species depends on the species depends on the production of gametes with production of gametes with balanced sets of balanced sets of chromosomeschromosomes

Bivalents- pairs of Bivalents- pairs of synapsed homologous synapsed homologous chromosomes that ensure chromosomes that ensure balanced gametesbalanced gametes

Fig. 13.33 b

Fig. 13.33 c

Some polyploids have agriculturally desirable traits Some polyploids have agriculturally desirable traits derived from two speciesderived from two species

Amphidiploids created by Amphidiploids created by chromosome doubling in chromosome doubling in germ cellsgerm cells

e.g., wheat – cross e.g., wheat – cross between tetraploid wheat between tetraploid wheat and diploid rye produce and diploid rye produce hybrids with desirable hybrids with desirable traitstraits

Fig. 13.34

Deletions Deletions remove remove genetic genetic

material material from from

genomegenome

Fig. 13.2

Phenotypic consequences of Phenotypic consequences of heterozygosityheterozygosity

Homozygosity Homozygosity for deletion is for deletion is often but not often but not always lethalalways lethal

Heterozygosity Heterozygosity for deletion is for deletion is often often detrimentaldetrimental

Fig. 13.3

Mapping distances affected in Mapping distances affected in deletion heterozygotesdeletion heterozygotes

Recombination between homologues can only occur if both Recombination between homologues can only occur if both carry copies of the genecarry copies of the gene

Deletion loop formed if heterozygous for deletionDeletion loop formed if heterozygous for deletion Genes within the loop cannot be separated by Genes within the loop cannot be separated by

recombinationrecombination

Fig. 13.4 a

Deletion loops in polytene Deletion loops in polytene chromosomeschromosomes

Fig. 13.4 b

Deletions in heterozygotes can Deletions in heterozygotes can uncover genesuncover genes

Pseudodominance shows a deletion has Pseudodominance shows a deletion has removed a particular generemoved a particular gene

Fig. 13.5

Deletions can be used to locate genesDeletions can be used to locate genes Deletions to assign Deletions to assign

genes to bands on genes to bands on Drosophila Drosophila polytene polytene chromosomeschromosomes

Complementation Complementation teststests

Deletion Deletion heterozygote heterozygote reveals reveals chromosomal chromosomal location of mutant location of mutant genegene

Fig. 13.6

Deletions to locate genes at the Deletions to locate genes at the molecular levelmolecular level

Labeled probe hybridizes to wild-type Labeled probe hybridizes to wild-type chromosome but not to deletion chromosome but not to deletion chromosomechromosome

Fig. 13.7 a

Molecular mapping of deletion Molecular mapping of deletion breakpoints by Southern blottingbreakpoints by Southern blotting

Fig. 13.7 b, c

Duplications add material to the Duplications add material to the genomegenome

Fig. 13.8 a,b

Duplication loops form when chromosomes pair in Duplication loops form when chromosomes pair in duplication heterozygotesduplication heterozygotes

In prophase I, the duplication loop can In prophase I, the duplication loop can assume different configurations that assume different configurations that maximize the pairing of related regionsmaximize the pairing of related regions

Fig. 13.8 c

Duplications can affect phenotypeDuplications can affect phenotype

Novel phenotypesNovel phenotypes More gene copiesMore gene copies Genes next to Genes next to

duplication duplication displaced to new displaced to new environment environment altering expressionaltering expression

Fig. 13.9

Unequal crossing over between duplications Unequal crossing over between duplications increases or decreases gene copy numberincreases or decreases gene copy number

Fig. 13.10

The effects of duplications and The effects of duplications and deletions on phenotpyedeletions on phenotpye

Heterozygosity creates imbalance in gene Heterozygosity creates imbalance in gene product altering phenotypes (some lethal)product altering phenotypes (some lethal)

Genes may be placed in new location that Genes may be placed in new location that modify expressionmodify expression

Deletions and duplications drive evolution Deletions and duplications drive evolution of the genomeof the genome

Inversions reorganize the DNA Inversions reorganize the DNA sequence of a chromosomesequence of a chromosome

180° rotation of 180° rotation of chromosomal chromosomal regions after regions after double-stranded double-stranded breakbreak

Rare crossover Rare crossover between related between related genes on a genes on a chromosomechromosome

Fig. 13.11a,b

An inversion can affect phenotype if An inversion can affect phenotype if it disrupts a geneit disrupts a gene

Fig. 13.11 c

Inversion heterozygotes reduce the Inversion heterozygotes reduce the number of recombinant progenynumber of recombinant progeny

Inversion loop in Inversion loop in heterozygote forms heterozygote forms alignment of alignment of homologous regionshomologous regions

Fig. 13.12

Gametes produced from pericentric and Gametes produced from pericentric and paracentric inversions are imbalancedparacentric inversions are imbalanced

Fig. 13.13

Pericentric inversion Paracentric inversion (cont’d) (cont’d)

Fig. 13.13 cont’d

Balancer chromosomes help Balancer chromosomes help preserve linkagepreserve linkage

Balancers carry Balancers carry multiple, overlapping multiple, overlapping inversionsinversions

Most contain a Most contain a dominant marker and dominant marker and recessive lethal recessive lethal mutation that mutation that prevents survival of prevents survival of homozygoteshomozygotes

Useful in genetic Useful in genetic manipulations and manipulations and mutant screensmutant screens

Fig. D.6

Translocations attach one part of a Translocations attach one part of a chromosome to anotherchromosome to another

Translocation – part Translocation – part of one chromosome of one chromosome becomes attached to becomes attached to nonhomologous nonhomologous chromosomechromosome Reciprocal translocation-Reciprocal translocation-exchangeexchange between between

nonhomologous chromosomesnonhomologous chromosomes

Robertsonian translocations can Robertsonian translocations can reshape genomesreshape genomes

Reciprocal exchange between acrocentric Reciprocal exchange between acrocentric chromosomes generate large metacentric chromosomes generate large metacentric chromosome and small chromosomechromosome and small chromosome Tiny chromosome may be lost from organismTiny chromosome may be lost from organism

Fig. 13.16

Banding patterns can Banding patterns can be used to analyze be used to analyze chromosomal chromosomal differences between differences between speciesspecies

Can also be used to Can also be used to reveal cause of reveal cause of genetic diseasegenetic disease e.g., Downs e.g., Downs

syndrome – 3 copies syndrome – 3 copies of chromosome 21of chromosome 21

Fig. 12.6 c

A closer look at karyotypesA closer look at karyotypes

Chronic myelogenous leukemia

Fig. 13.17

Heterozygosity for translocations diminishes Heterozygosity for translocations diminishes fertility and results in pseudolinkagefertility and results in pseudolinkage

Fig. 13.18 a.b

Three possible segregation patterns in a translocation Three possible segregation patterns in a translocation heterozygote from the cruciform configurationheterozygote from the cruciform configuration

Pseudolinkage –genes near breakpoints act as if linked

Fig. 13.18 c

Semisterility Semisterility results from results from translocation translocation heterozygotesheterozygotes < 50% of gametes < 50% of gametes

arise from arise from alternate alternate segregation and segregation and are viableare viable

Fig. 13.18 d

Translocation Down syndromeTranslocation Down syndrometranslocation of chromosome 21 is small and thus produces translocation of chromosome 21 is small and thus produces

viable gamete, but with phenotypic consequenceviable gamete, but with phenotypic consequence

Fig. 13.19

Transposable elements move from Transposable elements move from place to place in the genomeplace to place in the genome

Any segment of DNA that evolves ability to move Any segment of DNA that evolves ability to move from one place to another in genomefrom one place to another in genome

Selfish DNA carrying only information to self-Selfish DNA carrying only information to self-perpetuateperpetuate

Most are 50 – 10,000 bp in lengthMost are 50 – 10,000 bp in length Present hundreds of thousands of times in a Present hundreds of thousands of times in a

genomegenome ~ 7% of human genome are transposable ~ 7% of human genome are transposable

elementselements

Retroposons generate an RNA that encodes a Retroposons generate an RNA that encodes a reverse transciptase-like enzymereverse transciptase-like enzyme

Two typesTwo types Poly-A tail at 3’ Poly-A tail at 3’

end of RNA-like end of RNA-like DNA strandDNA strand

Long terminal Long terminal repeat (LTRs) repeat (LTRs) oriented in same oriented in same direction on either direction on either end of elementend of element

Fig. 13.23 a

Fig. 13.23 b

The process of LTR transpositionThe process of LTR transposition

Fig. 13.23

LINEs and SINEs in humansLINEs and SINEs in humans LINEs- LINEs- LLong ong ININterspersed terspersed EElementslements

Likely source of retrovirusesLikely source of retroviruses L1 family in humans, 6-7 kb in lengthL1 family in humans, 6-7 kb in length Encode reverse transcriptase-like enzymeEncode reverse transcriptase-like enzyme >20,000 copies in human genome>20,000 copies in human genome

SINEs-SINEs-SShort hort ININterspersed terspersed EElementslements appear to have evolved from cellular RNA species, appear to have evolved from cellular RNA species,

usually tRNAs usually tRNAs Depend on availability of reverse transcriptase Depend on availability of reverse transcriptase

produced elsewhereproduced elsewhere Alu family in humans, 300 bp in lengthAlu family in humans, 300 bp in length >500,000 copies in human genome>500,000 copies in human genome

Creation of LINE and SINE familiesCreation of LINE and SINE families

Fig. 21.18

Transposons encode transposase enzymes that Transposons encode transposase enzymes that catalyze events of transpositioncatalyze events of transposition

Fig. 13.24 a

TEs can generate chromosomal rearrangements and TEs can generate chromosomal rearrangements and relocate genesrelocate genes

Fig. 13.26

TEs can generate mutations in adjacent genesTEs can generate mutations in adjacent genesspontaneous mutations in white gene of Drosophilaspontaneous mutations in white gene of Drosophila

Fig. 13.25

Genomes often contain defective Genomes often contain defective copies of transposable elementscopies of transposable elements

Many TEs sustain deletions during Many TEs sustain deletions during transposition or repairtransposition or repair

If promoter needed for transcription is If promoter needed for transcription is deleted, TE can not transpose againdeleted, TE can not transpose again

Nonautonomous elements – need activity of Nonautonomous elements – need activity of intact copies of same TE for movementintact copies of same TE for movement

Autonomous elements – move by themselvesAutonomous elements – move by themselves Most SINEs and LINEs in human genome Most SINEs and LINEs in human genome

are defectiveare defective

P elements in DrosophilaP elements in Drosophila M strains of M strains of DrosophilaDrosophila have no P elements (most have no P elements (most

lab strains)lab strains) P strains have many copies of P elementsP strains have many copies of P elements Hybrid dysgenesis – defects including sterility, Hybrid dysgenesis – defects including sterility,

mutation, and chromosomal breakage from mutation, and chromosomal breakage from crosses between P males and M femalescrosses between P males and M females Promotes movement of P elements to new positionsPromotes movement of P elements to new positions

P-element transposons are critical P-element transposons are critical tools in molecular geneticstools in molecular genetics

Hybrid dysgenesisHybrid dysgenesis Males from Drosophila strains carrying P elements crossed to Males from Drosophila strains carrying P elements crossed to

females that lack P elementsfemales that lack P elements P element becomes highly mobile in germ line of F1 hybridsP element becomes highly mobile in germ line of F1 hybrids Chromosome breakage reduces fertility in hybridsChromosome breakage reduces fertility in hybrids Progeny of F1 flies carry many new mutations induced by P Progeny of F1 flies carry many new mutations induced by P

element insertionelement insertion Eggs produced by P female have repressor protein that Eggs produced by P female have repressor protein that

prevents transpositionprevents transposition Repressor coded for by alternatively spliced P element mRNARepressor coded for by alternatively spliced P element mRNA

Fig. D.7

Transformation: the introduction of Transformation: the introduction of cloned DNA into fliescloned DNA into flies

P-elements used as P-elements used as vectorsvectors

Insert fly DNA into Insert fly DNA into intact P element and intact P element and then into plasmidthen into plasmid

Inject into embryos Inject into embryos from M strain mothersfrom M strain mothers

Cross to P malesCross to P males

Fig. D.8a

Figure D.9

Transformation in plants by T-DNATransformation in plants by T-DNA

Bacterium Bacterium Agrobacterium Agrobacterium tumefacienstumefaciens is agent of is agent of transformationtransformation Transfer of T-DNA into Transfer of T-DNA into

genome of wounded plantgenome of wounded plant Antibiotic resistance Antibiotic resistance

markers engineered into markers engineered into plasmids provide selectionplasmids provide selection

Cells expressing the GUS Cells expressing the GUS reporter gene stain bluereporter gene stain blue

Fig. B.10 a

Transformation by Transposon Transformation by Transposon Tagging Tagging

Transposon tagging using transposable Transposon tagging using transposable elements from Corn and elements from Corn and ArabidobsisArabidobsis

Insertional mutagenesis allows generation Insertional mutagenesis allows generation of mutants of mutants

Transposon or T-DNA sequence can be Transposon or T-DNA sequence can be used to identify and clone gene of interestused to identify and clone gene of interest

PCR can be PCR can be used to find used to find

plants plants carrying a carrying a

mutated gene mutated gene of interestof interest

Fig. B.11 a-c

Fig. B.11 d,e

Fig. C.10

PCR can be used to find worms PCR can be used to find worms carrying a mutated gene of interestcarrying a mutated gene of interest