DNA/RNA Structure & DNA Replication. Genes are on Chromosomes Thomas Hunt Morgan Thomas Hunt Morgan –Worked with Drosophila –Demonstrated that genes are

DNA/RNA Structure & DNA/RNA Structure & DNA ReplicationDNA Replication

Genes are on ChromosomesGenes are on Chromosomes

Thomas Hunt MorganThomas Hunt Morgan– Worked with Worked with DrosophilaDrosophila– Demonstrated that genes are located on Demonstrated that genes are located on

chromosomeschromosomes– Is protein or DNA the genetic material Is protein or DNA the genetic material

found on the chromosomes?found on the chromosomes?– Many leading scientists through the Many leading scientists through the

1940’s thought that protein was the 1940’s thought that protein was the genetic material!genetic material!

Frederick GriffithFrederick Griffith

Streptococcus pneumoniae Streptococcus pneumoniae bacteriabacteria

Worked with 2 strains of the bacteria: R & S Worked with 2 strains of the bacteria: R & S

Harmless live bacteria mixed with heat-killed Harmless live bacteria mixed with heat-killed infectious bacteria caused disease in miceinfectious bacteria caused disease in mice

The substance passed from the dead The substance passed from the dead bacteria to the live bacteria bacteria to the live bacteria “transforming “transforming factor”factor”

Griffith & the “Transforming Griffith & the “Transforming Factor”Factor”

Transformation: change in genotype Transformation: change in genotype and phenotype due to the and phenotype due to the assimilation of external DNA by the assimilation of external DNA by the cellcell

Avery, McCarty, & MacLeod Avery, McCarty, & MacLeod (1944)(1944)

Purified DNA & proteins from Purified DNA & proteins from Streptococcus Streptococcus pneumoniaepneumoniae

Which one, DNA or protein, will transform Which one, DNA or protein, will transform non-pathogenic bacteria?non-pathogenic bacteria?

Injected protein into bacteria Injected protein into bacteria – No effectNo effect

Injected DNA into bacteria Injected DNA into bacteria – Transformed harmless bacteria into virulent Transformed harmless bacteria into virulent

bacteriabacteria Transforming Agent was DNA!!!Transforming Agent was DNA!!!

Alfred Hershey & Martha Alfred Hershey & Martha ChaseChase

Hershey & Chase (1952)Hershey & Chase (1952)

Blender experimentBlender experiment Worked with bacteriophage Worked with bacteriophage

– Viruses that infect bacteriaViruses that infect bacteria Grew phage viruses in Grew phage viruses in

2 media 2 media radioactively radioactively

labeled with either:labeled with either:– 3535S in their proteinsS in their proteins– 3232P in their DNAP in their DNA

Infected bacteria with the labeled phagesInfected bacteria with the labeled phages

Blender Experiment ResultsBlender Experiment Results

Radioactive phage & bacteria in blenderRadioactive phage & bacteria in blender Centrifuge the mixture so bacteria falls to Centrifuge the mixture so bacteria falls to

the bottomthe bottom 3535S phageS phage

– Radioactive proteins stayed in the supernatant; Radioactive proteins stayed in the supernatant; therefore, protein did NOT enter the bacteriatherefore, protein did NOT enter the bacteria

3232P phageP phage– Radioactive DNA stayed in the pellet; Radioactive DNA stayed in the pellet;

therefore, DNA did enter bacteriatherefore, DNA did enter bacteria DNA is confirmed as the “transforming DNA is confirmed as the “transforming

factor”factor”

Edwin ChargaffEdwin Chargaff

Chargaff’s Rules: DNA compositionChargaff’s Rules: DNA composition– Varies from species to speciesVaries from species to species– Amounts of 4 nitrogen bases are not equalAmounts of 4 nitrogen bases are not equal– Bases are present in the following ratio:Bases are present in the following ratio:

A = TA = T G = CG = C

Humans Humans A = 30.9%A = 30.9% T = 29.4%T = 29.4%

G = 19.9%G = 19.9% C = 19.8%C = 19.8%

Structure of DNAStructure of DNA

James Watson James Watson

& Francis Crick& Francis Crick

– Developed the Developed the

double helix model double helix model

of DNAof DNA

Other scientists working on the Other scientists working on the structure of DNA……..structure of DNA……..

Maurice Wilkins Linus Pauling Rosalind Franklin

Prokaryotic vs. Eukaryotic Prokaryotic vs. Eukaryotic DNADNA

Prokaryotic CellsProkaryotic Cells– Single, circular chromosomeSingle, circular chromosome– Also contain plasmids (small, extra-Also contain plasmids (small, extra-

chromosomal DNA molecules); also chromosomal DNA molecules); also found in viruses found in viruses

Eukaryotic CellEukaryotic Cell– Multiple linear chromosomesMultiple linear chromosomes

•Structure of DNAStructure of DNA

Double HelixDouble Helix Nucleotides linked togetherNucleotides linked together


Nucleotide consists of……….Nucleotide consists of……….– Deoxyribose Sugar (CDeoxyribose Sugar (C55HH1010OO44))

(green)(green)

– Phosphate GroupPhosphate Group

(blue)(blue)

– Nitrogen Base: Nitrogen Base:

purines & pyrimidines purines & pyrimidines (gold)(gold)


Nitrogen Base Pairing in Nitrogen Base Pairing in DNADNA

Purines Purines two-ringed two-ringed– AdenineAdenine– GuanineGuanine

Pyrimidines Pyrimidines one-ringed one-ringed– ThymineThymine– CytosineCytosine

Base-Pairing Rules:Base-Pairing Rules:– A : TA : T– G : CG : C

Hydrogen Bond

Bonding in DNABonding in DNA

Phosphodiester Bonds: Phosphodiester Bonds:

b/w sugar & phosphate b/w sugar & phosphate Hydrogen Bonds: b/w Hydrogen Bonds: b/w

nitrogen bases in the nitrogen bases in the

middle of the moleculemiddle of the molecule Which bonds are………Which bonds are………

strong? weak? strong? weak?

Phosphodiester Bond

Anti-parallel StrandsAnti-parallel Strands

Phosphate to sugar bond Phosphate to sugar bond

(phosphodiester bond) (phosphodiester bond)

involves carbons in a involves carbons in a

3’ & 5’ position3’ & 5’ position DNA has a “direction”DNA has a “direction” Complementary strands Complementary strands

run in run in opposite opposite directiondirection

“It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material” Watson & Crick

Structure of RNAStructure of RNA

Like DNALike DNA– Has a sugar, phosphate, & nitrogen baseHas a sugar, phosphate, & nitrogen base– Made up of nucleotidesMade up of nucleotides– Follows base-paring rulesFollows base-paring rules

Unlike DNAUnlike DNA– Sugar is ribose (not deoxyribose)Sugar is ribose (not deoxyribose)– Single-stranded (not double-stranded)Single-stranded (not double-stranded)– Adenine, Guanine, Cytosine, & URACIL Adenine, Guanine, Cytosine, & URACIL

(no thymine in RNA)(no thymine in RNA)

DNA vs. RNADNA vs. RNA

Types of RNATypes of RNA

mRNA (messenger) mRNA (messenger) – carries the DNA message from the carries the DNA message from the

nucleus to the ribosomenucleus to the ribosome tRNA (transfer)tRNA (transfer)

– Carries amino acids to the ribosomeCarries amino acids to the ribosome rRNA (ribosomal)rRNA (ribosomal)

– Makes up the ribosomeMakes up the ribosome

DNA ReplicationDNA Replication

Base Pairing allowsBase Pairing allows

each strand to serve each strand to serve

as a template for a as a template for a

new strandnew strand


Models of DNA ReplicationModels of DNA Replication conservative semi-conservative dispersiveconservative semi-conservative dispersive

Semi-Conservative Semi-Conservative ReplicationReplication

Meselson & Stahl (1958)Meselson & Stahl (1958)– Labeled the nucleotides of the parent strand with an Labeled the nucleotides of the parent strand with an

isotope of heavy nitrogen isotope of heavy nitrogen 1515NN– Labeled the “new” nucleotides with a lighter isotope Labeled the “new” nucleotides with a lighter isotope

1414NN Replicated strands are half Replicated strands are half 1515NN and half and half 1414NN


Parent molecule will split at the Parent molecule will split at the hydrogen bonds and each nucleotide hydrogen bonds and each nucleotide will pair with it’s complementary will pair with it’s complementary nucleotide.nucleotide.

Process of DNA ReplicationProcess of DNA Replication

Multiple enzymes control the Multiple enzymes control the replication of DNA……….replication of DNA……….– HelicaseHelicase– Single-stranded binding proteinSingle-stranded binding protein– DNA Polymerase IIIDNA Polymerase III– PrimasePrimase– DNA ligaseDNA ligase

DNA Replication: Getting DNA Replication: Getting StartedStarted


Origin of ReplicationOrigin of Replication: recognizable : recognizable base sequence where a protein binds base sequence where a protein binds to start replicationto start replication– Bacterial DNA Bacterial DNA 1 replication origin 1 replication origin– Eukaryotic DNA Eukaryotic DNA multiple replication multiple replication

originsorigins Replication ForkReplication Fork: Y-shaped area : Y-shaped area

where the DNA strands of DNA are where the DNA strands of DNA are growing fromgrowing from


Enzymes involved in “getting Enzymes involved in “getting started”…started”…

Helicase: starts the unwinding of the Helicase: starts the unwinding of the double helix; opens the DNA helixdouble helix; opens the DNA helix

Single-Stranded Binding Protein: Single-Stranded Binding Protein: holds the separated strands apart holds the separated strands apart

DNA Replication: DNA Replication: ElongationElongation

DNA Polymerase IIIDNA Polymerase III Adds nucleotides only to the 3’ endAdds nucleotides only to the 3’ end

DNA Replication: DNA Replication: ElongationElongation

Leading & Lagging StrandsLeading & Lagging Strands– Leading StrandsLeading Strands

Continuous synthesisContinuous synthesis – Lagging StrandsLagging Strands

Okazaki fragmentsOkazaki fragments Joined by ligase Joined by ligase

acts like glue to connectacts like glue to connect

Okazaki fragmentsOkazaki fragments

DNA Replication: PrimingDNA Replication: Priming DNA polymerase can onlyDNA polymerase can only extend an existing DNA extend an existing DNA molecule—it cannot molecule—it cannot

start a new one!start a new one! RNA primerRNA primer – made on – made on parent DNA strand byparent DNA strand by primaseprimase RNA primer is laterRNA primer is later removed byremoved by DNA polymerase IDNA polymerase I

DNA Replication EnzymesDNA Replication Enzymes

DNA PolymerasesDNA Polymerases

Don’t get them confused…………..Don’t get them confused………….. DNA Polymerase IDNA Polymerase I 20 bases/second20 bases/second Editing, repair, & RNA primer removalEditing, repair, & RNA primer removal

DNA Polymerase IIIDNA Polymerase III 1000 bases/second1000 bases/second Main DNA building enzymeMain DNA building enzyme

Editing/Proofreading DNAEditing/Proofreading DNA Lots of mistakes occur!Lots of mistakes occur! DNA polymerase I DNA polymerase I

removes incorrect basesremoves incorrect bases– Reduces the error rate Reduces the error rate

from 1 in 10,000 to from 1 in 10,000 to

1 in 100 million bases1 in 100 million bases Mismatch Repair:Mismatch Repair: -enzymes correct incorrectly-enzymes correct incorrectly

paired nucleotidespaired nucleotides 130 known DNA repair 130 known DNA repair

enzymes in humansenzymes in humans

Speed of DNA ReplicationSpeed of DNA Replication

E.coliE.coli can copy it’s entire genome of 5 can copy it’s entire genome of 5 million base pairs in under 1 hour!!!million base pairs in under 1 hour!!!

A human cell can copy its 6 billion A human cell can copy its 6 billion bases & divide into 2 cells in a few bases & divide into 2 cells in a few hourshours– Incredibly accurateIncredibly accurate– About 30 errors per cell cycleAbout 30 errors per cell cycle

The End-Replication The End-Replication ProblemProblem

Ends of the Ends of the chromosomes are chromosomes are destroyed with eachdestroyed with each replicationreplication No way to complete No way to complete

the 5’ends of the 5’ends of daughter DNA daughter DNA strandsstrands

What are telomeres?What are telomeres? Expendable non-coding Expendable non-coding

segments of DNA at the segments of DNA at the

ends of chromosomesends of chromosomes

– Short sequence of basesShort sequence of bases

repeated thousands of timerepeated thousands of time– TTAGGG in humansTTAGGG in humans– Prevent erosion of genesPrevent erosion of genes

TelomeraseTelomerase - enzyme in - enzyme in

certain cells that starts certain cells that starts

the lengthening of telomeres; only in germ-line cells!the lengthening of telomeres; only in germ-line cells!

Changes in Chromosome Changes in Chromosome StructureStructure

Documents

DNA/RNA Structure & DNA Replication. Genes are on Chromosomes Thomas Hunt Morgan Thomas Hunt Morgan –Worked with Drosophila –Demonstrated that genes are