DNA Chapter 2 – read principal points

Timeline for genetics

Deoxyribonucleic acid

Functional Properties1. Replication – DNA is

copied prior to cell division why?

2. Storage of information DNA inherited

from parent to offspring from cell to cell

Gene expression – Genes encode proteins

3. Mutation – DNA changes to allow variation and adaptation, the basis of evolution

A six-legged green frog. (Reproduced by permission of JLM Visuals http://www.isogenic.info/assets/images/autogen/a_image16.jpg

http://3quarksdaily.blogs.com/3quarksdaily/images/wolfe_seal_1.jpg

Neutral, harmful, adaptive?

DNA History

1869 Meischer extracted nuclein from pus

1900s – chromosomes discovered

The genetic material must have the 3 functional properties

microscopy.bio.cmich.edu

Griffith finds “transforming factor”1928 London

Streptococcus pneumoniae bacterium

pneumonia in mice, deadly to humans

sputum with bacteria

Smooth strain (IIIS) virulent polysaccharide capsule capsule allows bacteria to evade immune

system

Fluorescent stain of capsule

isolate live IIISfrom mouse

S pneumococcus kills mouse in 24 hours. But 100 million IIR strain bacterial cells is harmless

S R

(Research photographs of Dr. Harriet Ephrussi-Taylor, courtesy of The Rockefeller University.)

Appearance when grown on an agar plate

http://biology.kenyon.edu/courses/biol114/KH_lecture_images/How_DNA_works/how_DNA-works.html

Rough strain IIR is avirulent

IIR lacks capsule

-> isolate live IIR

The experiment:

Heat kill IIIS strain mouse ?

Heat killed IIIS strain + live strain IIR mouse ?

Which strain is isolated?

Griffith’s experiment and conclusion

A “transforming factor” in killed S strain transformed live R strain into S

DNA or protein?

1944 Avery, McCarty, Macleod

1. Heat kill IIIS

2. Remove lipids and sugars – how?

FYI iGenetics: DNA as Genetic Material: Avery’s Transformation Experiment

3. Divide into 3 and treat with: proteaseRNaseDNase

next, add live R cells to each

1952 Hershey and Chase

Used T2 bacteriophage + E. coli

A phage is a virus that infects bacteria

How phage work

1. phage adsorbs onto bacterial surface

2. Genetic material injected

3. Cell makes progeny phage

IS the genetic material DNA, or protein?

Experiment

1. Label phage protein with 35 S infect E. coli strip phage off cell surface

New phage are not radioactive

2. Label phage DNA with 32 P -> infect E.coli -> blend ->

New phage contain 32 P

http://osulibrary.orst.edu/specialcollections/coll/pauling/dna/pictures/hersheychase-experiment.html

Hershey and Chase conclusion

DNA is responsible for function and reproduction of phage virus = the genetic material

Structure of DNA = nucleotide polymers

NUCLEOTIDES

1. Nitrogenous base

Purines = guanine and adenine

G A

Purines attached to 1 carbon of sugar at 9 nitrogen, covalent bond, pyrimidines attached to 1 carbon at 1 nitrogen

How big IS a nucleotide? UTAH cell scale

PyrimidinesThymine TCytosine CRNA contains uracil U

2. Deoxyribose sugar

RNA (ribose) 2’ OH makes RNA less stable than DNA Sugar + base = nucleoside

3. Phosphate (PO4)

Nucleotide = base + sugar + phosphate

Phosphate covalently (phosphodietster bond) attached to 5’C of sugar Phosphodiester bond - Covalent bond between phosphate of one nucleotide and 3’ sugar carbon of another9 N (purine) or 6N (pyrimidine) covalently bonded to 1C of sugar

DNA is a polymer of nucleotides

polarity 5’carbon to 3’hydroxyl

DNA (double helix)Watson and Crick 1953

X-ray diffraction data RosalindFranklin, Maurice Wilkins

GETncm/justsaycust-recrate-itemcommunittg/stores/dtg/stores/d-favorite-listruejust-say-no

DNA properties include:

Complementary base pairing

1. Hydrogen bonds between complementary bases

How many bonds in a G-C pair?

A-T?

Which is stronger?

Complementary base pairs

Which are the G-Cpairs?

2. antiparallel stands5’ 3’ and 3’ 5’

3. Sugar phosphate backbones

4. Base composition

DNA 50% purine 50% pyrimidine

A = T G = C

A/T = 1 C/G = 1

A +T does not equal C+G

A + G = C + TChargaff

(1950)

5. DNA can denature and renature

Melt hydrogen bonds (chemical or heat)

And

5. Right handed helix6. Complete turn of the helix is 0.34 nm,

10 bases per turn7. Major and minor grooves

Major and minor grooves

Forms of DNA

B DNA right helix

10 bp/ turn

A DNA right helix

10.9 bp/ turn

Z DNA left helix

12 bp/turn (role?)

Cellular DNA closest to B DNA10.4 bp/turn

Replication of DNA by Complementary Base Pairing

HHMI interactive DNA replication advanced

Organization of DNA in chromosomes Genome

Full amount of genetic material in a single cell

Influenza ssRNA HIV ssRNA

Bacteriophage

ds DNA

Viral chromosomeSingle or double stranded DNA or RNACircular or linear

Herpes ds DNA

Parvovirus ssDNA

Genetic material in prokaryotes

1 (usually) chromosome Circular (most) chromosome Supercoiled DNA located in nucleoid region

Neisseria gonorrhoeae

E. coli = 4.6 million bp, circular chromosome

1500 um genome stuffed into a 1 um cell via supercoiling

E. Coli cells E. coli DNA map of chromosome

Some bacteria contain extra-chromosomal DNA called a plasmid

Eukaryotic Chromosomes

C value - Amount of DNA varies among species

The structure of chromatin

DNA + proteins Highly conserved

Histones and non-histones

Histone proteins basic

net + charge interacts with – charged DNA

Package DNA

Highly conserved

Non-histone proteinsvary among species

http://faculty.jsd.claremont.edu/jarmstrong/images/chromatin.gif

What do histones do?

1. pack DNA into chromatinCondense DNA 10,000X (2 meters nanometers)

1. Modifications to histone proteins affect gene expression

5 histone proteins

  Histone type #amino acidscontent_________________

H1  200-265 27%lysine, 2%arginine

H2A   129-155  11% lysine, 9% arginine  H2B  121-155  6% lysine, 6% arginine 

H3 135  10%lysine, 15% arginine  H4  102  11% lysine, 4% arginine 

Note: all are lysine/arginine rich, they contain other amino acids, but at small percentages.Basic, + charge

Gel electrophoresis

epigenetics Modification of histone proteins changes

gene expression. Chemical tags can be inherited and are

stable in cell division

Pbs: A Tale of Two MiceEpigenetics with deGrassi http://www.pbs.org/wgbh/nova/sciencenow/3411/02.html

Agouti Mice

Epigenetics and gene silencing

Mouse and twin studies

Diet and the epigenome

Methyl group tags

Chromosome packaging

1. Nucleosomes 1 nucleosome

“beads on a string” The 10 um chromatin fiber 2(H2A).2(H2B) 2(H3).2(H4) octomer

Dual role of nucleosomes

stable to shelter DNA and compact it labile to allow DNA information to be used

2. The 30 nm chromatin fiber

Histone H1 attaches linker DNA to nucleosomes

The 30 nm chromatin fiber

(11nm string-> 30 nm helical fiber)

See Science article

3. Further packaging into loops and scaffolds……

Summary of DNA packaging

30nm chromatin fiber

Nucleosomes

DNA helix

Metaphase chromosome is 10,000 X condensed compared to double helix

Condensed scaffold

The scaffold with loops

Euchromatin and Heterochromatin

1. Heterochromatin Condensed Transcriptionally inactive Ex. centromeres

Heterochromatin stains darkly

Heterchromatin example

Barr body (facultative, extent of inactivity varies)

Inactivated X chromosome in females

2. Euchromatin

Lighter staining regions of DNA that contain genes

Transcriptionally active

Unique Sequences and Repetitive DNA

What genes are on a chromosome?

http://www.dnalc.org/ddnalc/resources/chr11.html

Chromosome 11 flyover

Terms:

Transposon

Pseudogene

Olfactory

Polymorphism

1. Unique Sequence DNA

(1 to a few copies)

a.Genes Encode proteins

~60% of DNA

Only 2% of DNA is coding (H. sapiens)

Estimated 20,000 genes in humans

b. Gene families

encodes embryonic beta globin encode fetal beta globin is a pseudogene (not functional) encodes normal beta globin encodes normal adult beta globin

Example: Beta globin (encode subunits of hemoglobin)

2. Repetitive DNA

Repeated 10 – 1000sX in the genome

a. Dispersed repeated DNA

LINES = long interspersed elements

1000 – 7000 bp

Ex. humans have 500,000 copies of L1 = 15 % of genome.

some are transposons= copy and move

SINES

100 – 400 bp Ex. Alu repeats repeated 1 million times = 10% of genome

b. Tandem repeats

1 – 10 bp long

tandemly repeated

Centromeres, telomeres, rRNA genes

Ex. telomere sequence repeated 2000X

5'...TTAGGG TTAGGG TTAGGG TTAGGG TTAGGG TTAGGG..3' 3'...AATCCC AATCCC AATCCC AATCCC AATCCC AATCCC..5'

Ex. rRNA genes