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Welcome to Genetics (BIOL 364/564)

Dr. Carol Ely Hepfer

Textbook and Study Guide:

iGenetics: A Molecular Approach, third edition

Author: Peter J. Russel

Publisher: Pearson/Benjamin Cummings

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Materials Available at Desire 2 Learn (D2L)

In ‘Content’ Area

Syllabus, Handouts, PowerPoint slides, Pencasts

Discussions - Group Lockers, Drop Boxes

Announcements - Marauder email essential

PLEASE - LOG OUT!!!

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Handouts

Syllabus

Lecture Slides (available at D2L, not comprehensive)

Lab Handouts

*Alternative Transformation Protocol*

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What is Genetics?

Biological Study of Inheritance

Inherited traits are determined

by genes that are transmitted

from parents to offspring during

sexual reproduction.

Chi-Chon

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Areas of Study within Genetics

Population Genetics, Quantitative Genetics

Molecular Biology

Cytogenetics

TransmissionGenetics

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History of Genetics

~~ Agriculture, Breeding

1859 Charles Darwin - ‘Origin of the Species’, evolution, natural selection

1866 Gregor Mendel - Research published on inheritance in peas

1869 Friedrich Miescher - Discovered nucleic acids

1900 Hugo deVries, Carl Correns, Erich vonTschermak -Rediscovered Mendel

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History of Genetics

1908 Archibald Garrod - ‘Inborn Errors of Metabolism’,

defective enzymes cause human diseases

1928 Frederick Griffith - Phenotype transformation observed in bacteria

1944 Oswald Avery, Colin MacLeod, MacLyn McCarty - DNA is the genetic material in bacteria

1952 Alfred Hershey, Martha Chase - DNA is the genetic material in some viruses

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History of Genetics

1950s Rosalind Franklin, Maurice Wilkins - Xray crystallography of DNA

1953 James Watson, Francis Crick - Deduced three dimensional structure of DNA

1970s Numerous investigators - Deciphered the genetic code Molecular Biology begins

1990s Human genome project initiated - Genomics, Proteomics, etc.,

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Gene expression determines phenotype

Phenotype - outward appearance of an individual

Examples

Plant Height - Tall, Dwarf

Hair Texture - Curly, Wavy Straight

Yeast Metabolism - Produces ASN, Requires ASN

Bacterial Resistance - Tetracycline, Ampicillin

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Classification of Phenotypes

Wild type (+) = most common phenotype in population, ‘normal’

Mutant = unusual, variant form,

due to changes (mutations) in gene(s)

Examples:

Trait Wild type Mutant

Eye color Red White,

scarlet, brown

Coloration Pigmented Albino

Whippets

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What is a gene?

Mendelian

Cytogenetic

Molecular

Abstract unit of inheritance - A a B b

Location along a chromosome gene locus

Sequences of bases in DNA

5’ ATCGCTGTCAGTCCTAGA 3’ OR

5’ ATCGCTCTCAGTCCTAGA 3’

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Discovery that DNA is the genetic material

Genes - control phenotype, located on chromosomes

Chromosomes - composed of DNA, RNA, protein

1940’s -

Proteins favored as candidate - sufficiently complex

Nucleic acids - too simple

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Preliminary Evidence in favor of DNA

1. Genes and chromosomes - in nucleus, not cytoplasm

Proteins and RNA - in nucleus and cytoplasm

DNA - mostly in

nucleus

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Preliminary Evidence in favor of DNA

2. [DNA] # of chromatids

G1 cells - 1 X [DNA] (unreplicated chromosomes)

G2 cells -2X [DNA]

(replicated chromosomes)

[DNA] in diploid cells = 2x that in haploid cells

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Preliminary Evidence in favor of DNA

3. Chemical composition of DNA

Consistent within each species

Same in all cells of an individual

RNA and Proteins differ substantially

in different cells of the same

individual

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Preliminary Evidence in favor of DNA

4. UV light (260 nm)

Wavelength of maximum absorbance for

DNA

Highly mutagenic

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Griffith’s Transformation Experiment (1928)

Diplococcus pneumoniae

Antigen type (II or III)

Colony shape

Smooth (S) - encapsulated, virulent

Rough (R ) - no capsule, avirulent

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Griffith’s Transformation Experiment

What is going on?

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Interpreting Griffith’s Transformation Experiment

Could mutations in IIR cells transform them into IIIS?

Something from dead IIIS cells ‘transforms’ IIR into IIIS

Probability of one mutation~ 1 in 1,000,000

Probability of two mutations< 1 in 1012

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What is Griffith’s Transforming Principle?

Avery, MacLeod, McCarty (1944)

Extract from virulent (S) bacteria isolated

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Avery, Macleod, McCarty Experiment

Extract from III S

protease

Is it DNA or RNA?

RNase

DNase

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Hershey & Chase Experiment

Experimental organism - T2 bacteriophage

- composed of protein and DNA

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Hershey & Chase Experiment

Life cycle of T2 bacteriophage

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Hershey & Chase Experiment

Phage inject genetic information into bacteria

Is it their DNA or their protein?

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Hershey & Chase Experiment

Allow infection, separate bacteria from phage

Centrifuge

Centrifuge

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Identity of Genetic Material Established

Most organisms - DNA

Some viruses - RNA

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Chemical Composition and Structure

DNA and RNA - polymers of nucleotides

Each nucleotide - sugar, base, phosphate

Different functional group at carbon #2

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Chemical Composition and Structure

Nitrogenous Bases

Purines

Pyrimidines

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Chemical Composition and Structure

DNA

Nucleosides:

deoxyadenosine

deoxyguanosine

deoxycytidine

deoxythymidine

Nucleotides:

-monophosphate

dAMP, etc.

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Chemical Composition and Structure

RNA

Nucleotides

adenosine

monophosphate,

uridine

monophosphate,

etc.

AMP, UMP, etc.

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Chemical Composition and Structure

DNA and RNA

polymers of

nucleotides

3’-5’ phosphodiester

linkage

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RNA is normally single-stranded

Secondary structures - intrastrand H-bonding

Ex. Stem and Loop

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Evidence that DNA is double-stranded

Chemical composition

Chargaff (1947)

[C] =

[G]

[A] = [T]

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Evidence that DNA is helical

Physical chemistry - Franklin & Wilkins (1952)

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Evidence that DNA is helical

O

Xray diffraction

helical structure

20 A diameter

highly ordered

repeating units

3.4 A

34 A

Note: 1 A = 0.1 nm

O

O

O

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Watson and Crick

Model building to deduce structure (1953)

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Watson and Crick Model for DNA

DNA B

right-handed

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Specific Base Pairing in DNA

Hydrogen Bonding

A with T

C with G

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Arrangement of DNA strands

Antiparallel

Complementary

5’ TGTA 3’

3’ ACAT 5’

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Different Forms of DNA

A-DNA B-DNA Z-DNA

right-handedtight coiling

left-handedloose coiling

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Organization of Chromosomes - Viruses

dsDNA, ssDNA, dsRNA, ssRNA

circular, linear

single chromosome, segmented genome

T-even phages (T2, T4, T6) - dsDNA, 1 linear chromosome

X174 - ssDNA, one circular chromosome

Lambda () - dsDNA, alternates linear and circular

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Organization of Chromosomes - Prokaryotes

Most - single circle, dsDNA

Some - linear

Extra chromosomes, Plasmids

Cells Divide by Binary Fission

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Organization of Chromosomes - Prokaryotes

A. tumefaciens - 1 circular + 1 linear chromosome (3.0 Mb) (2.1 Mb)

E. coli - 1 circular chromosome + circular plasmids (4.6 Mb)

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Organization of Chromosomes - Prokaryotes

DNA supercoiled into nucleoid region of cell

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Organization of Chromosomes - Eukaryotes

Replicated metaphase chromosome

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Organization of Chromosomes - Eukaryotes

DNA supercoiled to fit into chromosomes

Relaxed DNA in haploid (1n) human cell = 1 meter long

DNA in largest chromosome = 82 mm long

Metaphase chromosome = 10 m long

Analogy: 25 miles of rope coiled into 2 ft x 16 ft canoe

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Organization of Chromosomes - Eukaryotes

Chromatin - DNA supercoiled with proteins

Histones - small, basic (+), bind DNA (-)

H1, H2A, H2B, H3, H4

Nonhistones - all other associated proteins

- many acidic (+), bind histones

- include those for repair, replication, etc.

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Nucleosome Structure

DNA (147 bp) wrapped around histone octamer

10 nm chromatin fiber - string of nucleosomes

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Chromatin Structure

10 nm chromatin fiber - condensed with H1

30 nm chromatin fiber - solenoid model

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Chromatin in Metaphase Chromosome

Metaphase chromosome with histones removed

Nonhistone scaffold

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Euchromatin and Heterochromatin

Euchromatin - staining intensity varies with cell cycle as chromatin condenses and

relaxes

actively transcribed

no repetitive sequences

Heterochromatin - remains condensed (darkly staining)

often replicates later

transcriptionally inactive

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Types of Heterochromatin

Constitutive - present in all cells at same location

usually repetitive DNA

Ex. centromere region

Facultative - varies with cell type, stage, location

Ex. inactive X chromosome

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Centromeric DNA

Repetitive sequences,

Constriction where sister chromatids are joined,

Important for chromosome segregation

Vary in size and sequence

Bind proteins, site of kinetochore formation

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Telomeric DNA

Ends of chromosomes, tandem repeats

Stability and replication of chromosome ends

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Unique Sequence DNA

One copy per haploid cell

Most protein encoding genes

Other sequences also unique

~ 55-60 % of human genome

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Repetitive Sequence DNA

Numerous copies in genome

Dispersed Repeats (INterspersed Elements)

LINEs - 1,000 to > 7,000 bp

LINE1 (~500,000 copies in mammals)

SINEs - 100 to 400 bp

AluI (one per 5,000 bp of human genome)

Tandem Repeats

Telomeric sequences - 1 - 10 bp long

rRNA and tRNA genes -