1 Welcome to Genetics (BIOL 364/564) Dr. Carol Ely Hepfer Textbook and Study Guide: iGenetics: A...

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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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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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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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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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Life cycle of T2 bacteriophage

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Phage inject genetic information into bacteria

Is it their DNA or their protein?

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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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Nitrogenous Bases

Purines

Pyrimidines

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DNA

Nucleosides:

deoxyadenosine

deoxyguanosine

deoxycytidine

deoxythymidine

Nucleotides:

-monophosphate

dAMP, etc.

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RNA

Nucleotides

adenosine

monophosphate,

uridine

monophosphate,

etc.

AMP, UMP, etc.

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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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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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DNA supercoiled into nucleoid region of cell

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Organization of Chromosomes - Eukaryotes

Replicated metaphase chromosome

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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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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 -

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