Animal Science 3213

Genetics of Agricultural Plants and Animals

Dr. Prather

Week 1

For Jan. 12 READ Chapter 10

For Jan. 13 READ Chapeter 11 pg 327-341

For Jan. 14 Read Chapter 12 & pg 47-66

Bring your colored pencils

For Jan. 15 READ pg 77-84

For Jan. 16 READ Chapters 13 & 14

Chapter 10

The Structure of Genetic Material

1) Discovery of DNA

-Heritable Traits
Stability
Accuracy of Replication
Capacity for Variation

2) Chemical Composition and Structure of DNA & RNA

-Nucleotides
Pentose Sugar (Fig 10.9)
*deoxyribose DNA (hydrogen attached to the 2' carbon)
*ribose RNA (hydroxyl group
attached to the 2' carbon)
Nitrogenous Base (Fig 10.10)
*Purines
Adenine (A)
Guanine (G)
*Pyrimidines
Cytosine (C)
Thymidine (T) ONLY in DNA
Uracil (U) ONLY in RNA

Base linked to sugar by 1’ carbon of pentose sugar (Fig 10.11)

dGMP dAMP dCMP cTMP = DNA

GMP AMP CMP UMP = RNA

-Polynucleotides- (Fig 10.11)
Linked by covalent bonds between a phosphate group of one nucleotide and the 3' carbon of the pentose sugar of the next nucleotide (a 5' - 3' linkage).

The bonds between the nucleotides are phosphodiester bonds.

The two ends of the polynucleotide chain are not the same. The 5' end has a phosphate group attached to the 5' carbon of the pentose sugar. The 3' end has a hydroxyl group (OH) attached to the sugar's 3' carbon. Thus the chain has polarity.

-Pairing of Complementary Bases (Fig 10.15)
Hydrogen Bonding
AT
CG
AU (RNA)

3) Forms of DNA & RNA

-Watson & Crick put the puzzle together.

-The polynucleotide strands are antiparallel and hydrogen bonds form between complementary base pairs (A and T; C and G). The strands are twisted in a helical configuration, with 10 base pairs between each turn of the helix (Fig 10.14, 10.16)

-Mainly the B form of DNA is found in the cell.

Species A G T C U

i 21 29 21 29 0

ii 29 21 29 21 0

iii 21 29 0 29 21

iv 21 21 29 29 0

v 21 29 0 21 29

Which species has RNA? DNA?
Which is double stranded? Single stranded?

Take Home Messages & You Had Better Know

-Genetic material must contain all the information for the cell structure and function of an organism.
-Genetic material must also replicate accurately so the progeny cells have the same genetic information as the parent cell.
-Genetic material must be capable of variation.
-Know the structure of A, T, C, G & U.
-Know the difference between DNA and RNA.

Chapter 11

Organization of DNA into Chromosomes

1) Bacteria DNA Packaging
-One circular supercoiled chromosome. (Fig 11.1) Circular double stranded DNA molecule that is compacted by supercoiling
-Same principle in eukaryotic chromosomes. (Positive and Negative supercoiling controlled by Topoisomerases)
-Some proteins similar to histones are in bacterial cells.

2) Bacteriophages
-Genetic material may be single- or double-stranded RNA or DNA, linear or circular.
-Chromosome consists of Protein and DNA

3) Structure of Eukaryotic Chromosomes
-Karyotype-Largest is usually labeled #1. Pairs of chromosomes; 1 from each parent. (Fig 11.11 & 13).
Metacentric
Submetacentric
Acrocentric
Telocentric

-G and Q banding identify the same regions of chromosomes (A and T rich regions)

-Fluorescence in situ hybridization (FISH), identifies specific regions of a chromosome, e.g. A specific probe for growth hormone will bind the gene on the chromosome #1 in cattle. Binding is mediated after removing the protein and allowing the base pairs to match-up.

-Amount of DNA
C value is the haploid amount of DNA for any given species, i.e. the amount of DNA in a sperm.
Humans 94 cm, ~3 feet.
Humans have ~2.7X10⁷ base pairs

4) Chromatin = Protein + DNA

-Euchromatin
Stains lightly, uncoiled during interphase, but condensed during mitosis.

-Heterochromatin
Stains darkly, more condensed, genetically inactive.
Found near centromeres, telomeres and species-specific locations.
*Constitutive heterochromatin, centromeres & telomeres.
*Facultative heterochromatin, has the potential to become condensed, e.g. X chromosome inactivation.

-Proteins
Histones- Sequences are very similar in all species.
*Very rich in arginine and lysine
*Have a net positive charge, thus bind to negatively charged DNA.
*Histones associate to form nucleosomes, Nucleosomes fold into chromatin fibers.
These chromatin fibers are attached to a protein scaffold (Fig 11.23).
*Nucleosome
Two copies of H2B, H4, H3, H2A thus an octamer.
H1 is a linker between nueleosomes.
Non-histones
*Scaffolds, DNA- and RNA polymerases, Topoisomerases, etc...
Chromosomes are composed of DNA, histone and non-histone chromosomal proteins.
Each chromosome consists of one linear, unbroken, double stranded DNA molecule, with coils and folds.
Histones are fairly constant from cell to cell, but the non-histone protein vary considerably.

5) Centromeres & Telomeres

-Centromeres
Sites at which chromosomes attach to the mitotic and meiotic spindle (Fig. 11.25).
Responsible for accurate segregation of the replicated chromosome to the progeny cells during meiosis and mitosis.
Without a centromere the chromosomes will randomly segregate.
Non-disjunction results from mistakes in segregation during meiosis (Fig 3.27).
Generally seen as a constriction on the chromosome where unique sequences are located_.
-Telomeres
Located at the ends of the chromosomes.
Often associated with the nuclear envelope.
Short tandemly repeated sequences and other repeated sequences further in from the ends.
required for replication and stability of the chromosome.
Heterochromatic
They are not 'sticky', so individual chromosomes do not stick together.

Take Home Messages & You Had Better Know

What is a Karyotype.
What is Banding.
Describe the amount of DNA in an organism.
What is Euchromatin versus Heterochromatin.
How do Histones package DNA.
What is the function of Centromeres & Telomeres.
Chromosome Scaffold
Topoisomerase II induces negative supercoils
Matrix Attachment Regions (MARs)
Active Genes

Chapter 12 & pg 47-66

DNA Replication, Mitosis & Meiosis

1) DNA replication occurs by a semiconservative mechanism (Meselson-Stahl Exp. (Fig 12.1)

2) Enzymes involved in DNA synthesis.
-DNA polymerase I takes dNTPs (dATP, dCTP, dTTP, dGTP), Mg²+ ions, a fragment of DNA and makes a copy, by catalyzing the formation of phosphodiester bonds between the 3'-OH group of the deoxyribose on the last nucleotide and the incoming 5'-phosphate of the dNTP precursor. Each incoming dNTP is selected by the DNA polymerase I according to the order of bases on the template strand (Fig 12.3). First dNTP binding acts as a primer to get the reaction started. Synthesizes 5' to 3'.

-DNA polymerase II

-DNA polymerase III
3 subunits constitute the catalytic core (alpha, episilon, theta [a , e , f ]).

Polymerases also have 3'-5' exonuclease activity to correct errors in polymerization (Tbl 12.2).
-Telomerase to replicate telomeres.

3) DNA replication in Eukaryotes

-Occurs in the nucleus during S phase of the cell cycle.
-Is initiated by RNA primers, occurs in the 5' to 3' direction, is catalyzed by DNA polymerases,
requires a large number of other enzymes and proteins (Fig 12.6, 12.8, 12.9, 12.12).
-Is semiconservative and continuous, discontinuous, and bidirectional. Replication is
initiated at the same time at many points (origins of replication) along the chromosome (Fig 12.18).
-Heterochromatin replicates later in S than does euchromatin.
-In Eukaryotes DNA polymerases are classified as in Tbl 12.4.

4) Packaging of newly replicated DNA.

-Histone synthesis is coordinated with DNA synthesis. Note that these histones must disassemble for
DNA synthesis to occur. Segregation of the histones is not clear.

Take Home and You Had Better Know

*DNA replication is semi-conservative.
*A large number of enzymes are involved in DNA replication, and each has a different function; primase, polymerase, ligase, etc..
*Histones must disassemble and then reassemble during replication.

5) Cell Cycle

-Chromosome Cycle

Interphase-
*G₁, In G₁ phase, the morphology of the chromosomes changes from condensed to a dispersed state.
This is a result of a change in the coiling of the fibers. The cell also prepares for S by producing RNA and protein.
*G₀, Is a quiescent phase of neither growing or progressing to S. It is usually induced by starvation of some nutrient.
*During G₁ there is a period called the 'restriction point'. Once the cell passes this point
(completes certain biochemical events), then it must enter S phase.
*S, The untwisting of the DNA and replication as previously described.
*G₂, This phase is generally short, the DNA begins to condense

Mitosis-

Chromosomes are characterized as highly condensed and segregate to the two daughter cells (Fig 3.5, 3.7).

*Prophase
Sister chromatids become detectable.
Mitotic spindle assembles outside the nucleus.
Microtubules (tubulin) and a centrosome (in animals centrioles are present).
Kinetochores form on the centromere and the microtubules attach.
Nuclear envelope breakdown begins.

*Metaphase
Begins when the nuclear envelop is completely broken down.
Centromeres become aligned.

*Anaphase
Sister chromatids separate at the centromere (disjunction). Pac-Man model.

*Telophase
Migration is complete and chromosomes begin to disassemble.
The nuclear envelop reforms.

*Cytokinesis
Division of the cell (animals) or formation of the wall (plants).

-Variations
No G₁ or G₂
Meiotic cell cycle with no DNA synthesis
No G₁, but tissue specific G₂.
DNA synthesis without mitosis- polytenization

6) Cell Growth Cycle

-Double everything in the cell
-Cytokinesis
-Growth Factors
Turn on or off gene
Phosphorylation
Ion fluxes
Cancerous growth
-Internal Factors
Cell Size regulates entry into S
Little or no cell growth in early embryos

7) Measurement of DNA

Quantity
Sperm = 1C. In G₁ then 2C. In G₂ then 4C

Quality
Sperm = 1N.
G₁, S, or G₂ = 2N.

Haploid = 1 complete set of chromosomes
Diploid = 2 complete sets of chromosomes (each pair are homologous chromosomes).

8) Crossovers

Areas of homology will recombine (Fig 12.21).
Occurs during prophase I.

9) Meiosis

One homologue from the father and one from the mother.
Homologues duplicate and remain together as sister chromatids.
Homologous Chromosome Pairing & Crossing over.
On average 2-3 crossover events occur per pair of chromosomes.
Here sister chromatids go to the same spindle pole.

Details
*Prophase I
Leptotene
-Pairing
-Crossing-over
Zygotene
-Synapsis (tight associations) Pachytene
-Synaptonemal complex is complete
Diplotene
-Chiasmata become visible (results of the crossover).
-Can be very long stage, in human females arrest occurs from the 7th month of fetal development until ovulation.
Diakinesis
-Chiasmata often terminalize, i.e. joined region moves to the telomeres.
*Metaphase I
Here sister chromatids remain attached.
*Anaphase I
Here sister chromatids go to the same spindle pole.
*Telophase I

*Prophase II
Chromosome contraction
*Metaphase II
Stage of arrest form most female mammals.
*Anaphase II
Centromeres split (similar to mitosis)
*Telophase II
Nuclear envelop reforms.

Meiosis generates 4 halpoid sets of chromosomes. (Fig 3.15 & 3.17).
SEX Determination Pg 77-84

1) Genotypic & Phenotypic Sex
Genotype is usually X or Y chromosomes. If a Y is present then it is male. If no Y is present then individual develops into a female.
XO are female and sterile "Turner syndrome" 1:10,000 females
XXY are males "Klinefelter syndrome" 1:1,000
Dosage compensation & X chromosome inactivation (Tbl 3.2)

Sry, Testis Determining Factor (TDF) and Tdf region of the Y chromosome. Some XX appear to be males, & some XY appear to be females.

2) Birds, butterflies, moths and some fish the male is the homogametic sex, i.e. ZZ; females are ZW.

3) Plants
Some are:
Dioecious, Male plants & Female plants (cedar trees).
Monoecious, Male organs and Female organs in different flowers on the same plant (corn).

4) Others
Marine worm:
-if free swimming settles alone, it becomes female;
-if it attaches to a female, then it becomes male.
Turtles
Some
Egg incubation >32 C, then female, <28 C, then male.
Snapping turtles are female if <20 or >30C, otherwise mainly males.

Take Home & You had Better Know

*Understand the difference between 'C' and 'N'.
*Understand why the products of first meiosis are not 1N or 2N.
*Understand what determines the 'sex' of a mammal.

Chapter 13 & 14

Transcription and Translation, or "DNA makes RNA makes Protein"

1) Transfer of genetic information from DNA to RNA.

*RNA polymerase acts like DNA polymerase and catalyzes the production of RNA (A,U,C,G).
*DNA unwinds at the promoter region and makes RNA 5' to 3’, using the 3' to 5' strand of DNA (Fig 13.1, 13.2, 13.3).

*messenger RNA (mRNA)

*transfer RNA (tRNA)

*ribosomal RNA(rRNA)

*small nuclear RNA (snRNA)

*Only mRNA codes for protein.

*RNA polymerases in eukaryotes (Tbl 13.1)
RNA polymerase I transcribes genes for rRNAs.
RNA polymerase II " "for mRNA .RNA polymerase III " "for 55 rRNAs,

tRNAs & snRNAs.

*Regulatory regions determine if transcription occurs.
Promoter- nearby regulation (Fig 13.6, 13.7). Transcription factors and regulatory factors.
Enhancer- distant regulation. Regulatory factors.

2) mRNA (structural genes)
mRNA vary in length due to length of the gene and protein.
mRNA has a 5' cap (Fig 13.11) and a 3' poly (A) tail.
mRNA must have introns spliced out before translation. This occurs with snRNPs
(small nuclear ribonucleoproteins) in a spliceosome (Fig 13.13).

3) rRNA
The eukaryotic ribosome has a size of 80S, and the subunits are 60S and 40S.
The large subunit contains 50 proteins, and 28S, 5.8S and 5S rRNA.
The small subunit contains 35 proteins and 18S rRNA.
rRNAs made by RNA polymerase I (18S, 5.8S and 28S rRNA are made as a single transcript
and then processed in the nucleolus (Fig 13.20).
5S rRNA is imported into the nucleolus where the ribosome is assembled.

4) tRNA
tRNA is 4S in size, and the leader and trailer are removed.
Molecules of tRNA bring amino acids to the ribosome.
The sequence 5'-CCA-3' is found at the 3' end of all tRNAs.

5) Proteins
Chains of individual amino acids linked by peptide bonds (Fig 14.1, 14.2, 14.3).
Primary structure- sequence of amino acids.
Secondary and tertiary structures after folding (Fig 14.4).
Function is determined by 3-dimensional structure.
Sequence of nucleotides (triplet code) in the mRNA specify the amino acid sequence.
Triplet code is (Fig 14.9):
-nearly universal
-degenerate, i.e. more than one codon for each amino acid.
-AUG is the start codon & codes for methionine
-Three stop codons (UAG, UAA, & UGA).

Protein synthesis occurs on the ribosomes

*Initiation
Methionine

*Elongation
First aminoacyl tRNA enters the P site
Second tRNA enters the A site (Fig 14.12).
Peptide bond is formed.
The chain moves down and a new tRNA enters the A site (Fig 14.17).

*Termination
Ribosome recognizes a stop codon and disengages from the reading frame of the mRNA (Fig 14.19).

Take Home & You had Better Know

DNA makes RNA makes Protein.
RNA synthesis is very similar to DNA synthesis.
Know the subclasses of RNA and their function.
Know what regulates RNA synthesis (promoter, enhancer, etc.).
Know what RNA's are processed.
Ribosomes contain rRNA and proteins.
Know what determines the amino acid sequence in a protein.
Know how a polypeptide is elongated and terminated.