GENETICS

History of discovery of DNA structure

Structure of the nucleic acids

What do geneticists do?

Genetics is often divided into several branches; Geneticists employ a wide variety of different techniques, depending on the area that they study. The following brief list describes some important branches and the principle techniques employed in each.• Molecular genetics• Cytogenetics• Population genetics• Developmental genetics• Human genetics• Applied genetics

What will we do in our class in genetics?All of those things!Work with DNAWork with cells and transform themWork with population genetics etc………Experiments, speakers, reading……Just having fun and learning!

Three broad areas of genetics:

• Molecular genetics – focuses on the biochemical understanding of the hereditary material

• Transmission genetics – explores the inheritance patterns of traits as they are passed from parents to offspring

• Population genetics – concerned with genetic variation and its role in evolution

We will begin with DNA….the stuff of genes!

Molecular Genetics

Reference Chapter 16 Campbell

• Early in the 20th century, the identification of the molecules of inheritance loomed as a major challenge to biologists

• When T. H. Morgan’s group showed that genes are located on chromosomes, the two components of chromosomes—DNA and protein—became candidates for the genetic material.

• Was it the DNA or the protein that was the genetic material?

Evidence That DNA Can Transform Bacteria

• The discovery of the genetic role of DNA began with research by Frederick Griffith in 1928

• Griffith worked with two strains of a pneumonia bacterium, one pathogenic (S) and one harmless (R)

Fig. 16-2

Living S cells (control)

Living R cells (control)

Heat-killed S cells (control)

Mixture of heat-killed S cells and living R cells

Mouse diesMouse dies Mouse healthy Mouse healthy

Living S cells

RESULTS

EXPERIMENT

When heat-killed remains of the pathogenic S strain were mixed with living cells of the harmless R strain, some living cells became pathogenic

What NEW genetic information was being transformed to the bacterial cells?

Griffith did not identify the

transformingsubstance.

In 1943, Avery, MacCleod, and McCarty proved that the “transforming principle”was DNA.

• In 1952, Hershey and Chase used bacteriophage viruses and proved that the hereditary material was indeed DNA.

• They labeled the protein with S35 and the DNA with P32. Only the cells that contained the P32 were radioactive, showing that DNA was the material that coded for heredity.Why did

they use phage

viruses?

Do you remember how viruses replicate?

Life cycle of bacteriophage viruses

Google Image Result for http://www.slic2.wsu.edu:82/hurlbert/micro101/images/101PhageLife.gif

Why did they use a blender?

supernatant

Lighter phage coats

After most biologists became convinced that DNA was the genetic material, the challenge was to determine how its structure accounts for its role.

So, the race began!

• Chargaff’s rules state that in any species there is an equal number of A and T bases, and an equal number of G and C bases

He used paper chromatography after purifying the bases from each species.

Chargaff’s results:

What about the double helical structure?

• Linus Pauling (1950’s) proposed that regions or proteins fold into a helical structure (What level of protein structure?) He used models to visualize a helical structure for molecules.

I sure would like to figure

out the structure of

DNA!

A more important development was the X-ray crystallography of the DNA molecule.• Rosalind Franklin, working in the same lab as Maurice Wilkins,

made remarkable images of the DNA molecule using crystallography.

What did the pattern show?

• Helical structure• Diameter was too wide to be a single strand• Helix contained about 10 base pairs per complete

turn

Watson and Crick built models based on what they found out from Pauling, Chargaff, Franklin, and Wilkins.

Was it really that

easy?

Today we rely on computer models more.

The Race for the Double HelixJames

Watson Jeff Goldblum

He was popular in DNA movies!

Talk about a genetic Mutant!

I knew it wasa bad thingto restore hisDNA!

The

The FlyJurassic Park

Nucleic Acid Structure

• Term derived from discovery of DNA by Friedrich Miescher in 1869. He identified a novel phosphorus-containing substance from the nuclei of white blood cells found in waste surgical bandages. He named this substance nuclein.

DNA and RNA are acidic molecules because they release hydrogen ions in solution and have a net negative charge at neutral pH, due to the phosphates.

• Nucleic Acids have levels of complexity similar to proteins.

• (a) Nucleotides are the repeating structural units

• (b) nucleotides are linked together in strands

• (c) Two strands of DNA (and sometimes RNA) interact to form a double helix.

• (d) A 3-dimensional structure exists due to folding and bending to enable bonding to proteins to form chromosomes.

Protein levels of complexity

Types and Functions of Nucleic Acids

• DNA stores genetic information used for the synthesis of proteins including enzymes and is found in the nucleus and mitochondria.

• RNA has several functions and is found in the nucleus, cytosol and mitochondria.

• Messenger RNA (mRNA) carries genetic information obtained from DNA to sites that translate the information into a protein.

• Transfer RNA (tRNA) carries activated amino acids to sites where the amino acids are linked together to form polypeptides.

• Ribosomal RNA (rRNA) is a structural component of ribosomes, which serve as the sites for protein synthesis.

• Small nuclear RNA (snRNA) is a component of small nuclear ribonucleoprotein particles. These particles process heterogeneous RNA (hnRNA, the immature form of mRNA) into mature mRNA.

• RNAi (interference RNA) process , microRNA’s (miRNAs) and small interence (siRNAs) are genomically encoded noncoding RNAs that help regulate gene expression, particularly during development.

• In some viruses (HIV, influenza, polio), RNA functions as the storage house of genetic information.

RNAi pathway

The nucleotide, the basic unit of DNA, RNA

A review of the bases

The Double Helix

Structural Features of the DNA molecule

1) The sequence or order of the nucleotides defines the primary structure of DNA and RNA. The nucleotides of the polymer are linked by phosphodiester bonds connecting through the oxygen on the 5' carbon of one to the oxygen on the 3' carbon of another. The oxygen atoms in the backbone give DNA and RNA "polarity". The backbone is negatively charged due to a negative charge on each phosphate. The bases form flattened planar structures and are oriented so that the flattened regions are facing each other, an arrangement referred to as base stacking. Base stacking (along with H bonding) stabilizes the double helix and excludes water molecules.

Base Stacking

The orientation of the nucleotides is the second important structural feature. Notice the orientation of the strands; they are antiparallel to each other. Therefore, a strand has a directionality to it.

The nucleotides within a strand are covalently attached to each other so the sequence of bases cannot shuffle around and become rearranged. This is the defining feature that allows DNA

to carry genetic information. The phosphodiester bond is

covalent.

These bases cannot move around!

The sequence of the nucleotides codefor genetic information.

Forms of DNA – differ in handedness, the length of the helix turn, the number ofbase pairs per turn, and difference in size ofmajor and minor grooves.

I sure thoughtDNA was in the A form.

RNA• The secondary structure of RNA consists of a single

polynucleotide. • RNA can fold so that base pairing occurs between

complementary regions. RNA molecules often contain both single- and double-stranded regions. The strands are antiparallel and assume a helical shape. The helices are of the A form . RNA molecules are typically much shorter than DNA molecules.

• The structure of t (transfer) and r(ribosomal) RNA consists of multiple, single stranded, stem-loop structures. The stems consist of helices formed by base pairing of complementary regions within the RNA. The secondary structure of tRNA and rRNA are important for their biological functions, mRNA also assumes some degree of secondary structure but not to the same extent as tRNA and rRNA.

• Some of the bases in DNA and RNA can be chemically modified via methylation (adding a CH3 group).

• Enzymes, similar to proteases, called exo- and endo-nucleases can cleave RNA and DNA. Exonucleases cleave nucleic acids from the ends. Endonucleases (restriction enzymes) recognize specific sequences of duplex DNA and cleave at a specific site within or near the recognized sequence. The sequences that are recognized range from four to eight base pairs in length. The resulting fragments can be joined to other fragments to create new combinations of DNA sequences (recombinant DNA).

Endonuclease

• Remember: Restriction enzymes were first isolated from bacterial cells that used them to defend themselves from enemy viruses and other bacteria. They are named for the bacteria cells that were isolated from.

Restriction Enzymes They recognize certain basesequences.

Making Recombinant DNA

Denaturing and Renaturing DNA

• DNA can be denatured into single strands and renatured back into a double helix. Reversible denaturation is essential for the biological processes of replication and transcription; and for molecular biological techniques such as Southern blotting and polymerase chain reactions (PCR's).

There are three ways to denature DNA:• Enzymatically• Chemically• With heat

• Because GC base pairs are held together with three hydrogen bonds (AT have only two) the higher the percentage of GC base pairs the higher the heat required to melt the DNA.

• For renaturation to take place the two strands of DNA must contact one another to initiate base pairing. Once this happens the two strands quickly reassociate along their entire length. Several things influence renaturation:

complexity, DNA concentration, cation concentration, and temperature

Cations such as sodium, potassium and magnesium decrease the intermolecular repulsion of the negatively charged phosphate backbones of the two DNA strands.

Takes more heat

Denaturation of DNATm is the melting temperature in which 50 % of the DNA is denatured.

PCR Polymerase Chain Reaction

Southern Blotting – isolating DNA fragments