DNA Bio 11 Oct 14, 2008

The pedigree below most likely depicts a ______ pattern of inheritance. A) Autosomal Dominant B) Autosomal recessive C) X-linked Dominant D) X-linked Recessive E) mitochondrial

Why are sex-linked conditions more common in men than in women? A) Men acquire two copies of the defective gene during fertilization. B) Men need to inherit only one copy of the recessive allele for the condition to be fully expressed. C) Women simply do not develop the disease regardless of their genetic composition. D) The sex chromosomes are more active in men than in women. E) None of the choices are correct.

Announcements Next exam: Oct 28 (2 weeks!) Mitosis, Meiosis, Genetics, DNA, Central dogma, Gene regulation, DNA technology Next quiz: Oct 16

Sex linked genes helped T.H. Morgan determine that chromosomes contained heritable factors T. H. Morgan initially showed that chromosomes determine sex Mutant genes showed that individual genes were on chromosomes as well

Chromosomes are made of more than one substance Protein and DNA Which substance held the information? DNA: 4 different bases Protein: 20 different amino acids Could it be both?

DNA DNA’s double helix was elucidated by James Watson and Francis Crick in 1953 DNA was known since early 1900’s

The Hershey-Chase experiment showed that DNA was the genetic material

Viruses are not cells

Viruses are in the gray area between living and nonliving things They have a completely regular structure They have genetic information They do not have a metabolism They cannot reproduce themselves Are they alive?

Basic life cycle of Viruses Viruses are obligate intracellular parasites They inject their genetic material into their host Host machinery is commandeered to mass-produce virus Viruses burst host cell to infect other cells

Viral Structure Genetic material (not necessarily DNA) A protein capsid coded for by the genetic material Some eukaryotic viruses have an envelope Various accessory proteins

Bacteria are infected by viruses, too

Bacteriophages Infect bacteria Cause formation of plaques on a lawn of agar in bacteria

Phage structure Looks like the lunar lander Head contains genetic material Tail fibers bind to surface antigens of host Tail injects genome into host

DNA contains phosphorus, Protein contains sulfur

How does the Hershey-Chase experiment show which substance is the genetic material?

But how does DNA work?

Scientists expected that form could elucidate function

Additional Evidence That DNA Is the Genetic Material: Chargaff’s rules Amount of C=G, A=T DNA composition varies from one species to the next made DNA a more credible candidate for the genetic material Gave an important clue about how DNA worked

Rosalind Franklin used X-ray crystallography to determine helical structure

X-ray crystallography is still used today

Nobel Prize for chemistry 2007 Roger Kornberg, Stanford- Structure of RNA polymerase

DNA structure

DNA (and RNA) Nucleotides have 3 parts

There are 4 DNA bases

Bases interact by H-bonding

DNA structure is antiparallel There is a 3’ end and a 5’ end Each strand is unidirectional Many enzymes that replicate DNA are unidirectional also

Hydrogen bonding between DNA bases A with T, C with G CG pairs have 3 bonds, AT have two A always binds with T C always binds with G

How does DNA replicate?

LE Conservative model. The two parental strands reassociate after acting as templates for new strands, thus restoring the parental double helix. Semiconservative model. The two strands of the parental molecule separate, and each functions as a template for synthesis of a new, comple-mentary strand. Dispersive model. Each strand of both daughter molecules contains a mixture of old and newly synthesized DNA. Parent cell First replication Second replication

Meselson-Stahl experiment They labeled the nucleotides of the old strands with a heavy isotope of nitrogen The first replication produced a band of hybrid DNA, eliminating the conservative model A second replication produced both light and hybrid DNA, eliminating the dispersive model and supporting the semiconservative model

LE Bacteria cultured in medium containing 15 N DNA sample centrifuged after 20 min (after first replication) DNA sample centrifuged after 40 min (after second replication) Bacteria transferred to medium containing 14 N Less dense More dense Conservative model First replication Semiconservative model Second replication Dispersive model

DNA replication

DNA replication is catalyzed with enzymes

LE New strand 5 end Phosphate Base Sugar Template strand 3 end 5 end 3 end 5 end 3 end 5 end 3 end Nucleoside triphosphate DNA polymerase Pyrophosphate

Antiparallel Elongation The antiparallel structure of the double helix (two strands oriented in opposite directions) affects replication DNA polymerases add nucleotides only to the free 3  end of a growing strand; therefore, a new DNA strand can elongate only in the 5  to  3  direction

Okazaki fragments fill in DNA on lagging strand DNA polymerase is unidiirectional Each strand separated at an Ori must be copied

Along one template strand of DNA, called the leading strand, DNA polymerase can synthesize a complementary strand continuously, moving toward the replication fork To elongate the other new strand, called the lagging strand, DNA polymerase must work in the direction away from the replication fork The lagging strand is synthesized as a series of segments called Okazaki fragments, which are joined together by DNA ligase

LE Parental DNA 5 3 Leading strand Okazaki fragments Lagging strand DNA pol III Template strand Leading strand Lagging strand DNA ligase Template strand Overall direction of replication

Priming DNA Synthesis DNA polymerases cannot initiate synthesis of a polynucleotide; they can only add nucleotides to the 3 end The initial nucleotide strand is a short one called an RNA or DNA primer An enzyme called primase can start an RNA chain from scratch Only one primer is needed to synthesize the leading strand, but for the lagging strand each Okazaki fragment must be primed separately

LE 16-15_1 5 3 Primase joins RNA nucleotides into a primer. Template strand 5 3  Overall direction of replication

LE 16-15_2 5 3 Primase joins RNA nucleotides into a primer. Template strand 5 3  Overall direction of replication RNA primer DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment.

LE 16-15_3 5 3 Primase joins RNA nucleotides into a primer. Template strand 5 3  Overall direction of replication RNA primer DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment. Okazaki fragment After reaching the next RNA primer (not shown), DNA pol III falls off.

LE 16-15_4 5 3 Primase joins RNA nucleotides into a primer. Template strand 5 3  Overall direction of replication RNA primer DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment. Okazaki fragment After reaching the next RNA primer (not shown), DNA pol III falls off After the second fragment is primed, DNA pol III adds DNA nucleotides until it reaches the first primer and falls off.

LE 16-15_5 5 3 Primase joins RNA nucleotides into a primer. Template strand 5 3  Overall direction of replication RNA primer DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment. Okazaki fragment After reaching the next RNA primer (not shown), DNA pol III falls off After the second fragment is primed, DNA pol III adds DNA nucleotides until it reaches the first primer and falls off DNA pol I replaces the RNA with DNA, adding to the 3 end of fragment 2.

LE 16-15_6 5 3 Primase joins RNA nucleotides into a primer. Template strand 5 3  Overall direction of replication RNA primer DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment. Okazaki fragment After reaching the next RNA primer (not shown), DNA pol III falls off After the second fragment is primed, DNA pol III adds DNA nucleotides until it reaches the first primer and falls off DNA pol I replaces the RNA with DNA, adding to the 3 end of fragment DNA ligase forms a bond between the newest DNA and the adjacent DNA of fragment 1. The lagging strand in the region is now complete.