Central dogma: Information flow in cells

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Central dogma Information flow in cells
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Nucleotides

• Pyrimidine bases Cytosine (C), Thymine (T),
  Uracil (U, in RNA)
• Purine bases Adenine (A), Guanine (U)

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Prokaryotic gene coding
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Eukaryotic processing of rRNA
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DNA Replication addition of a nucleotide
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DNA duplex formation
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A-T hydrogen bonding
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G-C hydrogen bonding
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3D structure of DNA
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Inverted repeats in DNA
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Formation of Stem-loops
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Sticky ends
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Hairpins
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Genetic Elements

• Prokaryotes Chromosome, plasmid, viral genome,
  transposable elements
• Eukaryotes Chromosomes, plasmid, mitochondrion
  or chloroplast genome, viral genome, transposable
  elements

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Melting of DNA

• Melting means separation of two strands from the
  heteroduplex
• Melting temperature of DNA is dependent on the
  relative number of AT and GC pairs
• Melted DNA can hybridize at temperatures below
  melting temperature
• This process can be used to test relatedness
  between species (interspecies DNA-DNA
  hybridization)
• It is also possible to reanneal DNA with rRNA to
  test relatedness of one species rRNA with the
  rRNA genes of another species

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Reannealing DNA
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DNA structure overview

• complementary strands (antiparallel)
• 3 Angstrom separationof hydrogen bonds
• sugar phosphate backbone held together
  with hydrogen bonding between bases
• size is expressed in nucleotide bases
  pairs. E. coli has 4600 kbp. (E. coli chromosome
  is gt 1mm, about 500X longer than the cell itself.
  How can the organism pack so much DNA into its
  cell?
• each bp takes up to 0.34nm, and each
  helix turn is 10bp(or 34 Angstroms), therefore
  how long is l kb of DNA? and how many turns does
  it have?
• inverted repeats, stem-loop, hairpins,
  sticky ends
• supercoiled DNA (DNA-binding proteins)
• relaxed, nicked circular DNA

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Supercoiled and relaxed DNA
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DNA Organization

• In prokaryotes naked circular DNA with negative
  supercoiling
• Negative supercoiling is introduced by DNA gyrase
  (topoisomerase II)
• Topoisomerase I relaxes supercoiling by way of
  single-strand nicks
• In eukaryotes linear DNA packaged around
  histones in units called nucleosomes
• The coiling around histones causes negative
  supercoiling

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Restriction and modification
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DNA Replication addition of a nucleotide
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Semiconservative replication
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Initiation of DNA replication
Origin of replication oriC 300bp Templates,
primers, polymerase, primase
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DNA Replication
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Bidirectional replication
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Okazaki fragments
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(No Transcript)
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Proofreading by DNA polymerase III
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Replication overview

• 1. origin of replication 300 bases, recognized
  by specific initiation proteins replication
  fork
• 2. bidirectional, therefore leading and
  lagging strands
• helicase unwinds the DNA a little
  (ATP-dependant)
• single-strand binding protein prevents
  single strand from reannealing
• Primase, DNA polymerase III and DNA
  polymerase I (also 5' to 3' exonuclease
  activity), ligase
• Okazaki fragments
• Topoisomerases, and supercoiling
  regulation
• 3. Proofreading (3 to 5' exonuclease
  activity by DNA pol III)

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DNA Sequencing
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Transcription

• RNA plays an important role
• tRNA, mRNA, rRNA
• Name three differences between chemistry of RNA
  and DNA
• RNA has both functional and genetic roles

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Initiation of Transcription
Pribnow boxtataat
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Transcription
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Completion of transcription
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Example of termination sequence
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More transcription

• Polycistronic mRNA
• How can mRNA be used in microbial ecology?
• Antibiotics and RNA polymerases

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RNA processing

• Removal of introns
• Ribozymes (nobel prize-Tom Cech and Sid Altman)
• RNA-splicing enzymes
• Origins of life? Which came first RNA or DNA?

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The genetic code

• Notice that the wobble base generally makes minor
  changes in the amino acid
• AUG is the start code (formyl methionine) for
  bacteria
• UAA, UAG, UGA are stop codons
• Specific tRNA for each other codon

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Codon and Anticodon Wobble
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tRNA associated with codon
60 specific tRNAs in prokaryotes
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mRNA, tRNA and ribosomes
Shine Dalgarno sequence GTP and Elongation
Factors (EF)
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Growing protein polymer
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Translocation
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Role of rRNA in protein synthesis

• Structural and functional role
• 16S rRNA involved in initiation
• Base pairing occurs between ribosome binding
  sequence on the mRNA and a complementary seq on
  the 16S rRNA
• 23S rRNA involved in elongation
• Interacts with EFs

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Chaperones (heat-shock proteins)
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Overview of today

• Summarized basic DNA structure
• DNA replication
• DNA sequencing
• Transcription
• RNA processing
• Translation
• Role of rRNA in protein synthesis