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Central dogma: Information flow in cells. Nucleotides. Pyrimidine bases: Cytosine (C), Thymine (T), Uracil (U, in RNA) Purine bases: Adenine (A), Guanine (U). Prokaryotic gene coding. Eukaryotic processing of rRNA. A-T hydrogen bonding. G-C hydrogen bonding. Genetic Elements.

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nucleotides
Nucleotides
  • Pyrimidine bases: Cytosine (C), Thymine (T), Uracil (U, in RNA)
  • Purine bases: Adenine (A), Guanine (U)
genetic elements
Genetic Elements
  • Prokaryotes: Chromosome, plasmid, viral genome, transposable elements
  • Eukaryotes: Chromosomes, plasmid, mitochondrion or chloroplast genome, viral genome, transposable elements
melting of dna
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
dna structure overview
**DNA structure overview**
  • complementary strands (antiparallel)
  • 3 Angstrom separation of 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 > 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?
  • supercoiled DNA (DNA-binding proteins)
dna organization
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
initiation of dna replication
Initiation of DNA replication

Origin of replication= oriC = ~300bp

Templates, primers, polymerase, primase

replication overview
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)
transcription
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
initiation of transcription
Initiation of Transcription

Pribnow box=tataat

more transcription
More transcription
  • Polycistronic mRNA
  • How can mRNA be used in microbial ecology?
  • Antibiotics and RNA polymerases
rna processing
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?
the genetic code
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
trna associated with codon
tRNA associated with codon

~60 specific tRNAs

in prokaryotes

mrna trna and ribosomes
mRNA, tRNA and ribosomes

Shine Dalgarno sequence

GTP and Elongation Factors (EF)

role of rrna in protein synthesis
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
overview of today
Overview of today
  • Summarized basic DNA structure
  • DNA replication
  • DNA sequencing
  • Transcription
  • RNA processing
  • Translation
  • Role of rRNA in protein synthesis