unit 1 prokaryotic genomes and genetic mapping

• Viral, Prokaryotic & Eukaryotic genomes

  All genomes are composed of ribo- or deoxyribo-nucleotides
• Bacteriophages

  Viruses that infect and replicate in bacteria
• 1. value

  The total DNA content of a genome, represents the number of bases in the single set (haploid) of chromosomes
• Table showing the characteristics of selected genomes

  • Type of Nucleic Acid
  • Number of Genes (protein estimates also acceptable)
  • C-value or Size - kilobases
• fd bacteriophage (host is E. coli)

  • ssDNA
  • 12
  • 6.4 kB

  
• The DNA in prokaryotic cells is found in the cytoplasm as a dense clump or clumps known as a nucleoid and occupy ~1/3 of the cell

  
• When stretched out, the E. coli chromosome is approximately 1.1 mm in length (about 1,000 times the size of a single cell)

  
• The nucleoid does not have a membrane and in E. coli consists of ~400 folded loops of ~40 kb (40,000 base pairs) each

  
• The compacted DNA is stabilized by the loops being held/anchored by an unknown mechanism by a core of proteins and possibly some RNA

  
• Negative supercoiling


  If DNA is a spiral staircase, it would be deemed to be negatively super-coiled if there was the same number of steps but at least one less 360 turn
• Positive supercoiling


  If DNA is a spiral staircase, it would be deemed to be positively super-coiled if there was at least one additional 360 turn
• How E. coli Topoisomerase II (eg. DNA gyrase) introduces negative supercoiling into DNA

  Describes the process

  
• How E.coli Topoisomerase I relaxes supercoiled DNA

  Describes the process

  
• Antibiotics that target DNA gyrase


  A class of antibiotics
• Negative supercoiling of the bacterial chromosome is important, it makes the DNA more compact, and also affects gene function
• Negative supercoiling creates tensions that may be released by DNA strand separation, which is necessary for many genetic functions such as replication and transcription

  
• Bacterial chromosomes


  • Generally circular and consist of double-stranded DNA
  • Have a unique single "origin of replication" (known as ori)
  • Replication is initiated at ori
  • Replication proceeds in a bi-directional manner
  • Completed at a unique termination of replication site (known as ter)
  • Effectively a single replicon
• If it takes 40 minutes to replicate the E. coli chromosome

  How can E. coli divide every ~20 minutes under optimal growth conditions?

  
• E. coli cells replicate their chromosome to produce two identical copies, and the cell then divides by binary fission

  
• The nucleoid (genome) is not randomly positioned within the bacterial cell and positioning depends on the cell cycle

  
• FtsZ protein


  Assembles into a ring at the future site of the septum and recruits other proteins, distantly related to a protein family known as 'tubulin' in eukaryotic cells

  
• Most prokaryotic cells (and viruses) have very little repetitive DNA sequences and have high gene densities, unlike eukaryotic cells that have large amounts of repetitive sequences
• The non-transcribed regions of DNA located between genes are termed intergenic regions

  
• Genomes of all species are not static – genes can be gained or lost, which is very important in human medicine when bacteria gain genes that code for antibiotic resistance
• Plasmids


  • Small molecules of circular DNA that exist independently of the host organism's chromosome
  • Can be as small as 1-2 kb or over 100 kb
  • Can vary in number from one copy to >300 copies per cell
  • Code for their own replication and usually one or more cellular traits that are not essential for normal metabolism, but confer an advantage to the cell (eg. antibiotic resistance)
  • Can be engineered for use as cloning vectors

  
• Key features for different types of plasmids

  • Fertility (F) Factor
  • Resistance (R)
  • Bacteriocinogen
  • Cryptic
  • Degradative
  • Virulence
• Why do plasmids threaten our continued and effective use of antibiotics?

  Plasmids can transfer antibiotic resistance genes between bacteria, reducing the effectiveness of antibiotics
• Transposable elements


  • Ubiquitous components of both prokaryotic and eukaryotic genomes
  • Repetitive sequences of DNA that can move (transpose) to different, non-homologous positions in the genome, creating large changes to the genome
• Types of transposable elements

  • Insertion sequences
  • Transposons
• When transposable elements are inserted into a new site, they may disrupt gene function by causing loss of function (LOF) mutations
• Insertion Sequences (IS)


  Structural features described and drawn
• Transformation


  • Process where a recipient bacterial cell takes up DNA from the environment
  • Discovered by Frederick Griffith in 1928
  • Cells with the ability to take up DNA from the environment are said to be competent
• Insertion Sequences (IS)


  Genetic elements that can move within and between genomes
• Insertion Sequences (IS)


  • Have a simple structure with only the genes required for transposition
  • Can move within and between genomes
• Transposons


  Genetic elements that can move within and between genomes
• Main types of transposons


  • Composite
  • Non-composite
• Composite transposons


  • Contain additional genes between the two insertion sequences
  • Can carry antibiotic resistance genes
• Non-composite transposons


  • Contain only the genes required for transposition, no additional genes
• Transformation is the process where a recipient bacterial cell takes up DNA from the environment
• Griffith discovered that live bacteria were "transformed" by heat-killed virulent bacteria, due to uptake of DNA by live bacteria