Level 2 Genetic Systems. M J Larkin

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Level 2 Genetic Systems.M J Larkin

• M J Larkin Biology Biochemistry. The Queens
  University of Belfast.

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Dr Mike Larkin - Communication

• MBC Room 113
• xt 2288 (Belfast 972288 Diverted to the DKB)
• DKB Questor Centre Room 318a
• 3rd Floor Microbiology Laboratory
• xt (90977) 4390 or (90977)4388 lab or (9033)
  5577 office
• Email m.larkin_at_qub.ac.uk
• Pigeon hole in Biology Biochemistry office
• ALWAYS LEAVE MESSAGE and HOW TO CONTACT YOU!

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Dr Mike Larkin - Communication continuedWWW
PAGES

• http//www.qub.ac.uk/mlpage/page1/index.html
• http//www.qub.ac.uk/mlpage/courses/level2/page.ht
  ml

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Course Theme reminder

• Half of the course dedicated to Microbial
  Genetics. Primarily Prokaryote Genetics.
• An introduction to bacteria and their genetic
  systems.

• Transfer of genes BETWEEN cells in bacteria
• Bacteriophages and plasmids (JQ)
• The fate of DNA WITHIN bacterial cells
• Mutation, recombination, repair, transposition
  (ML)
• Introduction to genomics and genome mapping
  (ML)

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Reading

• There are many texts with clear explanations e.g.
• Maloy S R et al (2001) Microbial Genetics. 4th
  Edition Jones and Bartlett. ISBN 0867202483
  QH434/MALO - A comprehensive text and good
  introduction 2004 edition due out
• Winter P et al (1998)Instant Notes in Genetics.
  OxfordBIOS Scientific. ISBN 1859961665
  QH440.2/WINT - Not a great coverage of bacterial
  genetics but useful nevertheless for
  recombination
• A good primer text for this part of the course is
  Dale J.W. (2004) Molecular Genetics of Bacteria.
  5th Edition Wiley ISBN 047085085X QH434.D35.
• ? DIRECTED READING will be indicated.

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Lectures

• Introduction. The basis of diversity
• Mutation, homologous recombination and repair (3
  lectures)
• Non-homologous recombination
• Mechanisms of transposition (2 lectures)
• Genomics and genome mapping
• Pre-genomic techniques
• Prokaryote genome sequencing (2 lectures)
• Revision tutorial

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Practicals Aims

• 1. Phage restriction
• Observe phage restriction and consider the
  implications of this phenomenon in nature.
• Acquire the skills needed to cultivate and titre
  phage in practice
• 2. Phage co-transduction
• Demonstrate the close linkage of two bacterial
  genes by transductional mapping
• Observe the connection between RecA function and
  DNA repair
• 3. Conjugation and transposition
• Demonstrate plasmid conjugation
• Demonstrate Insertion Sequence transposition/coint
  egration

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Introduction the basis of diversity.
LECTURE

• A. What you should know
• B. Terminology
• C. Breeding systems
• D. Types of mutation
• E. Reversion and suppression
• F. Mechanisms of mutation
• G. DNA damage repair
• H. Homologous recombination
• I. Homologous recombination and repair

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2
3
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Introduction

• A. What you should know.
• From Level 1 Courses
• The basis of the genetic code
• DNA replication
• The structure of the bacterial chromosome
• ? Maloy and Dale cover introductory material
  well.
• ? DNA structure and basis of code well covered
  in Winter A1

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Chromosomal replication is bidirectional
Cairns (1963) and Rodriguez (1973) Autoradiography
- 3H labelled DNA
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The structure of the bacterial Chromosome
Growing cells showing Geimsa stained
condensed chromosome
Lysed E.coli cell (lysozyme)
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Introduction cont.

• B. Terminology. See Practical schedule.
• The biggest source of confusion in practice
• Strains given numbers e.g. AB1157
• Mutant genotype written as e.g. arg lac Z
• Phenotype written as e.g. Arg or Lac Z
• (?C1857) or (P1Cm ) Lysogenic or cells carrying
  these phage
• (pOXKm) cells carrying this plasmid
• srlCTn10 indicates presence of transposon (or
  IS) in this gene
• Mutant or Mutation
• Auxotrophic or Prototrophic
• Mutant selection or mutant screening
• ? Dale Ch 2, Glossary and appendices are very
  useful.

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Introduction cont.

• C. Breeding Systems.
• Eukaryotes possess a SEXUAL CYCLE

?
?
DIPLOID 2n
HAPLOID 1n
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C. Breeding Systems. cont.

• Variation arises from from
• Mutation, recombination and random assortment

E.g Two Chromosomes randomly assorted.
4 x possibilities at Meiosis
HAPLOID 1n
DIPLOID 2n
3 Chromosomes --------- 16 possibilities n
chromosomes ---------- 2n possibilities
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C. Breeding Systems. cont.

• Humans have 223 POSSIBILITIES !
• Assuming 2 -3 crossovers per chromosome (i.e.
  about 60 in total) (223)60 variants possible
  in Meiosis !
• Mutation rate may be only 1 in 20,000 to 200,000
  per chromosome.
• Mutation adds variants at a low rate. If too
  high then a high GENETIC LOAD. Therefore
• Meiosis increases overall variation at a high
  frequency without a detrimental Genetic Load.

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C. Breeding Systems. cont.

• What about Prokaryotes ?
• No MEIOSIS possible
• Asexual reproduction
• Single Chromosome with plasmids
• Genetic recombination NOT associated with
  reproduction
• Mutation rate at single loci about 1 in 108 to
  1010 per cell per generation
• How is variability generated then ?
• At the gene transfer population GENE POOL level
• Complex mutation/recombination systems within
  cells

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The basis of diversity in bacteria

• Recombination
• Homologous
• Non-homologous
• Rearrangements/transpositions
• Deletions

• DNA Replication
• Mutation
• Repair

DNA Transformation
Plasmid Conjugation
Phage Transduction