Microbial Genetics, Characteristics of Prokaryotic DNA

The DNA in most prokaryotic cells exists as a single, covalently bonded circular piece of chromosome
Prokaryotic genes involved with the same process is clustered

1. Microbial Genetics

2. Introduction The DNA in most prokaryotic cells exists as a single, covalently bonded circular piece of chromosome Prokaryotic genes involved with the same process is clustered Prokaryotic DNA is tightly packed DNA occupies one-third space in the cell Fully extended it measures 1.5 mm long, i.e. 500X the length of bacteria ( E. coli ) It adjusts by supercoiling - a twisting and tight packing Twisting up of DNA; super coiled domain (coils folded to form loops of 10,000 bases); loop domain structure

3. Many prokaryotic cells also contains plasmids- any circular DNA molecule in cells that replicates independently of the chromosome and regulates its own replication so that the number o copies of the DNA molecule remains relatively constant Do not carry information for normal structure, growth or metabolism Plasmids account for 2% of total genetic information Some plasmids are large enough to integrate into chromosomes and excise from it many essential genes

4. Plasmid confers selective advantage and genetic flexibility F plasmids: allow transfer of genetic information from donor to recipient through a recombination process R plasmid(resistance factor): provides genes for antibiotic resistance, resistance to heavy metals (Silver and mercury) Provides genes for production of antibiotics and bacteriocins ; toxin production (anthrax)

5. Characteristics of prokaryotic and eukaryotic chromosomes Prokaryotic Organized in the nucleoid Chromosome usually circular, about 1.5 mm in length Single molecule of DNA per genetic trait: haploid 4,300 genes in E. coli Little protein present No dominance or recessive Introns very rare Replicate just prior to binary fission Single or multiple replication origins Mutation in DNA Eukaryotic Organized in nucleus Chromosome linear Two molecules of DNA per genetic trait: diploid; some haploid 35,000 genes in humans Histone protein present Genes may be dominant or recessive Introns present Replication just prior to mitosis Multiple replication origin Mutation in DNa

6. Bacterial genetics Manipulation of bacterial DNA to study the cellular function Bacteria are haploid Have only one copy or allele of each gene Short generation time Doubling time of E. coli is 20 minutes Asexual reproduction By cell division to form clones Colony growth on agar plates Colony purification of bacterial strains and mutants Serial dilution Selections under selective conditions Storing stocks of bacterial strains Genetic exchange: transformation, conjugation, transduction

7. Genetic recombination Formation of a new genotype by reassortment of genes following an exchange of genetic material between two different chromosomes which have similar genes at corresponding sites Genetic information in bacteria can be transferred vertically or horizontally Recombination important for survival, evolution, change in gene order within, repair of DNA damage

8. In bacteria genetic recombination results from three types of gene transfer Conjugation : transfer of genes between cells that re in physical contact with one another Transduction : transfer of genes from one cell to another by a bacteriophage Transformation : transfer of cell-free or ‘naked’ DNA from one cell to another

9. Homologous/general recombination: A process of genetic re-assortment between homologous DNA sequence from two different source or within the same genome Homologous DNA has the same or nearly the same sequence; this type of process occurs in “ crossing over” in classical genetics Site-specific recombination: integration of phage lambda into genome of bacteria Non- recA protein dependent recombination mediated by phage encoded protein and host encoded integration factor (IHF) Homologous (ectopic) recombination Recombination between different region in two DNA molecules because the same or similar sequence occur in more than one place in the DNAs Recombination

10. General mechanism of recombination

11. Basics of recombination Identical or very similar sequences in the crossover region Mandatory complementary base pairing between double stranded DNA molecule-synapse formation Recombination enzymes-DNA endonucleases and ligases Heteroduplex formation

12. Holiday double strand invasion model (Robin Holiday 1964) and Chi form

13. Migration of Holiday junction

14. Single strand invasion model

15. Double strand break repair model