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1 number 3 Done by Neda a Baniata Corrected by Waseem Abu Obeida Doctor Hamed Al Zoubi

2 Note: it is important to refer to slides. Bacterial genetics *The main concepts we will talk about in this lecture: We will talk about genome replication briefly, mutations, genetic exchange between bacteria, and finally we will talk about transposons briefly. Bacterial genome and replication: -Bacterial genome (chromosome): single circular double stranded DNA, found in a region called nucleoid. Bacterial genetic structure is represented by a genetic material consisting of: 1-chromosome: must be found in every bacterial genome. 2- Plasmids. 3-bacteriophage. 4-transposons. The last three are accessory components that might not be found in bacterial genetic material. *why we are interested in study of bacterial genetics? 1-Firstly, bacterial genetics is important in determining the structure of bacteria: genes are translated into proteins (sequence of amino acids).these proteins can have a structural

3 (being a component of the cell wall giving it more rigidity) or functional role (as enzymes or as pathogenic function like toxins). 2-It is important in determining antibiotic resistance of bacteria: to understand some of their mechanisms later on. 3-molecular cloning :sometimes we use bacterial genetics to clone some genes into the bacteria then bacteria will express it into proteins usually these proteins are useful to us, so we can extract it for using it for various aspects (vaccines,antibiotics,antigens, or for diagnosis). *Bacterial chromosome: -1300Mm long, super coiled. -it has (2-5) million nucleotide bases (nucleotide bases are: A, T, G, C base pairing: A T, G C) -Have enough DNA to encode around 1-3 thousand different genes according to bacteria types. -genes are expressed as required depending on the surroundings (nutrients, metabolites, temperature and pressure),or if some of the bacterial proteins are recognized by the immune system,the bacteria turns off the expression for these proteins. -Gene expression is controlled by many factors such as amount of end product metabolism and presence of repressors (a repressor is a DNA- or RNA-binding protein that inhibits the expression of one or more genes by binding to the operator or associated silencers.)

4 Genotype /wild type Represent the genetic structure of bacteria. All Inherited essential biological features & growth patterns. phenotype represent the formal characteristics of bacteria -phenotype determine a lot of characteristics of bacteria: biochemical rxns, irreversible factors that can be used to find bacteria.

5 *not every genotype is expressed to phenotype (they are expressed upon need as we mentioned earlier). Mutations:Mistakes or inaccuracies during DNA replication. Spontaneous mutations: it occurs naturally (not induced by an external factor), where its occurrence may result in a protein defect and the outcome will be significant. Induced Mutations: 1-chemicals, X-rays, UV light and viruses. 2-direct damage of nucleotides or alteration of nucleotide bonds. -these mutations could be induced in vitro and lead to dimerization of base pairs and this can affect replication. *In vitro: in laboratory or in animals. *In vivo: in human being. -Remember that: -A codon is a triplet of nucleotides that codes for one amino acid only, the same amino acid may be coded by more than one codon. - The translation begins from the start codon (AUG) and ends at the stop codon (UAA, UAG, UGA). -we call a triplet of nucleotides a codon if they exist on the mrna strand and we call it the reading frame.

6 Types of the mutation: 1-Multisite: affects many nucleotides by inversion, duplication or deletion. -Inversion for example occurs in bacteria to avoid recognition by the immune system. The gene which was recognized by the immune system will be changed by inversion, when threat disappears reinversion occurs. 2-Point mutation: affects single or few nucleotides by insertion, substitution, deletion.

7 Outcome of mutations: Outcomes of Base substitution which occurs due to DNA polymerase error or mutagens. 1)missense mutation : GGC Glycine AGC Serine DNA sequences change RNA sequences change Codes for different amino acids. 2)nonsense mutation : AAG lysine UAG stop codon DNA sequences change RNA sequences change Early stop codon introduced. (Translating stops protein incomplete so the bacteria loses the protein). 3)silent mutation: GGC, GGU both code for the same amino acid (glycine), So no effect on the amino acids sequence would occur. Frame-shift mutation : -One or more bases are added or deleted -Shift in the reading frame, Corrupting the reading of codons downstream leading to inactive protein. -Change in protein structure is significant to bacteria for example-if it is a toxin and after mutation become nontoxic it loses the pathogenicity trait- Can be lethal to bacteria. Or change in protein structure which is an antibiotic target causing resistance to antibiotics.

8 Genetic exchange: The importance of genetic exchange : 1-moving antibiotic resistance genes among bacteria. 2-moving virulence genes among bacteria. 3-changing the genetic makeup to avoid immunity. Mechanisms: 1-transformation: the uptake of naked DNA from the environment. -DNA source can be from dead bacteria cell and its DNA is free in the environment. -usually this mechanism occurs between the related bacteria. -a recipient cell must be competent; competent means: it has structures on its cell surface that can bind to DNA and take it up intracellularly). -Naked DNA will be up taken by competent bacterial cell through pores, a semi-permeable cytoplasmic membrane -then the DNA might be degraded or incorporated with bacterial chromosome to become a part of it. *normal competence: certain bacteria are naturally transformable. Example: streptococcus pneumoniae that has two populations classified according to the differences in their cellular Capsule: 1-encapsulated bacteria: stronger and more pathogenic 2-non encapsulated Bacteria: weak, is able to cause disease only if the immune system efficiency decreased. So the non-encapsulated kind can be transformed into the encapsulated, as a result the coming generation of these

9 transformed encapsulated bacteria will be stronger and more pathogenic. 2-trasduction: in this mechanism, bacteriophage is the vector of the genetic material. -Bacteriophage structure: head (contains the DNA), tail and fibres. -bacteriophages have two life cycles: 1-Lytic cycle: the phage injects its DNA in the bacterium resulting in the lysis of the bacterium and replication of its DNA material. 2-Lysogenic cycle: the phage injects its DNA and merges it with the bacterium DNA, the DNA of the virus replicates when the bacterium divides until there are better conditions for the bacterium DNA to separate from the bacterial genome and enter the lytic cycle to replicate and infect other bacteria.

10 -in the case of the lysogenic cycle, the virus genome is called prophage and the bacterium is called a lysogen, if the phage genome encodes an observable function, the lysogen will be altered in its phenotype (lysogenic conversion, example: cholera toxin).

11 Types of transduction: 1-generalized transduction (happens in the lytic cycle): usually when the phage attacks the bacterium and replicates, the newly synthesized bacteriophages contain only the DNA of the virus, but a few bacteriophages could carry the bacterial DNA in their capsids, and when attacking a new bacterium by lysogenic cycle the new host will contain part of the old bacterial genome adding a new property to the new host, for example adding antibiotic and toxin resistance. 2-specialized transduction (happens in the lysogenic cycle): The prophage integrates into the bacterial genome at a specific location, when a prophage is induced to lytic phase, it may drag along a piece of the bacterial genome next to the integration site and move that bacterial sequence into the new recipient host cell changing its genome(usually non medically significant).

12 3-conjugation: The importance of conjugation is to move certain features among bacteria such as moving antibiotic resistance genes that exist in the plasmids of certain bacteria(r plasmids). - mediated by plasmids: circular double-stranded DNA molecules that lie outside the chromosome and can carry many genes including those for drug resistance. -there are three possible states for plasmid conjugation: 1-conjugative: the plasmid contains a set of genes which promote sexual conjugation by sexual pilus with other bacteria, and the plasmid can move itself from the donor to the recipient cell. 2-mobilizable: the plasmid can only move with the help of a conjugative plasmid in the recipient cell. 3-non-transmissible: can t move by conjugation.

13 *steps of conjugation in general: 1-synthesis of the sex pilus. 2-cell to cell contact. 3-copying plasmid DNA and transfer of the copy into recipient cell. -bacteria containing a conjugative plasmid are called donor, male (F+). -bacteria receiving the plasmid are called recipient, female (F- ). *types of conjugation: 1) F+ conjugation: Genetic recombination in which there is a transfer of an F+ plasmid but not chromosomal DNA from a male donor bacterium to a female recipient bacterium. Other plasmids present in the cytoplasm of the bacterium, such as those coding for antibiotic resistance, may also be transferred during this process. 2)High frequency recombination (Hfr conjugation): In this type the plasmid ( F factor) integrates with the chromosome of the same bacteria(donor) before transport to the recipient cell, in this case the donor cell is called Hfr instead of F+, now the genome of the donor bacteria has the plasmid and the chromosome genes connected, the plasmid genes will initiate connection to other bacterium and try to transport the F factor to the other cell

14 ,but because the F factor is connected to the chromosome it will move the bacterial genes with it, but usually the connection breaks before moving all the genes, so the recipient bacteria will have some of the donor genes in addition to the F factor

15

16 3)F conjugation: in this type of conjugation the F factor merges with the chromosome of the same bacterium, picks up some genes and then breaks apart from the chromosome and now it is called the F factor, when the donor connects to a recipient cell it moves a copy of the F factor. 4-transposons: Jumping genes/copy and paste (Class1) or cut and paste (2) between plasmids or between chromosomes and plasmids. Medical importance: since many antibiotic resistance genes are encoded by transposons in antibiotic resistance. *it is considered fundamental transmission between plasmids and chromosomes. plasmids

17 THE END GOOD LUCK DOCTOR 2016

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