Microbial Genetics

Genetic material of Esch coli Single circular DNA 5x10 6 base pairs Codes for 2000 proteins haploid- single chromosome

Structure and Function of Genetic Material DNA & RNA DNA=deoxyribonucleic acid RNA=ribonucleic acid Basic building blocks: Nucleotides Phosphate group Pentose sugar Nitrogenous base DNA and RNA-polymers Nucleotides-monomers

Structure of DNA Double stranded (double helix) Chains of nucleotides 5’ to 3’ (strands are anti-parallel) Complimentary base pairing A-T G-C A-adenine T-thymine G-guanine C-cytosine

DNA Structure Phosphate- P Sugar- blue Bases- AT GC Hydrogen bonds-hold base pairs together A-T 5’ end-means P comes off 5’ carbon of deoxyribose sugar 3’means P comes of 3’ end of deoxyribose sugar Hydrogen bonds-hold base pairs together 5’ end-means P comes off 5’ carbon of deoxyribose sugar 3’means P comes of 3’ end of deoxyribose sugar

DNA Replication Bacteria have closed, circular DNA Genome: genetic material in an organism E. coli 4 million base pairs 1 mm long (over 1000 times larger that actual bacterial cell) DNA takes up around 10% of cell volume

DNA Replication-occurs at the replication fork 5’ to 3 ‘ DNA helicase-unzips + parental DNA strand that is used as a template Leading stand (5’ to 3’-continuous) *DNA polymerase-joins growing DNA strand after nucleotides are aligned (complimentary) Lagging strand (5’ to 3’-not continuous) *RNA polymerase (makes short RNA primer) *DNA polymerase (extends RNA primer then digests RNA primer and replaces it with DNA) *DNA ligase (seals Okazaki fragments-the newly formed DNA fragments) During replication, on parental ds DNA is converted into 2 identical ds daughter molecules… After DNA helicase unzips DNA strand, free nucleotides present in cytoplasm are matched up to exposed bases on the single stranded parental DNA Dna polymerase can add nucleotides only to the 3’ end Okazaki fragments-1000 nucleotides

Replication Fork

Protein Synthesis DNA------- mRNA------ protein transcription translation DNA ribosome Central Dogma of Molecular Genetics

Transcription One strand of DNA used as a template to make a complimentary strand of mRNA Promoter/RNA polymerase/termination site/5’ to 3’ Ways in which RNA & DNA differ: RNA is SS RNA sugar is ribose Base pairing-A-U instead of A-T mRNA-messenger RNA

Transcription

Types of RNA Three types: mRNA: messenger RNA rRNA: ribosomal RNA Contains 3 bases ( codon) rRNA: ribosomal RNA Comprises the 70 S ribosome tRNA: transfer RNA Transfers amino acids to ribosomes for protein synthesis Contains the anticodon (3 base sequence that is complimentary to codon on mRNA) mRNA-messenger

Genetic Code DNA: triplet code---- A T C mRNA: codon (complimentary to triplet code of DNA) U A G tRNA: anticodon (complimentary to codon) A T C

Genetic Code Codons: code for the production of a specific amino acid 20 amino acids 3 base code Degenerative: more than 1 codon codes for an amino acid Universal: in all living organisms 64 total codons 3 STOP codons uaa,uga,uag 1 start codon (for protein synthesis) aug (also a sense codon that forms aa)

Genetic Code

Translation Three parts: Initiation-start codon (AUG) Elongation-ribosome moves along mRNA Termination: stop codon (UGA) reached/polypeptide released and new protein forms rRNA=subunits that form the 70 S ribosomes (protein synthesis occurs here) tRNA=transfers amino acids to ribosomes for protein synthesis)

MUTATIONS Change in base sequence of DNA Different AA inserted in protein Altered phenotype

Mutations Changes in base sequence of DNA lethal and inheritable Can be: Harmful Lethal Helpful Silent Silent-neutral

Normal DNA Missense Mutation—base subsitution Missense-just one different amino acid formed-caused from a base substitution, single base is replaced with a different one

c ) Nonsense Mutation (base subsitution stop codon UGA) d) Frameshift Mutation (base inserted or deleted) Nonsense mutation-base substitution in the middle of the mRNA results in the formation of the stop codon/protein synthesis stopped Frameshift-1 or a few nucleotides are deleted or inserted-alters 3by3 transitional reading frame/produces inactive protein Base substitutions and frameshift mutations occur spontaneously or by chemicals in the environment

Causes of mutations Chemicals Radiations Viruses ---inserted bonds of wrong bases not AT but AC Base analogues are taken up – bromouracil instead of thymine Frame shift mutations----tobacco insert between bases Radiations X-rays breaks bonds of ribose phosphate chain, Free radicals—damage bases, H bonds of bases are altered by changing outer electrons of bases. UV—dimers of bases----replication inhibited Viruses ---inserted

Mutation due to transposons Transfers DNA in same bacteria from one site of chromosome to another or to plasmid Make copy and insert where Inserted can cause mutation

WHAT ARE TRANSPOSONS? Pieces of DNA Jumping genes Moves within DNA or Between DNA, Plasmid &Bacteriophage Replicate n make copy but not indepedently only with DNA Insertion sequences small transposons Code for drug resistsnce & enzymes,toxins

WHAT ARE PLASMIDS ? Extra chromosomal DS DNA in cytoplasm Capable of independent replication Integrate in DNA Transmissable & non transmissable Code for antibiotic resistance ,toxins,enzymes

Genetic Transfer in Bacteria Genetic transfer-results in genetic variation Genetic variation-needed for evolution Three ways: Transformation: genes transferred from one bacterium to another as “naked” DNA Conjugation: plasmids transferred to one bacteria to another via a pilus Transduction: DNA transferred from one bacteria to another by a virus Transduction-DNA passed from 1 bacteria to another in a bacteriophage (virus) and then incorporated into the host DNA

Transformation

Transduction by a Bacteriophage

Examples of transduction bacteriophage Botulinism toxin Corynebacterium toxin Diptheria toxin Erythrogenic toxin

Conjugation in E. coli F factor-plasmid F+ cell –donor/f- is recipient Hfr-when plasmid becomes incorporated into chromosome of bacteria, that cell is called a high frequency of recombination cell During conjugation, hfr cell can transfer chromosomal dna to f-cell

Conjugation continued…

Conjugation continued…

Bacteriophage containing host-cell DNA is involved in which of the following processes?  Transformation Conjugation Transduction Transcription Recombination

Plasmids is involved in which of the following processes?  Transformation Conjugation Transduction Transcription Recombination

Mobile genetic elements that code for antibiotic resistance genes in bacteria but are incapable of self-replication are plasmids R factor Temperate RNA phages Transposons Virulent DNA phages