speedrun bsci223 #2

lag phase

• Not actively growing and reproducing but recovering and trying to get used to new environment by synthesizing materials

log phase

• Exponential phase is the only time this growth equation is applicable (only time the doubling rate will be constant) 

• Metabolic rate is at its maximum rate

stationary phase

• Due to nutrient deficiency/ toxin accumulation

• Cells have either stopped growth or growth rate = death rate (flatline)

death phase

• Waste accumulates with no nutrients added 

• Cells begin to lyse faster than they are produced 

• Death rate > growth rate

Direct cell counts 

• manual

• no culturing

• fast

• Can't tell if cells are alive or dead with out special staining

Viable plate counts 

• Dilution plating: count # of colonies formed by viable cells

• Only includes cells able to reproduce and form colonies: CFU 

• requires media and time

Indirect cell counts (Turbidity)

• Amount of light transmitted through a culture (“shadow” of bacteria)

• More bacteria = Less light transmitted (More light is absorbed)

• rapid measuring

• Can’t tell if cells are alive or dead

DNA directionality

• Bases can only be added on the 3’ end so elongation of the DNA molecule is always going in a 5’ → 3’ direction

DNA Replication

• Copies information from one strand to a new, complementary strand 

• DNA Helicase unwinds DNA at oriC by breaking hydrogen bonds between bases 

• DNA Poly binds to each strand and adds nucleotides to 3’ end

• Terminus: DNA regions at which each replication fork ends during replication 

DNA Transcription

• Begins with promoters which have recognition sequences that RNA Poly recognizes 

• Binds to DNA with help from Sigma factors which guides to the promoter 

• Starts at +1 site (TSS), first nucleotide transcribed

• RNA Poly unzips DNA to expose template strand

• Builds complementary nucleotides from 5’ —> 3’

Inducible Operon (lac operon)

• usually inactive

• must be turned on by by inducer (lactose)

• produces proteins that transport and catabolize nutrients

Repressible Operon (trp operon)

• usually active

• must be turned off by repressor (trp)

• produces proteins that are anabolic

Lac operon

• Encodes enzymes necessary for metabolism of lactose 

• lacZ, lacY, lacA

• on when lactose presence

• off when lactose absent

Central Dogma

• DNA —> RNA —> Protein

• replication —> transcription —> translation

Lacl Repressor Protein

• Lactose absent

• The lac repressor activates

• It binds to the operator on the DNA

• This blocks RNA polymerase from successfully transcribing the lac genes

• Transcription is OFF

Silent mutation

• amino acid sequence not mutated

• geno —> C

• pheno —> NC

Missense mutation

• amino acid sequence mutated to a different one

• geno —> C

• pheno —> C (depends on where on dna sequence)

Nonsense mutation

• amino acid sequence mutated to stop codon

• geno —> C

• pheno —> C, results in lack of function

Base-excision direct repair

• Occurs when there is a small amount of damage on one DNA strand

• Excises mutated dna base and replaces it 

Light direct repair 

• Occurs when there is a small amount of damage on one DNA strand

• Photolyase breaks pyrimidine dimers 

Nucleotide-excision single strand repair 

• damage cant be repaired directly

• cuts away section of a single strand containing damage and uses complementary strand as template to repair gap

Mismatch single strand repair

• damage cant be repaired directly

• remove and replace mismatched bases

• must repair before new strand is methylated or cant tell old strand from new strand

Error prone repair

• damage so extensive it cant be corrected with normal mechanism

• SOS response, last resort before cell death

• fill in gaps with random sequences when no template

Lytic phase

• assembles new phages inside cell then releases

• kills host cell

Lysogenic phase

• integrate into host genome

• replicates when host divides

• can reactivate to become lytic due to mutagens

Transformation

• take up external DNA in environment

Transduction

• transfer by bacteriophage

Conjugation

• one way transfer of DNA by pilus of another cell of a related species

Sticky ends vs Blunt ends

• Sticky: single strand cut, creates overhanging sequences

• Blunt: double stranded cut, straight through DNA

Positive selection

• Kills all cells except those with desired phenotype

• Cell has ampicillin resistant gene (blaA) on a plate with ampicillin

Negative selection

• Kills all the cells with the desired phenotype

Screening

• everything survives but you include something else to differentiate cells

• adding X-gal to a plate with lactose so the cells with the lac operon will turn blue

• lacZ makes β-galactosidase which cleaves X-gal to turn blue

Mutation Rate