microbiology

• • Must obtain carbon in an organic form made by otherliving organisms
• • Such as proteins, carbohydrates, lipids, and nucleicacids

• • An organism that uses CO2 as its carbon source
• • Not nutritionally dependent on other living things

gain energy from chemical compounds

gain energy through photosynthesis

• • Oxygenic photosynthesis
• • Anoxygenic photosynthesis

• • lithoautotrophs
• • survive totally on inorganic substances

eats rocks

• • A kind of chemoautotroph
• • Produce methane gas under anaerobic conditions

• Majority of hetertrophes
• • Aerobic respiration
• – Two categories
• • Saprobes
• • Parasite

• – Free-living microorganisms that feed on organic detritusfrom dead organisms
• – Opportunistic pathogen
• – Facultative parasite

• – Derive nutrients from host
• – Pathogens
• – Some are obligate parasites

they are parasites that cause damage to cells (disease) or death.

does not require energy; substancesexist in a gradient and move from areas of higherconcentration toward areas of lower concentration

molecules propelled down a concentration gradient by random thermal motion.

diffusion of water

requires a carrier

requires energy and carrier proteins;gradient independent

transported molecule chemically altered

endocytosis, exocytosis, pinocytosis

solid and liquids materials are taken into the cell through membrane invagination or engulfment into a vesicle

releases enveloped viruses from the membrane of the host's cytoplasm

engulfment or endocytosis of liquid by extensions(microvilli) of the cell membrane

type of endocytosis in which the cell membrane actively engulfs large particles or cells into vesicles

When it has less solute outside the cell causing the cell to take in water and push against cell wall

when the cell has more solute outside the cell causing the cell to lose water and shrink.

When the cell has the same solute concentration inside and outside the cell

the relationship between two organisms that live together in a close partnership

members are obligatory, dependent; both members benefit

commensal member benefits,other member neither harmed nor benefited

parasite is dependent and benefits;host is harmed

• result when organisms attach to a substrate by some form of extracellular matrix that binds them together in complex organized layers
• • Dominate the structure of most natural environments on earth
• • Communicate and cooperate in the formation and function of biofilms – quorum sensing

Parent cell enlarges, duplicates its chromosome, and forms a central transverse septum dividing the cell into two daughter cells

Time required for a complete fission cycle

Each new fission cycle increases the population by a factor of 2

“flat” period of adjustment, enlargement; little growth

a period of maximum growth will continue as long as cells have adequate nutrients and a favorable environment

rate of cell growth equals rate of celldeath caused by depleted nutrients and O2, excretion of organic acids and pollutants

as limiting factors intensify, cells die exponentially

a graphical representation of the change in population size over time. It has 4 periods known as lag phase, exponential phase, stationary phase and death phase.

all chemical and physical workings of a cell

degradative;breaks the bonds of larger molecules forming smaller molecules; releases energy

biosynthesis;process that forms larger macromolecules from smaller molecules;requires energy input

• Enzymes are biological catalysts
• –Lower the energy of activation
• – The energy of activation is the resistance to a reaction
• – The enzyme is not permanently altered in the reaction
• • Enzymes use a specific lock and key fit with substrate

consist of protein alone

• – contain protein and nonprotein molecules
• – Apoenzyme is proteinportion
• – Cofactors is nonprotein portion
• • Metallic cofactors:iron,copper,magnesium
• – Coenzymes,organic molecules:vitamins

• – Site for substrate binding is active site,or catalytic site
• – A temporary enzyme-­‐substrate union occurs when substrate moves into active site=induced fit
• – Appropriate reaction occurs;product is formed and released

• • Transported extracellularly
• • They breakdown large food molecules or harmful chemicals
• • Cellulase,amylase,penicillinase

• • retained intracellularly and function there
• • Most enzymes are endoenzymes

• – Always present
• – Always produced in equal amounts or at equal rates
• – Enzymes involved in glucose metabolism

• – Not constantly present
• – Production is turned on(induced)or turned off(repressed) in response to changes in concentration of the substrate

• –Anabolic reactions to form covalent bonds
• – Between smaller substrate molecules
• – Require ATP
• – Release one molecule of water for each bond formed

• – Catabolic reactions that breakdown substrates into small molecules
• – Requires the input of water to break bonds

• • Temperature
• – Reduce activity
• – Denature
• • pH
• – Reduce activity
• – Denature
• • Substrate concentration
• – Reaches a saturation point

Competition for the active site. Inhibitor blocks the active site.

• • Allosteric enzymes
• • The binding by a substance to a site other than the active site which blocks the substrate from active substrate

• • Inhibits at the genetic level by controlling synthesis of key enzymes
• when too much product is being made, the product binds to another protein which binds to promoter and turns gene off

• Enzymes are made only when suitable substrates are present
• protein keeps geen off until a certain protein or sugar is present and binds to promoter and turns gene on causing it to start making the product. once enough product is made the protein or sugar breaks off and the gene turns off. this helps preserve energy.

consume energy

release energy

• – Always occur in pairs
• – There is an electron donor and electron acceptor which constitute a redox pair
• – Process salvages electrons and their energy
• – Released energy can be captured to phosphorylate ADP or another compound
• Na + e^- + Cl » Na⁺_oxidized Cl^- _reduced
• everytime you add on an e^- and H⁺ to a molecule it is reduced

• – Repeatedly accept and release electrons and hydrogen to facilitate the transfer of redox energy
• – Most carriers are coenzymes:
• • NAD+
• • FAD
• • NADP
• • coenzymeA
• • Compounds of the respiratory chain

• – Metabolic“currency”
• – Three part molecule consisting of
• • Adenine–anitrogenous base
• • Ribose–a 5-­‐carbonsugar
• • 3phosphate groups– ATP utilization and replenishment is a constant cycle in active cells
• – Removal of the terminal phosphate releases energy

• • Substrate-­ level phosphorylation – transfer of phosphate group from a phosphorylated compound(substrate) directly to ADP
• • Oxidative phosphorylation – series of redox reactions occurring during respiratory pathway
• • Photophosphorylation – ATP is formed utilizing the energy of sunlight

glycolysis, the Kreb’s cycle, respiratory chain

glycolysis, the TCA cycle, respiratory chain; molecular oxygen is not final electron acceptor

glycolysis, organic compounds are the final electron acceptors

glucose(6C) is oxidized and split into 2molecules of pyruvic acid (3C), NADH is generated

processes pyruvic acid and generates 3CO₂ molecules, three NADH and one FADH₂ are generated

accepts electrons from NADH and FADH; generates energy through sequential redox reactions called oxidative phosphorylation

• – Final processing of electrons and hydrogen and the major generator of ATP
• – Chain of redox carriers that receive electrons from reduced carriers (NADHandFADH2)
• – ETS shuttles electrons down the chain, energy is released and subsequently captured and used by ATP synthase complexes to produce ATP which is Oxidative phosphorylation

• as the electron transport carriers shuttle electrons, they actively pump hydrogen ions(protons) across the membrane setting up a gradient of hydrogen ions–proton motive force
• Hydrogen ions diffuse back through the ATP synthase complex causing it to rotate, causing a 3-dimensional change resulting in the production of ATP

• • Glycolysis
• – 2 NADH = 6 ATP
• – Plus 2 ATP
• • Pyruvate to Acetyl CoA
• – 2 NADH = 6 ATP
• • Kreb’s cycle
• – 6 NADH = 18 ATP
• – 2 FADH₂ = 4 ATP
• – Plus 2 ATP
• • Total = 38 ATP

• – Functions like aerobic respiration
• •Except it utilizes other electron acceptors rather than oxygen
• • Nitrate(NO3-­‐) and nitrite(NO2-­‐)
• • Used by anaerobes and facultative anaerobes

• – Incomplete oxidation of glucose or other carbohydrates in the absence of oxygen
• – Uses organic compounds as terminal electron acceptors
• – Yields a small amount of ATP
• – Recycles the NADH
• – Produces
• • Ethyl alcohol by yeasts acting on glucose
• • Formation of acid or acid and gas by bacteria

• the study of heredity
• • The science of genetics explores
• – Transmission of biological traits from parent to offspring
• – Expression and variation of those traits
• – Structure and function of genetic material
• – How this material changes

• sum total of genetic material of a cell(chromosomes+mitochondria/chloroplasts and/or plasmids)
• • Genome of cells – DNA
• • Genome of viruses – DNA or RNA
• – DNA complexed with protein constitutes the genetic material as chromosomes
• – Bacterial chromosomes are a single circular loop
• – Eukaryotic chromosomes are multiple and linear

• • Chromosome is subdivided into genes
• – The fundamental unit of heredity responsible for a given trait
• – Site on the chromosome that provides information for a certain cell function
• – Segment of DNA that contains the necessary code to make a protein or RNA molecule

• – Genes that code for proteins which are structural genes
• – Genes that code for RNA
• – Genes that control gene expression which are regulatory genes

All types of genes constitute the genetic makeup

The expression of the genotype creates observable traits

• Basic unit of DNA structure
• • Each nucleotide consists of 3 parts
• :– A 5 carbon sugar which is deoxyribose
• – A phosphate group
• – A nitrogenous base of adenine,guanine,thymine,cytosine
• • Nucleotides joined by phosphodiester bond
• • Two strands twisted into a doublehelix
• – They are anitiparallel

• • Helicase-­‐unwinds and unzips the double helix by binding hydrogen bonds
• • Topoimerase-­‐relieves supercoiling
• • Single Stranded Binding proteins
• • Primase-­‐makes a short RNA primer
• • DNA polymerase III-­‐adds nucleotides in a 5 ' to 3' directions
• • DNA polymerase I-­‐removes the primer and fills in the gaps
• • DNA polymerase II– has no known function
• • DNA Ligase-­‐joins the two ends after DNA Pol I

• DNA synthesis is always towards the replication fork
• – Leading strand is Synthesized continuously in 5'to3' direction
• – Lagging strand is synthesized 5'to 3' in short segments
• • Over all direction is 3' to 5'
• • Segments are called Okazaki Fragments

because each chromosome ends up with one new strand of DNA and one old strand.

• – Transcription
• • Is used to make RNA ( mRNA, tRNA, rRNA)
• – Translation
• • Is used to make protein from mRNA

• – Initiation
• • RNA polymerase finds the promoter before the gene and unwinds the DNA
• – Elongation
• starts 10 base pairs down from the promoter
• • NTPs are added in the 5’to 3‘ direction
• – Termination
• • Self termination with a stem loop
• • Enzyme dependent

• mRNA is composed of codons and the genetic code is universal, the genetic code is redundant
• • There is more than one codon per amino acid
• • There are 64 codons
• • Three are stop codons
• The tRNA is a stem loop structure and brings amino acids to the ribosome on the 3’end containing the anticodon
• The rRNA is used to make ribosomes

• – Initia-on
• • mRNA and ribosomes come together
• • Starts at the P site with the AUG start codon
• • First amino acid is methionine
• – Elongation
• • Add amino acids and form a peptide bond
• • Ribosome moves down them RNA to next codon
• – Termination
• • Ribosome reaches the stop codon
• – Causes them all to come apart

• – Transcription occurs in the nucleus
• – Translation occurs in the cytoplasm
• – Eukaryotic mRNA starts as premessenger RNA
• • Contains introns or intervening sequences
• • Contains exons that are expressed
• • Spliced by ribosomes
• • 3’PolyA tail is added
• • 5’methyl cap
• • Now it can leave the nucleus
• – Eukaryotic start codon is AUG, but it does not use formyl‐methionine

• – One or more pieces of DNA or RNA
• – Contains only genes needed for production of new viruses
• – Viral can have overlapping
• • Requires access to host cell’s genetics and metabolic machinery

• – Can perform transcription and translation at the same time
• – Poly cistronic mRNA
• • Genes are clustered together
• • In groups called operons
• • operons, a set of genes,all of which are regulated as a single unit

• operons, a set of genes,all of which are regulated as a single unit
• – Inducible operon
• •Operon is turned ON by substrate
• – Repressible operon
• • genes in a series are turned OFF by the product synthesized

• Lactose Operon : Inducible Operon
• Made of 3 segments:
• • Regulator–gene that codes for repressor
• • Control locus–composed of promoter and operator
• • Structural locus–made of 3 genes each coding for an enzyme needed to catabolize lactose
• –» β-­‐galactosidase–hydrolyzes lactose
• » permease–brings lactose across cell membrane
• » β-­‐galactosidasetransacetylase–uncertain function

• • Lac Operon is normally off
• – When there is no lactose
• • The repressor binds with the operator locus
• •Blocks transcription of downstream structural genes
• – Lactose turns the operon on
• • Lactose binds to repressor and changes its shape
• • Repressor comes off the operator
• • Now, RNA polymerase can bind to the promoter

• • Arginine Operon:Repressible
• – Normally on
• – Will be turned off when the product of the pathway is no longer required
• – When excess arginine is present it will bind to the repressor and changes its shape or the repressor binds to the operator and blocks arginine synthesis

• • Genetic recombination– Occurs when an organism acquires and expresses genes that originated in another organism
• • Horizontal genetic transfer:
• – Conjugation
• – Transformation
• – Transduction

• – Transfer of a plasmid
• • Plasmid is called a factor
• • Example is the F factor, which meands fertility factor
• – Done through pili
• – Can transfer factors for many different functions
• • Antibiotic resistance
• • Fertility
• • Enzymes

• – Can take up DNA from the environment
• – Chromosome fragments from alysed cell are accepted by a recipient cell
• – The genetic code of the DNA fragment is acquired by the recipient
• – Donor and recipient cells can be unrelated
• – Useful tool in recombinant DNA technology

Bacteriophage serves as a carrier of DNA from a donor cell to a recipient cell

• • Special DNA segments that have the capability of moving from one location in the genome to another – “jumping genes”
• • Cause rearrangement of the genetic material
• • Can move from one chromosome site to another, from a chromosome to a plasmid, or from a plasmid to a chromosome
• • Maybe beneficial or harmful

Use of an organism’s biochemical and metabolic pathways for industrial production

• – Intrinsic properties of DNA hold true even in a test tube
• • DNA heated from 90°C to 95°C
• – The two strands separate
• – The nucleotides can be identified, replicated,or transcribed
• • Slowly cooling the DNA
• –Allows complementary nucleotides to hydrogen bond
• – DNA will regain double-­stranded form

• • Restriction Endonucleases
• • Ligase
• • Reverse transcriptase

• – Recognize specific sequences of DNA
• – Break phosphodiester bonds between adjacent nucleotides
• – The enzymes can be used to cleave DNA at desired sites
• – Recognize and clip the DNA at palindrome base sequences
• – Used in the lab to cut DNA into smaller pieces
• – restriction fragments

• – Rejoins phosphate‐sugar bonds
• – Sticky ends‐cut by endonucleases
• – Used for final splicing of genes into plasmids and chromosomes

• – makes a DNA copy of RNA = cDNA
• – cDNA can be made from mRNA, tRNA,or rRNA
• – Provides a means of synthesizing eukaryotic genes

• • Gel Electrophoresis
• •Nucleic Acid Hybridization and Probes

• – Separates DNA fragments based on size
• – DNA samples are placed on soft agar gel and subjected to an electric current
• – Negative charge of molecule causes DNA to move toward positive pole
• – Rate of movement is dependent on size of fragment
• – larger fragments move more slowly
• – Fragments are stained for observation
• – Useful in characterizing DNA fragments and comparing for genetic similarities

• – Hybridization
• – Foundation for gene probes
• • Short fragments of DNA of a known sequence
• • Will base-­pair with a stretch of DNA with a complementary sequence,if one exists in the sample
• – Useful in detecting specific nucleotide sequences in unknown samples
• • DNA fragments are separated by electrophoresis
• • Southern blot is performed on the gel
• • Membrane from southern blot is incubated with probes
• • If probes are complimentary they will bind to DNA
• • This will isolate fragments from a mix of fragments and find specific gene sequences

• • DNA sequencing
• • PCR

• – Determining the actual order and type of bases for all types of DNA
• – Most common sequencing technique is Sange rtechnique
• • Test strands are denatured to serve as a template to synthesize complementary strands
• • Fragments are divided into tubes that contain:
• –Primers
• – DNA polymerase
• –All 4 nucleotides
• –dideoxynucleotide
• » A different one in each tube

• – Method to amplify DNA
• – Ingredients
• • DNA to be amplified
• • Taq Polymerase(fromThermusaquaticus)
• • Nucleotides
• – Repetitively cycled
• • Denaturation(94oC)
• • Priming(50to65oC)
• • Extension(72oC)
• – Each cycle doubles the number of copies for analysis

• • Recombinant DNA technology
• • Cloning Vectors

• – The intentional removal of genetic material from one organism and combining it with that of a different organism
• – Requirements to make RecDNA
• • Restriction endonucleases
• • Desired gene
• • Ligase
• • Vector(plasmidorvirus)
• • Cloning host(bacteriaoryeast)

• – Origin of replication is needed so it will be replicated
• – Vector must accept DNA of the desired size
• – Gene which confers drug resistance to their cloning host

• – Rapid overturn, fast growth rate
• – Can be grown in large quantities using ordinary culture methods
• – Nonpathogenic
• – Genome that is well delineated
• – Capable of accepting plasmid or bacteriophage vectors
• – Maintains foreign genes through multiple generation
• – Will secrete a high yield of proteins frome xpressed foreign genes
• • Producing a synthetic peptide using recombinant DNA technology and transformation

• • Enables large scale manufacturing of life‐saving hormones, enzymes, vaccines
• – Insulin for diabetes
• – Human growth hormone for dwarfism
• – Erythropoietin for anemia
• – Factor VIII for hemophilia
• – HBV vaccine

• • DNA Finger printing –Every individual has a unique sequence of DNA
• • Methods used include restriction endonucleases, electrophoresis, hybridization, and Southernblot
• • Types of analysis
• • SNP – single nucleotide polymorphism
• • Markers
• • VNTRs
• • Microsatellite polymorphisms

• • Identify hereditary relationships
• • Study inheritance of patterns of diseases
• • Study human evolution
• • Identify criminals or victims of disaster
• • Analysis of mitochondrial DNA isused to trace evolutionary origins
• • Micro array analysis – track the expression of thousands of genes

start with 6 c glucose which gains 2 phosphates from ATP which becomes ADP. this results in a 6c sugar phosphate which splits into 2 3c sugar phosphates. An electron and NAD+ is added to each 3c sugar phosphate this produces NADH then ADP picks up 2 phosphates becoming 2ATP and lose of 1 H20 molecule and formation of pyruvate. in aerobic respiration the pyruvate with Co2 is converted to acetyl CoA in the transition step before entering the Krebs cycle. in fermentation the pyruvate with Co2 is converted to acid and gas with no fnal electron receptor. Net results is 2NADH which equals 6 ATP and 2 ATP.

• Viral genes can be RNA or DNA and overlap
• Eukaryote have introns and exons, introns have to be taken out with rbosomes then the exons ned to be put back together wth a 5' prime cap and a 3' polyA tail end then leave the nucleus.
• Prokaryotes have operons, more than 1 gene per promoter and operator. It translates and transcribes more than 1 gene at a time into protiens simutaneously. During transcription into mRNA, each gene has its own start codon.
• [_promoter ][ _operator ]---[gene]--[gene]--[gene] »[ _mRNA ] [gene] [gene] [gene]
• -----------------operon---------------------- ==proteins= 1 ====2 -----3

Method to amplify DNA. Ingredients primer, DNA to be amplified, Taq (DNA polymerase that works when heated up), nucleotides. Ingredients are put in tube then heated making a double strand copy then heated up again to denature to separate strands. It is then cooled down making primer bind, heated up again so Taq works making 2 double stranded DNA. This repeated over and over again.

Ingredients needed restriction endonucleases (used to cut open plasmid), desired gene, ligase, plasmid (vector), bacteria or yeast (cloning host). Add cDNA or DNA from other pathogen and plasmid that has been cut open with restriction endonucleases plus contain the gene for antibodic resistance, then ligase which ligates genes together then add to bacteria or yeast (cloning host) and the plasmids will stick to bacteria. It is then heat shocked which cause transformation (bacteria suck up DNA). Plate bacteria on agar with antibodics where they will grow up (only bacteria with gene will grow). These bacteria are used to grow large batches of bacteria with new gene that can be used to make a synthetic peptide.

• – Recognize specific sequences of DNA
• – Break phosphodiester bonds between adjacent nucleotides
• – The enzymes can be used to cleave DNA at desired sites
• – Recognize and clip the DNA at palindrome base sequences
• – Used in the lab to cut DNA into smaller pieces called restriction fragments

An operon is a set of genes that are regulated as a single unit. When there is no lactose present, a repressor protein from a repressor gene(located elsewhere on the chromosome) binds to the control locus(promoter and operator) blocking the transcription of downsteam structural genes. when lactose enters the cell it turns on the operon by binding to the repressor causing it to change shape and fall off the operator. This allows RNA polymerase to bind to the promoter and start transcription. The repression of the genes prevents the production of enzymes for breaking down lactose when lactose isn't present. This helps conserve energy by only producing enzymes when necessary.

• 1.At the origin helicase unwinds the double helix asnd topimderase relieves supner coiling creating replicatieon forks at both of the replication bubble.
• 2. single stranded binding proteins stops hydrogen bonding.
• 3. leading strand is replicated in a continuous 5' to 3' strand by the laying down of primase making a RNA primer the DNA poly III adds nucleotides creating a new strand.
• 4. lagging strand is made in discontinuous 5' to 3' short segment because it goes from 3' to 5' and DNA has to replicated from 5' to 3'. primase lays RNA primer then DNA poly III lays nucleotides to 3' primer end in short segments called Okazaki fragments.
• 5. DNA I removes RNA primer and fills in gaps
• 6. DNA ligase joins DNA poly I and DNA poly III fragments together.
• 7. Process is terminated with either a stem loop or an enzyme causes RNA poly and RNA to come apart.

• Transcription is used to make RNA which occurs in the nucleus in 3 steps.
• 1. Initiation - RNA polymerase fnds the promoter before the gene and unwinds the DNA
• 2. Elongation - Starts 10 based pairs down from the promoter. NTPs are added in the 5' to 3' direction.
• 3. Termination - Occurs at the end of the gene by self termination with a stem loop or an enzyme dependent action. Self termination occurs when RNA start to H+ bond with itself creating a stem loop causing RNA polymerase to stop then come off, setting the RNA free. An enzyme dependent action occurs when an enzyme attaches to the RNA then goes up the RNA this makes the DNA, RNA polymerase, and RNA come apart.

• Translation turns amino acids into protiens occurs in the cytoplasm in 3 steps.
• 1. Initiation - The mRNA and ribosomes come together. Translation starts when a tRNA enters the P site and binds with the mRNA start codon AUG.
• 2. Elongation - The next tRNA