NOCTI Biotechnology Review Notes

Standard 1-Work Habits

  • Professional work habits include knowledge of procedures, practicing safety, attention to detail, and organization.
  • Maintaining a work area involves cleaning, equipment calibration, glassware cleaning, proper equipment placement, tidiness in storage, up-to-date labeling, purchasing of general items, instrument maintenance, and internal OHS audits.
  • Effective communication, staying current, and using effective suppliers are essential.
  • Task management includes using organizers, mixing and labeling frequently used reagents, and efficient troubleshooting by identifying the problem, listing possible explanations, collecting data, eliminating possibilities, and identifying the root cause.

Teamwork

  • Reduces error, increases safety, and aligns individuals with a common mindset and goals.
  • Requires assigned roles, maximizing team member strengths, shared accountability, and open communication.

Standard 2-Knowledge of Biotechnology

  • Biotechnology utilizes biological systems to develop products, like using bacteria to produce insulin or genetically modifying crops.
  • Key historical milestones include animal domestication, fermentation for food production, and early vaccination efforts.
  • The National Institute of Health (NIH) funds and oversees research and regulates gene therapy and GMOs.
  • The Centers for Disease Control and Prevention (CDC) works to prevent health, safety, and security threats.
  • The Food and Drug Administration (FDA) ensures the safety and efficacy of drugs, biological products, medical devices, food, cosmetics, and radiation-emitting products.
  • The Occupational Safety and Health Administration (OSHA) ensures safe working conditions through standards, training, and education.

Key Scientific Discoveries

  • James Watson and Francis Crick discovered the double helix structure of DNA.
  • Rosalind Franklin created X-ray crystallography images used to determine DNA structure.
  • Francis Crick determined the central dogma of molecular biology: DNA to RNA to proteins.
  • Erwin Chargaff found that the amount of T equals A, and C equals G, leading to base pairing rules.
  • Alexander Fleming discovered penicillin, and Jonas Salk developed the polio vaccine.

Processes

  • Central dogma: DNA →\rightarrow mRNA →\rightarrow Protein.
  • Polymerase Chain Reaction (PCR):11. Denaturation (95°C), 22. Annealing (55°C), 33. Extension (72°C) using Taq polymerase to add dNTPs, doubling DNA each cycle.
  • Restriction endonucleases cut DNA at specific sequences.
  • Gel electrophoresis separates DNA fragments by length, with negatively charged DNA moving towards a positive charge.
    • Restriction Fragment Length Polymorphism (RFLP) shows DNA sequence differences.
    • SDS-PAGE denatures proteins and coats them with a negative charge for electrophoresis.

Drug Development Phases

  • Discovery and Development: Research begins in the lab.
  • Preclinical Research: Lab and animal testing for safety.
  • Clinical Research: Testing on people for safety and efficacy.
  • FDA Review: Examination of data for approval.
  • FDA Post-Market Safety Monitoring: Monitoring safety after public use.

Clinical Trial Phases

  • Phase I: Safety (20-80 participants, months).
  • Phase II: Efficacy (100-300 participants, up to 2 years).
  • Phase III: Safety, efficacy, dosing (1,000-3,000 participants, 1-4 years).
  • Phase IV: Long-term effectiveness, cost-effectiveness (thousands of participants, 1+ years).

Bioindustries & Bioremediation

  • Include biochemicals, biofuels, bioplastics, bioremedies, biomaterials, and textiles.
  • Bioremediation treats contaminated media by stimulating microorganisms to degrade pollutants.

Bioterrorism

  • Involves the intentional release of biological agents.

Biotechnology Careers

  • Biomedical Engineer: Design solutions for biology and medicine. Median Pay: 88,040.</li><li>Biochemist:Studychemicalpropertiesoflivingthings.MedianPay:88,040.</li> <li>Biochemist: Study chemical properties of living things. Median Pay:91,190.
  • Medical Scientist: Conduct clinical research to improve patient health. Median Pay: 82,090.</li><li>Biological/ClinicalTechnician:Collectsamplesandanalyzeresults.MedianPay:82,090.</li> <li>Biological/Clinical Technician: Collect samples and analyze results. Median Pay:51,770.
  • Microbiologist: Study viruses, bacteria, and the immune system. Median Pay: 69,960.</li><li>ProcessDevelopmentScientist:Overseemanufacturingprocessestoincreasequalityandefficiency.MedianPay:69,960.</li> <li>Process Development Scientist: Oversee manufacturing processes to increase quality and efficiency. Median Pay:94,739.
  • Biomanufacturing Specialists: Guarantee product purity, safety, and quality. Median Pay: 80,629.</li><li>BusinessDevelopmentManager:Providemarketanalysisandcompetitiveintelligence.MedianPay:80,629.</li> <li>Business Development Manager: Provide market analysis and competitive intelligence. Median Pay:113,769.
  • Director of Product Strategy/Commercialization: Responsible for commercialization strategy. Median Pay: 124,726.</li></ul><h3id="standard3laboratoryknowledgeandskills">Standard3−LaboratoryKnowledgeandSkills</h3><ul><li>EquipmentValidation:Verifiesinstrumentperformancemeetsspecifications,e.g.,calibratingbalancesormicropipettes.</li></ul><h3id="commonlabequipment">CommonLabEquipment</h3><ul><li>Safetyglasses:Protecteyesfromchemicals,sharpobjects,andheat.</li><li>Erlenmeyerflask:Usedtoreducespillsandaidpouring;approx.volumes.</li><li>Beaker:Transfers/containsliquids,mixessolutions.</li><li>Graduatedcylinder:Measuresliquidvolumes.</li><li>Pipette:Transfersspecificliquidvolumes.</li><li>Inoculatingloop:Transfersmicroorganisms,sterilizablebyflame.</li><li>Bunsenburner:Alaboratoryheatsource;sterilizestools.</li><li>Testtube:Containsspecificvolumesofliquids.</li><li>Gloves:Preventabsorptionofharmfulmaterialsintotheskin.</li><li>Pipettebulb:Usedwithpipettestotransferliquids.</li><li>Dropperbottle:Transferssmallliquidvolumes.</li><li>Petridish:Growsculturesofmicroorganisms.</li><li>Serologicalpipette:Transfersliquidvolumesusingaseptictechnique.</li><li>Micropipette:Transfersspecificliquidvolumes;usesdisposabletips.</li><li>Vortexmixer:Vortexesliquids.</li><li>Fumehood:Mechanicallyventedareaforvolatilechemicals.</li><li>Incubator:Providesatemperature−regulatedareaforgrowingmicroorganisms.</li><li>Thermocycler:CyclestemperaturesforPCR.</li><li>Horizontalelectrophoresis:SeparatesDNAfragmentsusingagarosegels.</li><li>PAGE(verticalelectrophoresis):Separatesproteinsusingpolyacrylamidegels.</li><li>Centrifuge:Separatessubstancesbydensity.</li><li>Autoclave:Sterilizesinstrumentswithhighheatandpressure(121°C,15psi,15−20min).</li><li>Microscopeobjectives:Varymagnification(e.g.,4X,10X,40X,100X).<ul><li>Totalmagnification:(ocular)X(objective)e.g.,10X40=400X.FineFocusforspecimenparts;CoarseFocususedfirst.</li></ul></li><li>Diaphragm:Adjustslightpassingthroughthestagetoilluminatethespecimen.</li></ul><h3id="cellwalls">CellWalls</h3><ul><li>Gram(+)cellwalls:thickpeptidoglycanlayer,teichoicandlipoteichoicacids.Gram(−)cellwalls:thinpeptidoglycanlayer,outermembranewithlipopolysaccharides,porins.</li></ul><h3id="computerskills">ComputerSkills</h3><ul><li>Cutting/copying/pastingstoresdataintheclipboard.</li><li>Excel:Use“=”beforeenteringaformula,e.g.,=(A1+B1+C1)/3foraverage.</li></ul><h3id="emailetiquette">EmailEtiquette</h3><ul><li>Useproperemailetiquette:formalsubject,introduction,text(completesentences),salutation,andattachments.</li></ul><h3id="preparingasolution">PreparingaSolution</h3><ul><li>Toprepareasolution,youmustconsidermolarmassanddesiredmolarity.</li><li>Example:Toprepare1literof1.00MNaCl,weighout58.44gNaCl,dissolveindistilledwater.</li><li>Fora10</ul><h3id="standardcurve">StandardCurve</h3><ul><li>Astandardcurvegraphsknownpropertiestodeterminepropertiesofunknownsamples.</li><li>Spectrophotometermeasureslightabsorbance,detectingwavelengthsfrom400−800nm.</li></ul><h3id="spectrophotometry">Spectrophotometry</h3><ul><li>Steps:Warmupunit,setwavelength,useblankcuvettefor0</ul><h3id="serialdilutions">SerialDilutions</h3><ul><li>Usedtocalculateunknownconcentrations.</li></ul><h3id="experimentalvariables">ExperimentalVariables</h3><ul><li>Independentvariable:manipulated.</li><li>Dependentvariable:measured.</li><li>Controlledvariable:keptconstant.</li><li>Controlgroup:notexposedtoindependentvariable.</li><li>Experimentalgroup:exposedtoindependentvariable.</li></ul><h3id="dnamanipulationtechnologies">DNAManipulationTechnologies</h3><ul><li>RecombinantDNA:vectoranddonorDNAdigestedwithrestrictionenzyme,overhangsalign,DNAligasesealstheoverhangs.</li><li>DNAExtraction:detergentslyseplasmaandnuclearmembranes,enzymesbreakdowncellcontaminants.</li></ul><h3id="dnatransfer">DNATransfer</h3><ul><li>Transformation:DNApickedupfromsurroundings.</li><li>Transduction:DNAtransferredbyavector.</li><li>Conjugation:DNAtransferredbetweenbacteriaviapili.</li></ul><h3id="error">Error</h3><ul><li>Understandingaccuracyandprecision:<ul><li>Lowaccuracywithlowprecision.</li><li>Lowaccuracywithhighprecision.</li><li>Highaccuracywithlowprecision.</li><li>Highaccuracywithhighprecision.</li></ul></li></ul><h3id="conversions">Conversions</h3><ul><li>Conversionstoremember:<ul><li>124,726.</li> </ul> <h3 id="standard3laboratoryknowledgeandskills">Standard 3-Laboratory Knowledge and Skills</h3> <ul> <li>Equipment Validation: Verifies instrument performance meets specifications, e.g., calibrating balances or micropipettes.</li> </ul> <h3 id="commonlabequipment">Common Lab Equipment</h3> <ul> <li>Safety glasses: Protect eyes from chemicals, sharp objects, and heat.</li> <li>Erlenmeyer flask: Used to reduce spills and aid pouring; approx. volumes.</li> <li>Beaker: Transfers/contains liquids, mixes solutions.</li> <li>Graduated cylinder: Measures liquid volumes.</li> <li>Pipette: Transfers specific liquid volumes.</li> <li>Inoculating loop: Transfers microorganisms, sterilizable by flame.</li> <li>Bunsen burner: A laboratory heat source; sterilizes tools.</li> <li>Test tube: Contains specific volumes of liquids.</li> <li>Gloves: Prevent absorption of harmful materials into the skin.</li> <li>Pipette bulb: Used with pipettes to transfer liquids.</li> <li>Dropper bottle: Transfers small liquid volumes.</li> <li>Petri dish: Grows cultures of microorganisms.</li> <li>Serological pipette: Transfers liquid volumes using aseptic technique.</li> <li>Micropipette: Transfers specific liquid volumes; uses disposable tips.</li> <li>Vortex mixer: Vortexes liquids.</li> <li>Fume hood: Mechanically vented area for volatile chemicals.</li> <li>Incubator: Provides a temperature-regulated area for growing microorganisms.</li> <li>Thermocycler: Cycles temperatures for PCR.</li> <li>Horizontal electrophoresis: Separates DNA fragments using agarose gels.</li> <li>PAGE (vertical electrophoresis): Separates proteins using polyacrylamide gels.</li> <li>Centrifuge: Separates substances by density.</li> <li>Autoclave: Sterilizes instruments with high heat and pressure (121°C, 15psi, 15-20 min).</li> <li>Microscope objectives: Vary magnification (e.g., 4X, 10X, 40X, 100X).<ul> <li>Total magnification: (ocular) X (objective) e.g., 10 X 40 = 400X. Fine Focus for specimen parts; Coarse Focus used first.</li></ul></li> <li>Diaphragm: Adjusts light passing through the stage to illuminate the specimen.</li> </ul> <h3 id="cellwalls">Cell Walls</h3> <ul> <li>Gram (+) cell walls: thick peptidoglycan layer, teichoic and lipoteichoic acids. Gram (-) cell walls: thin peptidoglycan layer, outer membrane with lipopolysaccharides, porins.</li> </ul> <h3 id="computerskills">Computer Skills</h3> <ul> <li>Cutting/copying/pasting stores data in the clipboard.</li> <li>Excel: Use “=” before entering a formula, e.g., =(A1+B1+C1)/3 for average.</li> </ul> <h3 id="emailetiquette">Email Etiquette</h3> <ul> <li>Use proper email etiquette: formal subject, introduction, text (complete sentences), salutation, and attachments.</li> </ul> <h3 id="preparingasolution">Preparing a Solution</h3> <ul> <li>To prepare a solution, you must consider molar mass and desired molarity.</li> <li>Example: To prepare 1 liter of 1.00 M NaCl, weigh out 58.44 g NaCl, dissolve in distilled water.</li> <li>For a 10% (w/w) NaCl solution, mass out 10 g NaCl, dissolve in water, and bring to 100 mL.</li> </ul> <h3 id="standardcurve">Standard Curve</h3> <ul> <li>A standard curve graphs known properties to determine properties of unknown samples.</li> <li>Spectrophotometer measures light absorbance, detecting wavelengths from 400-800 nm.</li> </ul> <h3 id="spectrophotometry">Spectrophotometry</h3> <ul> <li>Steps: Warm up unit, set wavelength, use blank cuvette for 0% absorbance/100% transmittance, insert sample, record readings.</li> </ul> <h3 id="serialdilutions">Serial Dilutions</h3> <ul> <li>Used to calculate unknown concentrations.</li> </ul> <h3 id="experimentalvariables">Experimental Variables</h3> <ul> <li>Independent variable: manipulated.</li> <li>Dependent variable: measured.</li> <li>Controlled variable: kept constant.</li> <li>Control group: not exposed to independent variable.</li> <li>Experimental group: exposed to independent variable.</li> </ul> <h3 id="dnamanipulationtechnologies">DNA Manipulation Technologies</h3> <ul> <li>Recombinant DNA: vector and donor DNA digested with restriction enzyme, overhangs align, DNA ligase seals the overhangs.</li> <li>DNA Extraction: detergents lyse plasma and nuclear membranes, enzymes break down cell contaminants.</li> </ul> <h3 id="dnatransfer">DNA Transfer</h3> <ul> <li>Transformation: DNA picked up from surroundings.</li> <li>Transduction: DNA transferred by a vector.</li> <li>Conjugation: DNA transferred between bacteria via pili.</li> </ul> <h3 id="error">Error</h3> <ul> <li>Understanding accuracy and precision:<ul> <li>Low accuracy with low precision.</li> <li>Low accuracy with high precision.</li> <li>High accuracy with low precision.</li> <li>High accuracy with high precision.</li></ul></li> </ul> <h3 id="conversions">Conversions</h3> <ul> <li>Conversions to remember:<ul> <li>1000 \mu = 1 milli</li><li>milli</li> <li>1000milli=milli =1gram,liter,meter</li><li>gram, liter, meter</li> <li>1000 \mu l = 1ml</li><li>ml</li> <li>1000ml=ml =1L$$

Calculating Averages

  • Calculating averages, also known as the mean, is the sum of numbers divided by the number of data points.

Aseptic Technique

  • Aseptic technique ensures no contamination of specimens and prevents the spread of infectious agents.
  • Guidelines: PPE, no food/drink, tie hair back, disinfect surfaces, wash hands, and sterilize loops.
  • Isolation streaking dilutes a sample to produce individual colonies.

Aerobic vs Anaerobic

  • Aerobic: with oxygen (e.g., cellular respiration).
  • Anaerobic: without oxygen (e.g., glycolysis, fermentation); anaerobic bacteria grown in sealed tubes with reducing agent.

cDNA

  • Making complementary DNA: isolate mRNA, add reverse transcriptase, digest mRNA, add DNA polymerase.

Standard 4-Ethics

  • HIPAA: health information portability and accountability act protects individually identifiable health information.
  • Genetic Ethics: Ethical concerns include insurance and employment discrimination, and companies profiting from genetic data.
  • Stem cells: Ethical debates surround embryonic stem cell use vs. adult stem cells.
  • Plagiarism: any use of information without citation or permission.

Types of Plagarism

  • Complete plagiarism, Source-based plagiarism, Direct plagiarism, self Plagiarism, paraphrasing Plagiarism, Mosaic plagiarism, Accidental plagiarism, Inaccurate authorship

Standard 5-Safety

  • The health, fire, specific and instability hazards are represented by a blue, red, white and yellow diamond, respectively.
  • Handwashing is the most effective way to prevent the spread of disease.
  • Spills: use PPE, spill kits, report injuries, restrict access, eliminate ignition sources.
  • Emergency Planning and Community Right-to-Know Act (EPCRA) helps communities plan for chemical emergencies.
  • Blood Spills: Prevent contact, contain and remove, disinfect, dispose in biohazard bags, sanitize area.
  • Copyright protects original works; a trademark protects names, slogans, or logos; a patent protects new inventions.
  • Personal protective equipment (PPE) minimizes exposure to hazards.

Fire Safety symbols

  • Fire extinguisher, Flammable, No Open Flames, Oxidizing Material, Fire Blanket, Fire Hose

Disease

  • Chronic, acute, ideopathic, hereditary.

Standard 6-Document Your Work

  • Use a laboratory notebook to document your work and to create a table of contents.
  • Date everything.
  • If a mistake is made, cross out the mistake with a single line and make the correction.

Standard 7-Equipment and Instrumentation

  • Acids are solutions with a pH less than 7. Add/liberate hydrogen ions (H+) in solutions. Remove hydroxide ions (OH-) from solutions. Can be used to lower the pH of a solution.
  • Bases are solutions with a pH greater than 7. Bases produce hydroxide ions (OH-). Can be used to raise the pH of a solution.

Chromatography

  • Uses a matrix to separate a mixture into its different components. The matrix can be a gel (column chromatography-we used this to purify GFP) or paper (you might have used this to separate photosynthetic pigments. Planar and thin layer chromatography are other types)

Growing Pathogens

  • Grown best at 37 degrees C, since this is normal human body temperature.

Valumetric Glassware

  • includes graduated cylinders, beakers, volumetric pipets, burets and volumetric flasks.
  • Graduated cylinders, beakers and Erlenmeyer flasks have less accuracy than volumetric glassware.

Balances

  • Be sure you understand how to use a balance before taking mass measurements.
  • Never place a sample directly on the balance.

Autoclave

  • provides a physical method for disinfection and sterilization, operating at high temperature and pressure to kill microorganisms and spores.

PH Meter

  • pH meter must be calibrated before you start.
  • Start at pH 4, then pH 7.

Biological Safety Cabinet

  • Performed laboratory procedures that could create airborne biohazards in a biological safety cabinet

Gas Chromatography

  • the mixture of interest is vaporized and carried through a stationary phase (usually a metal or glass separation column) with an inert gas, usually nitrogen or helium.

Liquid Chromatography

  • the mixture of interest is dissolved in a liquid and passed through a solid stationary phase, which is often made of a silica material.

Bradford Protein Assay

  • Used to determine concentration of proteins in a solution