IntroductiontoBiologicalScience生物科技概論By:ChungW.Wu,M.S.,R.PhExt:---52131AncientBiologicalScience1.FoodPreparationBeer,Cheese2.FermentationMicrobialFood--------------Perfume3.AlcoholFermentationDistillationAlcoholConcentration21stWorldWarGermanyGlucose---------GlycerolBritishSugar---------------------Acetone+ButanolClostridiumacetobutylicumCarbonDioxideHydrogenGas:Explosive32ndWorldWar1940FlemingPenicilliumnotatumPenicillin:BroadSpectrumAntibiotic2mg/lPenicilliumchrysogenumUV,X-Ray—Mutation20mg/lStreptomycesgriseusStretomycin4GeneticEngineeringDNARecombinationRecombinantDeoxyribonucleicAcidChemicalComposition1.Phosphate2.Pentose3.NitrogenousBaseAdenineThymineCytosineGuanine5PrinciplesofDNARecombinantDNASectioncan’tReplicatebyitself.1.CloningVector

具有複製起始能力之載體2.Plasmid

能獨(dú)立複製之環(huán)狀DNA分子3.RestrictionEndonuclease(DNase)RecognitionSite4.StickyEnds6ExpressionofClonedGenes1.Transcription2.Translation3.Post-Translation

7TheMolecularBasisofHeredity:DNA&RNA

NuclearComponentsDNA→Transcription→RNA→Translation→AminoAcids→Polypeptide→ProteinsRNA:SingleStrandAdenine,Uracil,Cytosine,GuaninemRNAtRNArRNA8

DNATranscription1.DuplexDNASeperation→2SingleDNAStrandsExposingtheNitrogenousBaseSequencesOnlyOneofthe2DNAStrands,WhichServesasATemplate,orPattern(Gene),fortheSynthesisofRNA.9

2.ComplementaryTTTTCTAAACGTAAAAGATTTGCAmRNAUUUUCUAAACGU4x4x4=6464Possible3-LetterWords10TranslationProteinSynthesis20WordsintheFormof20CommonA.A.EachTripletCodesforanAminoAcids11Ribosome:SynthesisofProteinCellularOrganellethatServesastheMeetingPlaceformRNAandtRNAthatcarryA.A.toFormPeptideUUUUCUAAACGUPhe--Ser--Lys—Arg12WhatisGeneCloning?InsertDNAFragmenttoCircleDNAMolecule------ChimeraRecombinantDNA13PCR:PolymeraseChainReaction1995KaryMullis1.DenaturationofDNA94C2.AnnealingofthePrimers(Oligonucleotides)50---60C3.SynthesisofNewDNA74C4.RenaturationofDNARepeattheCycle25-30Times14GeneIsolationbyCloning1.DNAFragments2.ConstructRecombinantDNA3.IntroduceintoBacteria4.PlateOut5.EachColonycontainsMultipleCopiesofjustone15GeneIsolationbyPCR1.PolymeraseChainReaction2.XSeveralMillion16TheCommercialExploitationofMicroorganism1.BacillusthuringiensisNaturalInsecticideThuringiensisGene

選殖共生於玉米根部可以防治玉米害蟲172.AureobasidiumpullulansPolysaccharidesPullulan183.AcinetobactercalcoaccticusPolysaccharidesEmulsan19BiotransformationBioconversion1.Steroid------Cortisone37stepsUSD200dollars/g11stepsUSD6dollars/g20Glucose—Sorbitol-----Sorbose-----VitaminCGlucose—Sorbitol4chemicalstepsBiotransformation1chemicalstep21Examples:1.PhenylpyruvicAcid-----Phenylalanine2.Cinnamicacid-----PhenylalaninePhe+AsparticAcid----Aspartame160XSweetness22BiotransformationGrowingCell—休眠CellorKilledCell23Enzymes:1.Glucose---FructoseGlucoseIsomeraseArthrobactersp.Bacillussp.242.Penicillin----6-APCPenicillinAmidaseEscherichiacoliThroughDNARecombinant,theyieldOfEnzyme:20g/l253.Chymosin

小牛胃中凝乳素Milk---------Cheese(withFlavor)26Protein:1.GrowthHormoneFishCow:secretionofmilkHuman272.Insulin3.Interferon4.VaccineGMPRequirement28CellandEnzymeImmobilizationMethodsofImmobilization1.PhysicalAdsorptionPhysicalBindingCellorEnzyme以吸附方式被固定裝置在管柱中容易吸附也容易脫離尤其在基質(zhì)添加時CellorEnzyme容易Autolyse造成Activity喪失292.Crosslinking

以GlutaraldehydeToluenediisocyanate

進(jìn)行交聯(lián)形成不溶解性間質(zhì)但CellorEnzymeActivity仍會流失303.EntrapmentLatticeCollagenGelatin,AgarPolyacrylamideCelluloseToformPolymerMatrix31ChoiceofImmobilizationMethodsStabilityCostRegenerabilityCommercialization32MonoclonalAntibodyPolyclonalAntibodyEpitopes抗原決定位Chimeric嵌人化Humanized擬人化33TotalHumanMonoclonalAntibodyMakingAHybridomaAntigen----AnimalImmunizationBloodCollectionSpeenofAnimalRBC&Lymphocyte34ApplicationofMonoclonalAntibody1.DiagnosticKit2.增加敏感度與再現(xiàn)性3.Therapy35Examples:1.HepatitisVirus2.Polyoma3.Adenovirus36Isolation&PurificationofMABsImmunoaffinityChromatography1.Specificity2.Elution----SharpPeak3.IndependentTargetProtein37ImmunotoxinsasTherapeuticRicin----A,BPolypeptide----S----S----A,BChainEnterCellPlasmRibosome---Dysfunction---Cell’sDeath38Immunotoxin:MABscombinewithA,BChain----AttackCancerCellStableinHumanBody392.MABs+Radioiso