Lecture 11 - bacterial and virul genetic systems

Lecture 11 - bacterial and virul genetic systems -...

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Unformatted text preview: Lecture11 May24,2011 Chapter8 BacterialandViralGeneticSystems 1.Introductionofthebacterialsystem 2.Threemechanismsofbacteriagenetransfer 3.Viralsystem  Whyusebacterialandvirusesforgeneticstudy? Bacteria E.colicelldividesevery20 ­30minat37°C Cangroweasily Intesttubeswithliquidmedium  ­Formassivegrowth Onpetriplateswithgrowthmediumsuspendedinagar  ­ Formcolonyforisolateorcounting  ­ Colony:visibleclumpofgeneticallyidenticalcells  BacterialPhenotypes Wildtype:Prototrophicstrain Bacteriacanusethesimpleingredientstosynthesizeallthe compoundsthattheyneedforgrowth Minimalmedium:containsonlythenutrientsrequiredby prototrophicbacteria Mutantstrain:anxotrophicstrain Bacterialackoneormoreenzymesnecessaryforsynthesizing essentialmolecules Completemedium:containsallsubstancesandother supplementsrequiredbybacteriagrowthandreproduction  BacterialPhenotypes Bacterialphenotypesaredesignatedbythreeletters. +or designatethepresenceorabsenceofaspecificcharacter. Lac ­=inabilitytouselactose Lac+=abilitytouselactose rorsrepresentresistantorsusceptibletoacompound. Amp ­r=ampicillinresistant Amp ­s=ampicillinsusceptible BacterialGenome SinglecircularDNAmoleculeofmillionsbp Manybacteriapossessplasmid SmallcirculardsDNAofthousandsbp withfunctionalgenes From1to50copiesof plasmidpercell BacterialGenome Plasmidinbacteriahad ownoriginof replication. Theirreplicationis independentofthe chromosomalDNA. Havegenesprovide antibioticresistance Theyusuallydonot carryessentialgenes forbacterialsurvival. Extensivelyingenetic engineering.  BacterialGenetics Threemechanismsforgeneticmaterialexchanginginbacterium Conjugation Geneticmaterialpassesdirectlyfromonbacteriumto another Transformation BacteriumtakesupDNAfromgrowingmedium Transduction BacterialvirusescarryDNAfromonebacteriumtoanother Conjugation ProcessinwhichDNAistransferredfromadonorcelltoa recipientcellbycell ­to ­cellcontact. Allowsfortheexchangeofgeneticmaterialbetweendifferent bacterialstrains. Lederbergand  Y10strainlackthreeenzymesand  NOgrowthon minimalmedia. Y24lackanotherthreeenzymes  MixofY10andY24cangrowon minimalmedia Indicatedthatgeneticexchange occurredbetweentwobacterial strains. Didnotshowhowtheexchange occurred. MixY10andY24 Growthon minimalmedia.  ­tube experiment Membranepreventing directcontactbetween differentstrains IndicatedthatCell ­to ­Cell contactwasnecessaryfor conjugation Conjugation Conjugationinvolvesphysicalcontactviaapilus(conjugationtube) TheFplasmidinE.coliencodesthegenesneededtomakethepilus Ffactorsarelow ­copy ­numberplasmids Theyreplicateoncepercellcycleandsegregatetobothdaughtercellsduring celldivision. Fplasmids(100Kb)canoccasionally integrateintothebacterial chromosome(4.6Mb) ConjugationProcess TheFcantransferredfromdonor(F+)totherecipient(F ­)cellstrain ssDNAistransferred(rollingcirclereplication) BecomesdsDNAinrecipient,whichisthenF+ TransformationProcess AdonorDNAmolecule(freeDNAmolecule)istakenupfromthe surroundingmediumandincorporatedintothegenomeofa recipientcell. TheDNAthatistakenupcanalsobeaplasmid.Aslongastheplasmidcontainsan ori,itwillreplicateinthedonorcell TransformationProcess PieceofdonordsDNAbindstorecipientcellsurface. OnestrandofdonorDNAthenmovesacrossthecellmembrane. TheotherstrandofthedonorDNAisdegraded ThessDNAinrecipientpairswiththehostcellDNA. ThedonorDNAisrecombinedandintegratedintotherecipientDNA. » onemutantstrandandoneoriginalstrand. » Heteroduplexregion. Celldivisiongenerateatransformedcellandanon ­transformedcell Transduction DNAistransferredfromonebacterialcelltoanotherbya bacterialvirus=bacteriophage Bacteriophagetwoalternativelifecycles Make sure you know the difference between lytic and lysogenic TransductionProcess Bacterialgenesareincorporatedintoaphagecapsidbecauseof errorsthatoccurduringthevirallifecycle. Thebacteriophage,throughitsnormalinfectionprocess,injects thesegenesintoanotherbacteria,transferringgeneticmaterial fromonebacterialstraintoanother. Virus Caninfectbacteria,insects,plants,animals,andhumans Presentinlargenumbers evenatmodestmutationrates, variantswithalteredpropertieswillappear.Insomecasesthis leadstoseriousoutbreaksofinfectiousdiseases(HIV,ebola, hanta,WestNile,avianbirdflu,andH1N1)  Functionally,theyarereplicatingstructuresofDNAorRNA wrappedinaproteincoat Virusesarethereforenotconsideredlivingorganisms.They canbeconsideredgeneticparasites. Ofmajormedicalandeconomicimportance ClassificationofViruses DNAviruses ssanddsDNAviruses RNAviruses Plus ­strandRNA:canbetranslateddirectly Minus ­strandRNA:complementarystrandneedstobe synthesizedfortranslation Retroviruses RNAgenomesareconvertedtodsDNAbyreverse transcriptase. Integrateintohostgenome Transcriptionbyhosttranscriptionalmachinery  VirusGenome DNAVirus, X174wasthefirstviral genometobesequenced:5.3kbwith 9genes Singlestranded,circular;(+)strandin virion;complementarystrandismade inhost(E.coli),byhostenzymes!! dsDNAistranscribed. Overlappingreadingframes! VeryefficientuseofDNA Retrovirus Structure Replicationof retroviruses HIV HumanImmunodeficiencyVirus(HIV) ­causeacquiredimmune deficiencysyndrome(AIDS) Retroviruswithtwotypes:HIV ­1andHIV ­2;severalsubtypes thatwastransferredtohumanswhoateaninfected chimpanzee(HIV ­1)orSootyMangabeys(HIV ­2)andthatwas abletoreplicateinahumanhost Attacksoneofthebloodcells:T ­lymphocytes,involvedin fightingpathogens,verycentraltotheimmunefunction. Theviruscanspreadfrominfectedindividualsvianeedles, contactwithblood,bloodtransfusion,unprotectedsex http://www.avert.org/origins.htm ThelikelyoriginofHIV Anillustrationofhow recombinationandmutation, undercertaincircumstances,can leadtotheformationofnewviral strains. Evolutionatwork Otherexamplesincludebirdflu transmissionofSwineinfluenza H1N1(pigstohumans)orAvian influenzavirusAH5N1(birdsto humans) Transmissionofebolavirusvia fruitbats ...
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This note was uploaded on 12/16/2011 for the course AGR 3303 taught by Professor Gallio during the Summer '08 term at University of Florida.

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