med 4耶鲁大学公开课生物医学工程Word下载.docx
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med 4耶鲁大学公开课生物医学工程Word下载.docx
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regoingtocontinuetalkingtodayaboutDNA.Inparticular,we'
regoingtofocustodayonsortofhowtomanipulateanduseDNAinsomeapplications,andthisisahugeareaofscienceandtechnology.Youknowthis,youcan-it'
shardtopickupanewspaperoranewsmagazinewithouthearingsomenewapplicationofDNAtechnology.WhatI'
mgoingtodoisfocusonacouplebasicthingsthatturnouttobereallyimportantforgeneralapplications,andthenI'
lltalknotintoomuchdetailaboutafewapplicationsofDNAthatyou'
reprobablyfamiliarwithtotrytogiveyouaframeworktohangthison.
This,IthinkChapter3,describesfairlywellthethingswe'
lltalkabouttoday.You'
veprobablyalreadynoticedthatthere'
ssomematerialthat'
ssetasidefromthetextinboxes,andIencourageyoutoparticularlylookatthoseboxesforthischapter.There'
soneonDNAfingerprinting,forexample,thatgivesyoualittlebitmoredetailaboutthatspecifictechnique;
anotheroneonproductionoftherapeuticproteins.I'
lltalkaboutthesebothabittodaybutIencourageyoutoreadthoseformoreinformation.
Iwanttostartwhereweleftofflasttime.WetalkedaboutthestructureofDNA,howitworksintermsofaphysicalchemicalmodeloftheDNAmolecules.Wetalkedaboutbasepairingandhowthatleadstothisprocessofhybridizationorveryspecificmatchingbetweencomplimentarystrands.WetalkedverysuperficiallyaboutthebiologicalprocessgoingfromDNAtoprotein,sotheprocessoftranscription,RNAprocessing,andtranslationtoproduceaprotein.Iendedwiththispicturethatshowsyoualittlebitaboutcontrolofgeneexpression.Theimportantconceptisthat,whileeverycellinyourbodyhasthecapabilityofmakingalltheproteinsthatareneededthroughoutyourbody,noteverycellisdoingthatatanygiventime.Onlycertaingenesarebeingexpressedandit'
sthefamilyofgenesthatarebeingexpressedinacell,likewisethefamilythatarenotbeingexpressed,thatdetermineswhatacellislike,howitfunctions.What'
scalledthephenotypeofacellandwe'
lltalkmoreaboutthisnextweekwhenwestarttalkingaboutcellsandalittlebitaboutcellphysiology.
Therearemultiplemechanismsthatacellcanusetodecidewhichgenesitisexpressingatanyonetimeandwhichonesarenotexpressed.Ishowedyouthisinthispicturehereandthoselevelsofcontrolcanbeattheleveloftranscription.TherearemoleculesincellsthatgivetheDNAthesignalthatit'
stimetotranscribeandexpressagene,thosearecalledtranscriptionfactors,we'
lltalkaboutthemabitlater.There'
sinterferingwithRNAprocessing,andI'
mgoingtotalkaboutthatinthenextcoupleofslidesbecausethere'
sacoupleofnewmethodsfor-orpotentiallyinterferingwithgeneexpressioninlivinganimalsthathavebeendevelopedbasedonchanging-interferingwithDNAtranscriptionandtheabilityofmessengerRNAtobetranslated.Youcouldinterfereattheselaterlevelsaswell,forexample,byaugmentingRNAdegradation.IfthemessengerRNAforproteinisnotpresentinacell,thenthatcan'
tbetranslated,obviously,andtheproteincan'
tbemade.
ThesetwonewmedicaltherapiesthatImentionedarebasedoninterferingwiththebiologyofRNA,andoneisolderthantheotherandtheolderoneiscalledanti-sensetherapy.Whenyouthinkaboutageneoratranscript,themessengerRNAcopyofagene,youknowthatforeverysequenceofanucleicacidthere'
sacomplimentarysequence.Nowofthetwocomplimentarysequences,oneofthemencodesthegene.Oneofthemhastherightsequenceofcodonstospecifytheaminoacidsequenceoftheprotein,andtheotheronehasacomplimentarysequence.Youknowfromourdiscussionlasttimethatthesetwocomplimentarystrandsarenotmirrorimagesofoneanother,they'
renotidentical,they'
recomplimentary.Theyfaceintheoppositedirectionandyoucouldpredictthepropertiesofonefrombasepairingrulesoftheotherbutthey'
renotthesame.Oneofthestrandsencodestheprotein,encodesfortheprotein,theotherdoesnot.Theonethatdoesiscalledthesensestrandandtheotheroneiscalledtheanti-sensestrand,antimeaningit'
sthecomplimentanditwillhybridizetothesense.
Whatifyouknewwhatthesequenceofagenewas?
Agene,let'
ssayit'
sthegeneforinsulin.I'
llusethatoneastheexamplebecauseit'
safamiliaronetomostpeopleandyouknowthattheproteininsulinismadeonlyinyourpancreasandcertaincellsofthepancreas.SothatmeansthosecellsarecontinuallymakingmessengerRNAandthatmessengerRNAisbeingconvertedintoprotein.Well,whatifyouknewthesequenceforthemessengerRNAthatmadeinsulinandyoudesignedanothersinglestrandedDNAorRNAmoleculethatwastheexactopposite,ortheexactcompliment,Ishouldsay,ofthatstrand?
Soyoumadesomehowananti-sensepolynucleotidetotheinsulingeneorsomefractionoftheinsulingene.
Well,thatanti-sensestrandisshownhereastheredandthecellisnaturallymakingtheblueorsensestrandofmessengerRNAforaparticularprotein.Ifsomehowyoucouldtakeyouranti-sensemoleculesthatyou'
vemadeandyoucouldgetthemintocells,thenbythisprocessofhybridizationtheywouldnaturallyformapairlikethis.Theywouldnaturallyhybridizeandformaduplex,oradoublestrandednucleicacid.Whenit'
sdoublestrandedthisgenecan'
tbetranslated,becauseyouhavetohavethesinglestrandinorderforthetransferRNAtobindandforthisprocessoftranslationtotakeplace.Whatyoucoulddothen,ifyoucoulddelivertheseredcoloredmoleculeshereisyoucouldstopspecificallytheexpressionofthisparticulargeneintheseparticularcells.
Now,whatarethechallengesthere?
You'
vegottobeabletomakethisstuffandyou'
vegottobeabletomakeitinlargequantitiesandwe'
lltalkabouthowtomakenucleicacidsinlargequantitiesalittlebitlaterinthelecture.You'
vealsogottogetitintothecell.Itturnsoutthatgettinglargemoleculeslikethis,particularlylargechargedmoleculeslikenucleicacids,insideofcellsisnotsoeasy.We'
lltalkaboutthatalittlebitlateraswell.Infact,we'
lltalkaboutthatconceptthroughoutthecoursebecauseoneofthebigchallengesofmakingthesesortofnewbiologicaltherapiesworkinpeopletotreatdiseasesisgettingtherightmoleculesintotherightcells,attherightperiodoftime.
NowIgaveyoutheexampleofinsulinandyouprobablywouldn'
twanttostopinsulinproduction.Thatmightnotbeagoodthingtodo.Whatifthisisagenethat'
scausingacelltobecancerous?
Itwasagenethatwascausingacelltobemalignantandtodividewithoutcontrol,forexample.Thenyoucouldimagineblockinggeneexpressionwouldbeatherapy.
Student:
[inaudible]
Youwouldn'
twanttostopinsulin,forexample.Infactwhatyoumightprefertodoisstartinsulinproductionandwe'
lltalkaboutwaystodothatinjustaminute.Thesearewaystostopagenefrombeingexpressed,andthereturnsoutthere'
slotofapplicationsinthat,lotsofdiseasesresultfromtheunwantedexpressionofcertainkindsofgenesandcancerisprobablythebestexampleofthat,buttherearemany.
AnewerversionofthisthatworksinasimilarwaybutadifferentwayiscalledRNAinterference,anditturnsoutthatthisisanaturalmechanismthatcellshave.It'
samechanismthattheyhaveevolvedinordertopreventforeigngenesfromenteringacellandbeingexpressed.YouhavemechanismsinsideyournaturalmechanismsinsideyourcellthatallowthecellstodegradeunwantedRNAsequences.ThosemechanismsarecalledRNAinterference.Youmighthaveheardaboutthisbecauseit'
sbeenquiteanactiveareaofscience.
ItturnsouttoactivateRNAinterference,youdeliverdoublestrandedRNA.CertaindoublestrandedRNAsequenceswillcauseinthecellaprocessofdegradationofveryspecificRNAsequence.Thisinvolvesmechanismsthatarestillbeingunderstood,butifyou'
vestudiedsomebiologyorreadaboutthisyou'
veheardabouttheproteincomplexcalledDicer.DicerisaninternalcellularmechanismfordegradingRNA'
s.YoumighthavealsoheardabouttheRISCcomplex,ortheRNAsilencingcomplex,andthesearethebiologicalmechanismsthatareinvolvedhereandonlyshownbyorangearrowsonthisslide.TheendresultisyoucandesignnowveryspecificdoublestrandedRNAsequences,thatwhendeliveredintocellsagainwillactivatethisprocessofnaturaldegradationofanexistingmessengerRNA.Ofcourse,ifyoudegradethemessengerRNAatarapidratethanyou'
llstopexpressionofthecells.
Nowthenicethingaboutthisisthatthedegradationmechanismsseemtopersistforsomeperiodoftime,beyondthetimeatwhichyoudeliverthedoublestrandedRNA;
whereas,obviously,thismechanismhereisonlygoingtoexistforaslongastheanti-sensesequenceispresent.Sothismightbealongerlasting,morepermanentformofeliminationofexpressionofaparticulargene.
Ijustwantedtointroducethoseconceptsbecauseyou'
vereadaboutthem;
we'
llbetalkingmoreaboutRNAinterferenceinparticularaswegoonthroughthecourse.TherestofthetimeIwanttotalkaboutexpressionofgenes,ofnewgenes.Takingforeigngenes,genesthataren'
tnaturallyexpressedormightnotevenexistinsideacellandputtingthemthereandputtingthemthereinawaywheretheywork,andbyworkmeaningthegenegetsexpressedortranslatedintoaprotein.
I'
mgoingtostartbytalkingaboutaveryspecificandinterestingformofdoublestr
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