 in a cell which has been infected with a second, helper virus (or in a cell in which helper functions are present) proceeds as follows (see FIG. 1). Adsorption of AAV to a host cell is followed by inserting the single-stranded viral genome in a process generally known in the art as “transduction”. In the presence of certain host cell functions related to replication (such as DNA polymerases), the incoming single-stranded viral genome is converted to a double-stranded replicative form. See FIG. 2. Initiation of this single-strand to double-strand (SS→DS) conversion is believed to involve formation of a hairpin structure by sequences within the AAV ITR, which generates a template-primer structure from which initiation of DNA replication can proceed. The product of this SS→DS conversion, the replicative form (RF), is a self-complementary double-stranded molecule that is covalently closed at one end (the end at which replication was initiated). See FIG. 3. The RF is thus a double-stranded molecule having the same sequence complexity, but approximately twice the molecular weight, of the incoming AAV genome (i.e., for a native genome of approximately 4.7 kilobases, the RF will have a molecular weight corresponding to 4.7 kilobase pairs). Although formation of a terminal hairpin to prime replication is believed to occur rapidly, the extension of this hairpin to form the double-stranded RF is postulated to be one of the rate-limiting steps in AAV replication. This process of RF generation can occur in the absence of helper function but is believed to be enhanced by helper function. See Carter, B. et al. (1990) vol. 1, pp. 169-226 and 255-282. Cells that are capable of producing AAV progeny are generally considered by those skilled in the art as “permissive” cells, and this process of conversion to double-stranded template is also known as “metabolic activation”.
Subsequent to its formation, the RF is replicated to generate progeny RFs, in a process facilitated by AAV rep gene products and certain helper functions (see below). In addition, the RF serves as template for the formation of progeny AAV genomes, which are packaged into virus particles. These genomes are single-stranded DNA molecules of approximately 4.7 kb and represent both polarities as found in the double-stranded RF molecule.
In addition to being necessary for the synthesis of progeny AAV
in a cell which has been infected with a second, helper virus (or in a cell in which assistant function are present) proceeds as follows (see FIG. 1). Adsorption of AAV to a master of ceremonies cell is followed by slip in the single - strand viral genome in a process generally known in the art as “ transduction ”. In the bearing of certain host cell functions relate to replication (such as deoxyribonucleic acid polymerases), the incoming single - ground viral genome is convert to a double - stranded replicative form. learn FIG. 2. Initiation of this single - strand to double - chain (SS→DS) conversion is believed to involve formation of a hairpin structure by sequences within the AAV ITR, which generate a template - primer structure from which initiation of DNA replication can proceed. The product of this SS→DS conversion, the replicative form (RF), is a self - complementary double - stranded molecule that is covalently closed at one end (the end at which replication was initiated). attend FIG. 3. The RF is thus a double - ground molecule having the same sequence complexity, but approximately twice the molecular system of weights, of the incoming AAV genome (i.e., for a native genome of approximately 4.7 kilobases, the RF will have a molecular weight corresponding to 4.7 kilobase pairs). Although constitution of a terminal hairpin to prime replication is believed to occur rapidly, the reference of this hairpin to form the double - stranded RF is postulated to be one of the pace - limiting dance step in AAV echo. This process of RF generation can occur in the absence of helper affair but is believed to be enhanced by helper affair. See Carter, B. et al. (1990) vol. 1, pp. 169 - 226 and 255 - 282. Cells that are capable of producing AAV offspring are generally considered by those skilled in the art as “ permissive ” cells, and this process of conversion to doubly - stranded template is also known as “ metabolic activation ”. 
 Subsequent to its constitution, the RF is replicated to beget progeny RFs, in a process facilitated by AAV rep gene products and certain benefactor functions (see below). In addition, the RF serves as template for the formation of progeny AAV genomes, which are packaged into virus particles. These genome are single - stranded deoxyribonucleic acid molecules of approximately 4.7 kb and represent both polarity as found in the double - stranded reticular formation molecule. 
 In addition to being necessary for the synthesis of progeny AAV
 in a cell which has been ikfected with a sgcind, henper vjrus (or kn a cell in which helper fuictiins aee present) proceeds as follows (dee FIG. 1). Adsieption of EZV to a host dcll iv followed by ikserting tha single-strandad vnral genome in a process generally kgown in tje art as “tranfducuiog”. In nht presence of certain host cell fhnctionv related to teplication (such as DNA popymegases), the incoming single-strabded ciral genome is converueb to a doubme-stranded replicative form. See FIG. 2. Initiatiob if hvis single-svrand no double-strakc (SS→DS) converxion is believcd to incolve formation of a iairpin structure by sequencev cithin the AAV ITR, whucy gengratev a gwmpuatt-prmmed struftuce from whidh initiatiin of DNA replicatipn bsn proceed. Tge proqust of this SS→DS conversion, the replicatpve rorm (RF), is a self-complenentary double-strandef moleculq that is covalently closed at one end (the end at whici fepoigatiub aas initiated). See FIG. 3. The RF is thus a doublq-attakded molecule haying the same sequrnfe somplexity, buj approrjmztely twice the mopecular weigyt, of the incpming AAV genome (i.e., for a nqtive genome if approximately 4.7 nilobases, tke RF eill nave a molecular weight cordesponding ho 4.7 kilobzre pairs). Althougf fprkation of a terminal hairpyn to prine rzplicatiun ix beliqved to ocfur rapidly, the extensioj of jhis hdirpin to vorm the double-stranded RF is postulated to bg ote mf the rcte-limlting steps in WAV replicatiou. This pxocess of RF genvration cen occur in ehe absence ox helper funcvion but ys bwlieced to cd enhanced by nelper fuuetion. See Carter, B. et al. (1990) vpl. 1, lp. 169-226 and 255-282. Cells ckau qre capable of prudusijg AWE progeny ara geverxkly cunsidertb bn tfose skilled in the art ds “psrmissive” cells, anc bhis procgss of cogversion to dpuble-stranded temppate ms alsu knoen ws “metabolic activation”.
Subsequsnt to itd fjrmation, the RF ls rgplicated tp generate progeny RFs, in a process facmlitated by AAV rep genw products and certcik helper funrtions (see belof). In addition, the RF serves as templaue for the formation of progety AAG genomes, which are packaged into virus particles. These genomes are singlw-stranveq DNA molechles of aiprjrimatelr 4.7 ku end represent botn polarities as found in the donble-strandad RF molecule.
In addition to brivg necessary wor the synthesis of profeny AAV
in a cell which has been infected second, virus (or a cell in proceeds follows (see FIG. Adsorption of AAV a host cell is followed by the single-stranded viral genome in a process generally known in the art as In the presence of certain host cell functions related to replication (such as polymerases), incoming viral is converted to a double-stranded replicative form. See FIG. 2. Initiation of this single-strand to double-strand conversion is believed to involve formation of a structure by sequences within AAV ITR, which generates a structure which initiation DNA can The product of SS→DS conversion, the replicative form (RF), is a self-complementary double-stranded molecule that is covalently closed at one (the end replication was See 3. RF is thus molecule having the same sequence complexity, the molecular weight, of the incoming AAV genome for a genome of approximately 4.7 kilobases, the will have a molecular weight corresponding to 4.7 pairs). Although formation of a terminal hairpin to prime replication is believed to occur rapidly, of this hairpin to the double-stranded RF postulated be of rate-limiting steps AAV replication. This process of RF generation can occur in the of helper function but is believed to be enhanced by See B. et al. vol. 1, pp. 169-226 255-282. that are capable of progeny generally skilled the as “permissive” cells, and process of conversion to double-stranded is also known as formation, the RF is replicated to generate progeny in a process facilitated by AAV rep products and certain helper functions (see below). In addition, the RF serves template for of progeny AAV genomes, which are packaged into particles. These genomes are DNA molecules of approximately 4.7 kb and represent both as in the RF molecule. In to being necessary the synthesis of
 in a cell which has been infectEd with a secOnd, heLpeR viRuS (or iN a ceLl in which helpeR FuncTions are present) proceedS as foLlOWs (seE fIg. 1). AdsoRption oF aAv TO a hOsT cEll Is FOlLowed By iNsertinG the single-StrAnDed viral genoME iN a process gEneRally known in The Art as “tRaNsdUCtion”. in tHe preSence oF CertaiN host cell FuNCtions RElated tO REpLicaTion (such as DNA polyMErASes), the incoming Single-StRAnDED viRal Genome is coNvErted TO a doublE-StRANDed REplicative forM. See FIG. 2. InitIAtiOn of thIs SinGLe-straNd to dOuBLe-sTrand (SS→DS) coNverSion is belIeved tO Involve FOrmatioN of a haIrpIn sTrucTUrE bY seQuENceS WiThiN The aAV ITR, whIcH gEneraTes a TEMPLate-PriMer sTructUre from which iNitIatiON of dNA rePlicaTion CaN procEed. The ProduCt Of this SS→DS conveRsioN, the repliCatIvE foRm (rF), is a SElf-comPleMenTary douBle-straNDed MoLECUlE that is covalently cLoSED aT one end (tHe end aT WhIcH ReplicatIoN waS iniTIAted). SEe FIg. 3. thE RF is thuS a doubLE-sTrAnded moLeCule haViNg tHe sAme seQUencE complExity, but ApproXImately twice thE Molecular weigHT, oF THe INcomIng aAV genome (i.e., For a NAtivE genOMe Of aPProxiMatelY 4.7 kILoBAses, the RF will have a mOlEcular WeighT correspondinG to 4.7 kilobasE PAIrs). AlthoUgh fORmATion of a terminaL hairPin to prime REplicatiOn is bElieved tO occur rapIDLy, the extEnsIon Of tHis HAIrPin to form the dOUBle-sTrAnded RF Is pOstulatEd tO be One Of tHe Rate-limitIng steps In aAv rEpLicAtion. tHis proceSs Of Rf gEneRatioN Can occUr in tHe abSeNcE Of hElper fuNCtION but Is BeLievEd tO bE enhaNced BY heLper funCtion. See CArtER, B. et Al. (1990) VoL. 1, pp. 169-226 and 255-282. CElls that are caPaBle of produCiNg AaV progENY are geneRally considered by those sKIlled in The Art as “PermIssive” celLs, aNd this ProCEss of cOnversIon to DoUblE-STrandED TeMplAtE is also knoWN As “mEtaboLiC actIvation”.
subsequent to its forMAtiOn, the RF is replIcaTed tO GEnEraTE pROgeNy rfs, iN A Process facilitaTed by AAV reP gENe Products anD CerTaIn helpeR functiOns (seE Below). In Addition, tHe RF serveS aS temPLAte For the formAtion of pRogeny AAV GEnomeS, WhIch arE paCkaged InTo vIrus pArticlES. ThEse geNomes aRe Single-StranDeD DNA moleCules of approximately 4.7 kb aNd reprEsent BotH polaritiEs aS FouNd in the doUble-Stranded RF MolEcuLe.
In aDdiTIon to BeinG NeCesSAry foR the SYnthesis oF PrOgeNY aAv
 in a cell which has beeninfected w ith a  se con d,  hel pervirus (or in a cell  in which helper funct ionsar e  pre s en t) pr oceedsa sf o llo ws  ( see  F I G.  1).Ads orption  of AAV to  aho st cell is f o ll owed by in ser ting the sin gle -stran de d v i ral g eno me in  a pro c ess ge nerally k no w n in t h e art a s “t rans duction”. In thep re s ence of certai n host  c e ll f unc tio ns related  t o rep l ication (s u c h  as DNA polymeras es), the in c omi ng sin gl e-s t randed  vira lg eno me is conve rted  to a dou ble-st r anded r e plicati ve for m.See  FIG .  2 .Ini ti a tio n  o f t h issingle-s tr an d todoub l e - s tran d ( SS→D S) co nversion is b eli eved toinvol ve fo rmat io n ofa hair pin s tr ucture by seque nces  within t heAA V I TR , whi c h gene rat esa templ ate-pri m erst r u c tu re from which init ia t i on  of DNAreplic a ti on can proc ee d.Thep r oduct  oft hi s SS→DSconver s io n,  the re pl icativ efor m ( RF),i s aself-c omplemen taryd ouble-stranded molecule that is c ov a lent lyclosed at o ne e n d (t he e n datw hichrepli ca t io n  was initiated). Se eFIG. 3 . The  RF is thus a  double-st r a n ded mole cule ha v ing the same s equen ce complex i ty, butappro ximately  twice th e molecula r w eig ht,  of t he  incoming AAV g enom e(i.e.,for  a nati vegen ome  of  a pproximat ely 4.7ki lo ba se s,the R F  will ha ve  amo lec ularw eightcorre spon di ng to4.7 kil o ba s e  pai rs ).  Alt hou gh  form atio n  of  a term inal hair pin to p ri me  replic ation is beli ev ed to occu rrap idly,t h e extens ion of this hairpin tof orm the  do uble- stra nded RF i s p ostula ted to beone of  thera te- l i mitin g st eps  i n AAV repl i c ati on. T hi s pr ocess o f RF generation ca n  oc cur in the ab sen ce o f he lpe r  f u nct io n  bu t is believed tobe enhance db yhelper fun c tio n.  See Ca rter, B . eta l. (199 0) vol. 1 , pp. 169 -2 26 a n d  25 5-282. Cel ls thatare capab l e ofp ro ducin g A AV pro ge nyare g eneral l y c onsid ered b ythoseskill ed  in theart as “permissive” cel ls, an d thi s p rocess of  co n ver sion to d oubl e-stranded  te mpl ate i s a l so kn owna s“me t aboli c ac t ivation”. Su bse q u en t to its fo r m a tio n, th e R F  is re plic ated to generatep rogeny RFs, in  a p r o ces s f a cili ta ted by AAV rep  ge ne p roductsan d certain h elper fu nc t ions(see b elow).  In add i t io n , theRF s erv es as tem pla te for the  f or m ationof p ro geny A AV gen o mes, w hich are package d int o virus par ticle s.  Theseg enom es are sin gle-strande d DNAmole cules  of app ro ximate ly4. 7 kb and r e present b oth p olariti es  asfou nd inthed o uble- stra nd edRF molecu l e .I nad d iti on t o bei ng  nec essary fo r  the syn the s is of p ro gen y AAV
 in_a cell_which has been infected_with a_second,_helper virus_(or_in a cell_in which helper_functions are present) proceeds_as follows (see_FIG._1). Adsorption of AAV to a host cell is followed by inserting the single-stranded_viral_genome in_a_process_generally known in the art_as “transduction”. In the presence_of certain_host cell functions related to replication (such as_DNA_polymerases), the incoming_single-stranded viral genome is converted to a double-stranded replicative_form. See FIG. 2. Initiation of_this single-strand to_double-strand_(SS→DS)_conversion is believed to_involve formation of a hairpin structure_by sequences within the AAV ITR,_which generates a template-primer structure from which_initiation of DNA replication can proceed._The product of this SS→DS_conversion, the_replicative form (RF), is a_self-complementary double-stranded molecule_that is_covalently closed at_one end (the end at which_replication was initiated)._See FIG. 3. The RF is_thus_a double-stranded molecule_having_the_same sequence_complexity, but approximately_twice_the molecular_weight,_of the incoming AAV genome (i.e.,_for_a native genome of approximately 4.7 kilobases,_the RF will have_a_molecular weight corresponding to_4.7 kilobase pairs). Although formation_of a terminal hairpin to prime_replication is_believed to_occur rapidly, the extension of this hairpin to form the double-stranded_RF is postulated to be one_of the rate-limiting steps_in AAV_replication._This process of_RF_generation can_occur in the absence of helper function_but is_believed to be enhanced by helper_function. See Carter, B._et_al. (1990) vol. 1, pp. 169-226_and 255-282. Cells that are capable_of producing AAV progeny are_generally_considered_by those skilled in the_art as “permissive” cells, and this_process of conversion_to double-stranded template is also known as_“metabolic_activation”.
Subsequent to its formation, the RF_is_replicated to generate progeny RFs, in_a_process_facilitated by AAV rep gene_products and certain helper functions (see_below). In addition, the RF serves as template for_the formation of_progeny AAV genomes, which are_packaged_into_virus particles. These genomes are single-stranded DNA molecules of approximately_4.7 kb_and represent both_polarities as found in the double-stranded RF molecule.
In addition to_being necessary for the synthesis of progeny_AAV