LOCUS       Exported                8543 bp ds-DNA     circular SYN 20-MAY-2019
DEFINITION  non-standard AAV2 rep-AAV-PHP.eB cap plasmid with AAV cap expression
            controlled by a tTA-TRE amplifcation system.
ACCESSION   .
VERSION     .
KEYWORDS    pUCmini-iCAP-PHP.eB
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8543)
  AUTHORS   Chan KY, Jang MJ, Yoo BB, Greenbaum A, Ravi N, Wu WL, 
            Sanchez-Guardado L, Lois C, Mazmanian SK, Deverman BE, Gradinaru V
  TITLE     Engineered AAVs for efficient noninvasive gene delivery to the 
            central and peripheral nervous systems.
  JOURNAL   Nat Neurosci. 2017 Aug;20(8):1172-1179. doi: 10.1038/nn.4593. Epub 
            2017 Jun 26.
  PUBMED    28671695
REFERENCE   2  (bases 1 to 8543)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, May 21, 2019 from SnapGene Viewer 4.3.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..8543
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     99..121
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     113..130
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     114..130
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     140..159
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        140..158
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    239..2101
                     /label=AAV2 Rep
     misc_feature    2121..2867
                     /label=tTA
     CDS             2748..2858
                     /codon_start=1
                     /product="3 tandem repeats of the minimal activation domain
                     of herpes simplex virus VP16 (Cheng et al., 2013)"
                     /label=VP48
                     /translation="DALDDFDLDMLPADALDDFDLDMLPADALDDFDLDML"
     misc_feature    2894..3206
                     /label=pA_BGH
     promoter        complement(2897..2915)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2898..2915)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(2935..2952)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2941..3165
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    3227..3245
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3264..3282
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3301..3319
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3338..3356
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3375..3393
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3412..3430
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3449..3467
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3486..3504
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    3533..3827
                     /label=AAV5 Rep fragment
     misc_feature    3579..3827
                     /label=p41 promoter
     misc_feature    3828..4131
                     /label=AAV2 Rep - intron donor sites
     misc_feature    3851..3852
                     /label=splice donor
     misc_feature    4143..4146
                     /label=minor VP1 intron acceptor site
     misc_feature    4148..6379
                     /label=AAV9 Cap
     misc_feature    4172..4173
                     /label=major splice acceptor
     CDS             5906..5938
                     /codon_start=1
                     /label=AAV-PHP.B4
                     /translation="AQTLQIPFKAQ"
     misc_feature    5912..5928
                     /label=7mer insertion
     misc_feature    6386..6441
                     /label=AAV2 fragment
     misc_feature    6399..6404
                     /label=polyA
     misc_feature    6474..6604
                     /label=p5
     rep_origin      complement(6788..7376)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6868..6887)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7547..8407)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     8170..8189
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8408..8512)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ccaatgatac gcgtcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
       61 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
      121 gacggccagt gagcgcgcgt aatacgactc actatagggc gaattgggta catcgacggt
      181 atcgggggag ctcgcagggt ctccattttg aagcgggagg tttgaacgcg cagccgccat
      241 gccggggttt tacgagattg tgattaaggt ccccagcgac cttgacgagc atctgcccgg
      301 catttctgac agctttgtga actgggtggc cgagaaggaa tgggagttgc cgccagattc
      361 tgacatggat ctgaatctga ttgagcaggc acccctgacc gtggccgaga agctgcagcg
      421 cgactttctg acggaatggc gccgtgtgag taaggccccg gaggctcttt tctttgtgca
      481 atttgagaag ggagagagct acttccacat gcacgtgctc gtggaaacca ccggggtgaa
      541 atccatggtt ttgggacgtt tcctgagtca gattcgcgaa aaactgattc agagaattta
      601 ccgcgggatc gagccgactt tgccaaactg gttcgcggtc acaaagacca gaaatggcgc
      661 cggaggcggg aacaaggtgg tggatgagtg ctacatcccc aattacttgc tccccaaaac
      721 ccagcctgag ctccagtggg cgtggactaa tatggaacag tatttaagcg cctgtttgaa
      781 tctcacggag cgtaaacggt tggtggcgca gcatctgacg cacgtgtcgc agacgcagga
      841 gcagaacaaa gagaatcaga atcccaattc tgatgcgccg gtgatcagat caaaaacttc
      901 agccaggtac atggagctgg tcgggtggct cgtggacaag gggattacct cggagaagca
      961 gtggatccag gaggaccagg cctcatacat ctccttcaat gcggcctcca actcgcggtc
     1021 ccaaatcaag gctgccttgg acaatgcggg aaagattatg agcctgacta aaaccgcccc
     1081 cgactacctg gtgggccagc agcccgtgga ggacatttcc agcaatcgga tttataaaat
     1141 tttggaacta aacgggtacg atccccaata tgcggcttcc gtctttctgg gatgggccac
     1201 gaaaaagttc ggcaagagga acaccatctg gctgtttggg cctgcaacta ccgggaagac
     1261 caacatcgcg gaggccatag cccacactgt gcccttctac gggtgcgtaa actggaccaa
     1321 tgagaacttt cccttcaacg actgtgtgga caagatggtg atctggtggg aggaggggaa
     1381 gatgaccgcc aaggtcgtgg agtcggccaa agccattctc ggaggaagca aggtgcgcgt
     1441 ggaccagaaa tgcaagtcct cggcccagat agacccgact cccgtgatcg tcacctccaa
     1501 caccaatatg tgcgccgtga ttgacgggaa ctcaacgacc ttcgaacacc agcagccgtt
     1561 gcaagaccgg atgttcaaat ttgaactcac ccgccgtctg gatcatgact ttgggaaggt
     1621 caccaagcag gaagtcaaag actttttccg gtgggcaaag gatcacgtgg ttgaggtgga
     1681 gcatgaattc tacgtcaaaa agggtggagc caagaaaaga cccgccccca gtgacgcaga
     1741 tataagtgag cccaaacggg tgcgcgagtc agttgcgcag ccatcgacgt cagacgcgga
     1801 agcttcgatc aactacgcgg acaggtacca aaacaaatgt tctcgtcacg tgggcatgaa
     1861 tctgatgctg tttccctgca gacaatgcga gagactgaat cagaattcaa atatctgctt
     1921 cactcacggt gtcaaagact gtttagagtg ctttcccgtg tcagaatctc aacccgtttc
     1981 tgtcgtcaaa aaggcgtatc agaaactgtg ctacattcat cacatcatgg gaaaggtgcc
     2041 agacgcttgc actgcttgcg acctggtcaa tgtggacttg gatgactgtg tttctgaaca
     2101 ataaatgact taaaccaggt atgagtcggc tggataaatc taaagtcata aacggcgctc
     2161 tggaattact caatgaagtc ggtatcgaag gcctgacgac aaggaaactc gctcaaaagc
     2221 tgggagttga gcagcctacc ctgtactggc acgtgaagaa caagcgggcc ctgctcgatg
     2281 ccctggccat cgagatgctg gacaggcatc atacccactt ctgccccctg gaaggcgagt
     2341 catggcaaga ctttctgcgg aacaacgcca agtcattccg ctgtgctctc ctctcacatc
     2401 gcgacggggc taaagtgcat ctcggcaccc gcccaacaga gaaacagtac gaaaccctgg
     2461 aaaatcagct cgcgttcctg tgtcagcaag gcttctccct ggagaacgca ctgtacgctc
     2521 tgtccgccgt gggccacttt acactgggct gcgtattgga ggaacaggag catcaagtag
     2581 caaaagagga aagagagaca cctaccaccg attctatgcc cccacttctg agacaagcaa
     2641 ttgagctgtt cgaccggcag ggagccgaac ctgccttcct tttcggcctg gaactaatca
     2701 tatgtggcct ggagaaacag ctaaagtgcg aaagcggcgg gccggccgac gcccttgacg
     2761 attttgactt agacatgctc ccagccgatg cccttgacga ctttgacctt gatatgctgc
     2821 ctgctgacgc tcttgacgat tttgaccttg acatgctccc cgggtaaatg catgaattcg
     2881 atctagaggg ccctattcta tagtgtcacc taaatgctag agctcgctga tcagcctcga
     2941 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     3001 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     3061 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     3121 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     3181 gaaccagctg gggctcgaat caagctatca agtgccacct gacgtctccc tatcagtgat
     3241 agagaagtcg acacgtctcg agctccctat cagtgataga gaaggtacgt ctagaacgtc
     3301 tccctatcag tgatagagaa gtcgacacgt ctcgagctcc ctatcagtga tagagaaggt
     3361 acgtctagaa cgtctcccta tcagtgatag agaagtcgac acgtctcgag ctccctatca
     3421 gtgatagaga aggtacgtct agaacgtctc cctatcagtg atagagaagt cgacacgtct
     3481 cgagctccct atcagtgata gagaaggtac cccctatata agcagagaga tctgttcaaa
     3541 tttgaactga ctaagcggct cccgccagat tttggcaaga ttactaagca ggaagtcaag
     3601 gacttttttg cttgggcaaa ggtcaatcag gtgccggtga ctcacgagtt taaagttccc
     3661 agggaattgg cgggaactaa aggggcggag aaatctctaa aacgcccact gggtgacgtc
     3721 accaatacta gctataaaag tctggagaag cgggccaggc tctcatttgt tcccgagacg
     3781 cctcgcagtt cagacgtgac tgttgatccc gctcctctgc gaccgctagc ttcgatcaac
     3841 tacgcagaca ggtaccaaaa caagtgttct cgtcacgtgg gcattaatct gattctgttt
     3901 ccctgcagac aatgcgagag aatgaatcag aactcaaata tctgcttcac tcacggacag
     3961 aaagactgtt tagagtgctt tcccgtgtca gaatctcaac ccgtttctgt cgtcaaaaag
     4021 gcgtatcaga aactgtgcta cattcatcat atcatgggaa aggtgccaga cgcttgcact
     4081 gcctgcgatc tggtcaatgt ggatttggat gactgcatct ttgaacaata aatgacttaa
     4141 gccaggtatg gctgccgatg gttatcttcc agattggctc gaggacaacc ttagtgaagg
     4201 aattcgcgag tggtgggctt tgaaacctgg agcccctcaa cccaaggcaa atcaacaaca
     4261 tcaagacaac gctagaggtc ttgtgcttcc gggttacaaa taccttggac ccggcaacgg
     4321 actcgacaag ggggagccgg tcaacgcagc agacgcggcg gccctcgagc acgacaaagc
     4381 ctacgaccag cagctcaagg ccggagacaa cccgtacctc aagtacaacc acgccgacgc
     4441 cgagttccag gagcggctca aagaagatac gtcttttggg ggcaacctcg ggcgagcagt
     4501 cttccaggcc aaaaagaggc ttcttgaacc tcttggtctg gttgaggaag cggctaagac
     4561 ggctcctgga aagaagaggc ctgtagagca gtctcctcag gaaccggact cctccgcggg
     4621 tattggcaaa tcgggtgcac agcccgctaa aaagagactc aatttcggtc agactggcga
     4681 cacagagtca gtcccagacc ctcaaccaat cggagaacct cccgcagccc cctcaggtgt
     4741 gggatctctt acaatggctt caggtggtgg cgcaccagtg gcagacaata acgaaggtgc
     4801 cgatggagtg ggtagttcct cgggaaattg gcattgcgat tcccaatggc tgggggacag
     4861 agtcatcacc accagcaccc gaacctgggc cctgcccacc tacaacaatc acctctacaa
     4921 gcaaatctcc aacagcacat ctggaggatc ttcaaatgac aacgcctact tcggctacag
     4981 caccccctgg gggtattttg acttcaacag attccactgc cacttctcac cacgtgactg
     5041 gcagcgactc atcaacaaca actggggatt ccggcctaag cgactcaact tcaagctctt
     5101 taacattcag gtcaaagagg ttacggacaa caatggagtc aagaccatcg ccaataacct
     5161 taccagcacg gtccaggtct tcacggactc agactatcag ctcccgtacg tgctcgggtc
     5221 ggctcacgag ggctgcctcc cgccgttccc agcggacgtt ttcatgattc ctcagtacgg
     5281 gtatctgacg cttaatgatg gaagccaggc cgtgggtcgt tcgtcctttt actgcctgga
     5341 atatttcccg tcgcaaatgc taagaacggg taacaacttc cagttcagct acgagtttga
     5401 gaacgtacct ttccatagca gctacgctca cagccaaagc ctggaccgac taatgaatcc
     5461 actcatcgac caatacttgt actatctctc tagaactatt aacggttctg gacagaatca
     5521 acaaacgcta aaattcagtg tggccggacc cagcaacatg gctgtccagg gaagaaacta
     5581 catacctgga cccagctacc gacaacaacg tgtctcaacc actgtgactc aaaacaacaa
     5641 cagcgaattt gcttggcctg gagcttcttc ttgggctctc aatggacgta atagcttgat
     5701 gaatcctgga cctgctatgg cctctcacaa agaaggagag gaccgtttct ttcctttgtc
     5761 tggatcttta atttttggca aacaaggtac tggcagagac aacgtggatg cggacaaagt
     5821 catgataacc aacgaagaag aaattaaaac tactaacccg gtagcaacgg agtcctatgg
     5881 acaagtggcc acaaaccacc agagtgccca aacgttgcag attcctttta aggcacaggc
     5941 gcagaccggt tgggttcaaa accaaggaat acttccgggt atggtttggc aggacagaga
     6001 tgtgtacctg caaggaccca tttgggccaa aattcctcac acggacggca actttcaccc
     6061 ttctccgctg atgggagggt ttggaatgaa gcacccgcct cctcagatcc tcatcaaaaa
     6121 cacacctgta cctgcggatc ctccaacggc cttcaacaag gacaagctga actctttcat
     6181 cacccagtat tctactggtc aagtcagcgt ggagatcgag tgggagctgc agaaggaaaa
     6241 cagcaagcgc tggaacccgg agatccagta cacttccaac tattacaagt ctaataatgt
     6301 tgaatttgct gttaatactg aaggtgtata tagtgaaccc cgccccattg gcaccagata
     6361 cctgactcgt aatctgtaag tcgacttgct tgttaatcaa taaaccgttt aattcgtttc
     6421 agttgaactt tggtctctgc gaagggcaat tcgtttaaac ctgcaggact agaggtcctg
     6481 tattagaggt cacgtgagtg ttttgcgaca ttttgcgaca ccatgtggtc acgctgggta
     6541 tttaagcccg agtgagcacg cagggtctcc attttgaagc gggaggtttg aacgcgcagc
     6601 cgccaagccg aattctgcag atatcacatg tcctaggaac tatcgatcca tcacactggc
     6661 ggccgctcga ctagagcggc cgccaccgcg gtggagctcc agcttttgcg gaccgaatcg
     6721 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     6781 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     6841 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     6901 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     6961 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     7021 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     7081 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     7141 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     7201 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     7261 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     7321 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     7381 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     7441 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     7501 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     7561 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     7621 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     7681 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     7741 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     7801 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     7861 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     7921 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     7981 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     8041 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     8101 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     8161 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     8221 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     8281 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     8341 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     8401 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     8461 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     8521 cccgaaaagt gccacctgac gtc
//
