LOCUS       sequence-201340-        6165 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An AAV genome encoding Cre-dependent expression of the fluorescent 
            protein EYFP from the CAG promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6165)
  AUTHORS   Challis RC, Ravindra Kumar S, Chan KY, Challis C, Beadle K, Jang MJ,
            Kim HM, Rajendran PS, Tompkins JD, Shivkumar K, Deverman BE, 
            Gradinaru V
  TITLE     Systemic AAV vectors for widespread and targeted gene delivery in 
            rodents.
  JOURNAL   Nat Protoc. 2019 Feb;14(2):379-414. doi: 10.1038/s41596-018-0097-3.
  PUBMED    30626963
REFERENCE   2  (bases 1 to 6165)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10"; journalName: 
            "Nat Protoc"; date: "2019-02"; volume: "14"; issue: "2"; pages: 
            "379-414"
COMMENT     Sequence Label: pAAV-CAG-DIO-EYFP
FEATURES             Location/Qualifiers
     source          1..6165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        210..513
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        515..792
                     /label=chicken beta-actin promoter
     primer_bind     1122..1144
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1206..1225
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1292..1325)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1376..1409)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(1418..2131,2132..2134,2135..2137))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1463..1484)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1812..1831
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2071..2092
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2134..2143
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2150..2183
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    2234..2267
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2292..2880
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2345..2365)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2763..2774)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2912..3388
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3111..3130)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3428..3568
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3428..3557
                     /label=AAV2 ITR
     rep_origin      3643..4098
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3730..3749)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3940..3961
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4115..4134)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4234..4256
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4294..4312)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4380..4484
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4485..4553,4554..5345)
                     /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     complement(4703..4722)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5516..6104
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6005..6024
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtaacact agttattaat agtaatcaat tacggggtca
      181 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      241 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      301 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      361 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      421 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      481 tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc
      541 actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta
      601 ttttgtgcag cgatgggggc gggggggggg ggggggcgcg cgccaggcgg ggcggggcgg
      661 ggcgaggggc ggggccgggg gctgcggaga ggtgcggcgg cagccaatca gagcggcgcg
      721 ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc
      781 gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc
      841 tcgcgccgcc cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac
      901 ggcccttctc ctccgggctg taattagcgc ttggtttaat gacggcttgt ttcttttctg
      961 tggctgcgtg aaagccttga ggggctccgg gagcgccggc aggaaggaaa tgggcgggga
     1021 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccct ctccagcctc ggggctgtcc
     1081 gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1141 gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1201 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tgcggcccaa
     1261 cggtaccgga tcctctagag tcgactccgg aataacttcg tataggatac tttatacgaa
     1321 gttatgcaga atggtagctg gattgtagct gctattagca atatgaaacc tcttaataac
     1381 ttcgtatagc atacattata cgaagttatg gcgcgcctta cttgtacagc tcgtccatgc
     1441 cgagagtgat cccggcggcg gtcacgaact ccagcaggac catgtgatcg cgcttctcgt
     1501 tggggtcttt gctcagggcg gactggtagc tcaggtagtg gttgtcgggc agcagcacgg
     1561 ggccgtcgcc gatgggggtg ttctgctggt agtggtcggc gagctgcacg ctgccgtcct
     1621 cgatgttgtg gcggatcttg aagttcacct tgatgccgtt cttctgcttg tcggccatga
     1681 tatagacgtt gtggctgttg tagttgtact ccagcttgtg ccccaggatg ttgccgtcct
     1741 ccttgaagtc gatgcccttc agctcgatgc ggttcaccag ggtgtcgccc tcgaacttca
     1801 cctcggcgcg ggtcttgtag ttgccgtcgt ccttgaagaa gatggtgcgc tcctggacgt
     1861 agccttcggg catggcggac ttgaagaagt cgtgctgctt catgtggtcg gggtagcggg
     1921 cgaagcactg caggccgtag ccgaaggtgg tcacgagggt gggccagggc acgggcagct
     1981 tgccggtggt gcagatgaac ttcagggtca gcttgccgta ggtggcatcg ccctcgccct
     2041 cgccggacac gctgaacttg tggccgttta cgtcgccgtc cagctcgacc aggatgggca
     2101 ccaccccggt gaacagctcc tcgcccttgc tcaccatggt ggcgctagca taacttcgta
     2161 taaagtatcc tatacgaagt tatttgcctt aacccagaaa ttatcactgt tattctttag
     2221 aatggtgcaa agaataactt cgtataatgt atgctatacg aagttatgaa ttcgatatca
     2281 agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta
     2341 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta
     2401 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt
     2461 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg
     2521 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt
     2581 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag
     2641 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc
     2701 cttggctgct cgcctatgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc
     2761 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc
     2821 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc
     2881 atcgataccg agcgctgctc gagagatcta cgggtggcat ccctgtgacc cctccccagt
     2941 gcctctcctg gccctggaag ttgccactcc agtgcccacc agccttgtcc taataaaatt
     3001 aagttgcatc attttgtctg actaggtgtc cttctataat attatggggt ggaggggggt
     3061 ggtatggagc aaggggcaag ttgggaagac aacctgtagg gcctgcgggg tctattggga
     3121 accaagctgg agtgcagtgg cacaatcttg gctcactgca atctccgcct cctgggttca
     3181 agcgattctc ctgcctcagc ctcccgagtt gttgggattc caggcatgca tgaccaggct
     3241 cagctaattt ttgttttttt ggtagagacg gggtttcacc atattggcca ggctggtctc
     3301 caactcctaa tctcaggtga tctacccacc ttggcctccc aaattgctgg gattacaggc
     3361 gtgaaccact gctcccttcc ctgtccttct gattttgtag gtaaccacgt gcggaccgag
     3421 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     3481 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     3541 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     3601 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     3661 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     3721 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     3781 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     3841 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     3901 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     3961 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     4021 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     4081 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     4141 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4201 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4261 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     4321 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     4381 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     4441 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     4501 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     4561 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     4621 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     4681 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     4741 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     4801 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     4861 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     4921 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     4981 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5041 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     5101 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5161 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     5221 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5281 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5341 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     5401 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     5461 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     5521 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     5581 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     5641 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     5701 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     5761 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     5821 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     5881 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     5941 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     6001 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     6061 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     6121 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
