LOCUS       sequence-43857-e        6051 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6051)
  AUTHORS   Xie J, Ameres SL, Friedline R, Hung JH, Zhang Y, Xie Q, Zhong L, Su 
            Q, He R, Li M, Li H, Mu X, Zhang H, Broderick JA, Kim JK, Weng Z, 
            Flotte TR, Zamore PD, Gao G
  TITLE     Long-term, efficient inhibition of microRNA function in mice using 
            rAAV vectors.
  JOURNAL   Nat Methods. 2012 Mar 4;9(4):403-9. doi: 10.1038/nmeth.1903.
  PUBMED    22388288
REFERENCE   2  (bases 1 to 6051)
  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: "10.1038/nmeth"; 
            journalName: "Nat Methods"; date: "2012-03-4- 4"; volume: "9"; 
            issue: "4"; pages: "403-9"
COMMENT     Sequence Label: pAAV CB6 FFLuc
FEATURES             Location/Qualifiers
     source          1..6051
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     enhancer        204..583
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          966..1098
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1081..1100
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     promoter        1143..1161
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1163..2815
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(1229..1247)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     primer_bind     complement(2888..2907)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2953..3008
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     repeat_region   3101..3230
                     /label=AAV2 ITR
     primer_bind     complement(3249..3266)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3249..3265)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3258..3280)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3407..3862
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3494..3513)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3704..3725
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3888..3992
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3993..4061,4062..4853)
                     /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(4211..4230)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5024..5612
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5513..5532
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    5900..5921
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5936..5941,5942..5959,5960..5966)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5974..5990
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5979..6001
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5998..6014
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5998..6014
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctaccag ggtaatgggg
      181 atcctctaga actatagcta gtcgacattg attattgact agttattaat agtaatcaat
      241 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      301 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      361 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      421 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      481 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      541 tacttggcag tacatctacg tattagtcat cgctattacc atgtcgaggc cacgttctgc
      601 ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt tattttttaa
      661 ttattttgtg cagcgatggg ggcggggggg gggggcgcgc gccaggcggg gcggggcggg
      721 gcgaggggcg gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc
      781 tccgaaagtt tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg
      841 cgcggcgggc gggagcaagc tttattgcgg tagtttatca cagttaaatt gctaacgcag
      901 tcagtgcttc tgacacaaca gtctcgaact taagctgcag aagttggtcg tgaggcactg
      961 ggcaggtaag tatcaaggtt acaagacagg tttaaggaga ccaatagaaa ctgggcttgt
     1021 cgagacagag aagactcttg cgtttctgat aggcacctat tggtcttact gacatccact
     1081 ttgcctttct ctccacaggt gtccactccc agttcaatta cagctcttaa ggctagagta
     1141 cttaatacga ctcactatag gcatggaaga cgccaaaaac ataaagaaag gcccggcgcc
     1201 attctatccg ctggaagatg gaaccgctgg agagcaactg cataaggcta tgaagagata
     1261 cgccctggtt cctggaacaa ttgcttttac agatgcacat atcgaggtgg acatcactta
     1321 cgctgagtac ttcgaaatgt ccgttcggtt ggcagaagct atgaaacgat atgggctgaa
     1381 tacaaatcac agaatcgtcg tatgcagtga aaactctctt caattcttta tgccggtgtt
     1441 gggcgcgtta tttatcggag ttgcagttgc gcccgcgaac gacatttata atgaacgtga
     1501 attgctcaac agtatgggca tttcgcagcc taccgtggtg ttcgtttcca aaaaggggtt
     1561 gcaaaaaatt ttgaacgtgc aaaaaaagct cccaatcatc caaaaaatta ttatcatgga
     1621 ttctaaaacg gattaccagg gatttcagtc gatgtacacg ttcgtcacat ctcatctacc
     1681 tcccggtttt aatgaatacg attttgtgcc agagtccttc gatagggaca agacaattgc
     1741 actgatcatg aactcctctg gatctactgg tctgcctaaa ggtgtcgctc tgcctcatag
     1801 aactgcctgc gtgagattct cgcatgccag agatcctatt tttggcaatc aaatcattcc
     1861 ggatactgcg attttaagtg ttgttccatt ccatcacggt tttggaatgt ttactacact
     1921 cggatatttg atatgtggat ttcgagtcgt cttaatgtat agatttgaag aagagctgtt
     1981 tctgaggagc cttcaggatt acaagattca aagtgcgctg ctggtgccaa ccctattctc
     2041 cttcttcgcc aaaagcactc tgattgacaa atacgattta tctaatttac acgaaattgc
     2101 ttctggtggc gctcccctct ctaaggaagt cggggaagcg gttgccaaga ggttccatct
     2161 gccaggtatc aggcaaggat atgggctcac tgagactaca tcagctattc tgattacacc
     2221 cgagggggat gataaaccgg gcgcggtcgg taaagttgtt ccattttttg aagcgaaggt
     2281 tgtggatctg gataccggga aaacgctggg cgttaatcaa agaggcgaac tgtgtgtgag
     2341 aggtcctatg attatgtccg gttatgtaaa caatccggaa gcgaccaacg ccttgattga
     2401 caaggatgga tggctacatt ctggagacat agcttactgg gacgaagacg aacacttctt
     2461 catcgttgac cgcctgaagt ctctgattaa gtacaaaggc tatcaggtgg ctcccgctga
     2521 attggaatcc atcttgctcc aacaccccaa catcttcgac gcaggtgtcg caggtcttcc
     2581 cgacgatgac gccggtgaac ttcccgccgc cgttgttgtt ttggagcacg gaaagacgat
     2641 gacggaaaaa gagatcgtgg attacgtcgc cagtcaagta acaaccgcga aaaagttgcg
     2701 cggaggagtt gtgtttgtgg acgaagtacc gaaaggtctt accggaaaac tcgacgcaag
     2761 aaaaatcaga gagatcctca taaaggccaa gaagggcgga aagatcgccg tgtaattcta
     2821 gattccgaga tatcggtaat gggccctcta gagtcgagga cggggtgaac tacgcctgag
     2881 gatccgatct ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcatc
     2941 tgacttctgg ctaataaagg aaatttattt tcattgcaat agtgtgttgg aattttttgt
     3001 gtctctcact cggaagcaat tcgttgatct gaatttcgac cacccataat acccattacc
     3061 ctggtagata agtagcatgg cgggttaatc attaactaca aggaacccct agtgatggag
     3121 ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc
     3181 cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ccttaattaa
     3241 cctaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa
     3301 cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc
     3361 accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc
     3421 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     3481 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     3541 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     3601 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     3661 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     3721 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     3781 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     3841 aaaatattaa cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     3901 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     3961 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     4021 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     4081 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     4141 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     4201 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     4261 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     4321 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     4381 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     4441 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     4501 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     4561 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     4621 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     4681 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     4741 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     4801 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     4861 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     4921 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     4981 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     5041 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     5101 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     5161 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     5221 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     5281 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     5341 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     5401 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     5461 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     5521 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     5581 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     5641 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     5701 tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg
     5761 agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc
     5821 ccgcgcgttg gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg
     5881 gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac
     5941 actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag
     6001 gaaacagcta tgaccatgat tacgccagat ttaattaagg ccttaattag g
//
