LOCUS       sequence-43858-e        6341 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 6341)
  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 6341)
  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-miR122
FEATURES             Location/Qualifiers
     source          1..6341
                     /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(3178..3197)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3243..3298
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     repeat_region   3391..3520
                     /label=AAV2 ITR
     primer_bind     complement(3539..3556)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3539..3555)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3548..3570)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3697..4152
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3784..3803)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3994..4015
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4178..4282
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4283..4351,4352..5143)
                     /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(4501..4520)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5314..5902
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5803..5822
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6190..6211
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6226..6231,6232..6249,6250..6256)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6264..6280
                     /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     6269..6291
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6288..6304
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6288..6304
                     /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 tatcgggccc gactgcagtt tcagcgtttg gaaccactga ggagtctgtg
     2881 acagacaacg gaggaatgtg aagggaaggt taataccctt gtgcctgaca gactttcctt
     2941 agcagagctg tggagtgtga caatggtgtt tgtgtccaaa ccatcaaacg ccattatcac
     3001 actaaatagc tactgctagg caatccgtcc actccacgcg tgacttgacg tctgttctct
     3061 ccgagcaaga agttctgtct tgtgctagag ttctccattg tgtgtaatgt aaagtcgggc
     3121 ccgcgatcgg taatgctcta gagtcgagga cggggtgaac tacgcctgag gatccgatct
     3181 ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg
     3241 ctaataaagg aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact
     3301 cggaagcaat tcgttgatct gaatttcgac cacccataat acccattacc ctggtagata
     3361 agtagcatgg cgggttaatc attaactaca aggaacccct agtgatggag ttggccactc
     3421 cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg
     3481 gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ccttaattaa cctaattcac
     3541 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     3601 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     3661 cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa
     3721 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     3781 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3841 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     3901 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     3961 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4021 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     4081 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     4141 cgcttacaat ttaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt
     4201 tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca
     4261 ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     4321 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     4381 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     4441 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     4501 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     4561 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     4621 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     4681 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     4741 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     4801 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     4861 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggcggataa
     4921 agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc
     4981 tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc
     5041 ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag
     5101 acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag accaagttta
     5161 ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa
     5221 gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     5281 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     5341 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     5401 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     5461 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     5521 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     5581 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     5641 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     5701 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     5761 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     5821 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     5881 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     5941 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     6001 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     6061 gtcagtgagc gaggaagcgg aagagcgccc aatacgcaaa ccgcctctcc ccgcgcgttg
     6121 gccgattcat taatgcagct ggcacgacag gtttcccgac tggaaagcgg gcagtgagcg
     6181 caacgcaatt aatgtgagtt agctcactca ttaggcaccc caggctttac actttatgct
     6241 tccggctcgt atgttgtgtg gaattgtgag cggataacaa tttcacacag gaaacagcta
     6301 tgaccatgat tacgccagat ttaattaagg ccttaattag g
//
