LOCUS       sequence-47857-e        6780 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 6780)
  AUTHORS   Watabe-Uchida M, Zhu L, Ogawa SK, Vamanrao A, Uchida N
  TITLE     Whole-brain mapping of direct inputs to midbrain dopamine neurons.
  JOURNAL   Neuron. 2012 Jun 7;74(5):858-73.
  PUBMED    22681690
REFERENCE   2  (bases 1 to 6780)
  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"; journalName: "Neuron."; 
            date: "2012-06-7"; volume: "74(5)"; pages: "858-73"
COMMENT     Sequence Label: pAAV-EF1a-FLEX-TVA-mCherry
FEATURES             Location/Qualifiers
     source          1..6780
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1450..1483)
                     /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(1534..1567)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1577..2284)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1855..1874)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2124..2142
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     protein_bind    2765..2798
                     /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    2849..2882
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2907..3495
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2960..2980)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3378..3389)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3527..4003
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3726..3745)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4043..4183
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4043..4172
                     /label=AAV2 ITR
     rep_origin      4258..4713
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4345..4364)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4555..4576
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4730..4749)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4849..4871
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4909..4927)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4995..5099
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5100..5168,5169..5960)
                     /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(5318..5337)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6131..6719
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6620..6639
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc
     1441 gactccggaa taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga
     1501 ttgtagctgc tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg
     1561 aagttatggc gcgatcttac ttgtacagct cgtccatgcc gccggtggag tggcggccct
     1621 cggcgcgttc gtactgttcc acgatggtgt agtcctcgtt gtgggaggtg atgtccaact
     1681 tgatgttgac gttgtaggcg ccgggcagct gcacgggctt cttggccttg taggtggtct
     1741 tgacctcagc gtcgtagtgg ccgccgtcct tcagcttcag cctctgcttg atctcgccct
     1801 tcagggcgcc gtcctcgggg tacatccgct cggaggaggc ctcccagccc atggtcttct
     1861 tctgcattac ggggccgtcg gaggggaagt tggtgccgcg cagcttcacc ttgtagatga
     1921 actcgccgtc ctgcagggag gagtcctggg tcacggtcac cacgccgccg tcctcgaagt
     1981 tcatcacgcg ctcccacttg aagccctcgg ggaaggacag cttcaagtag tcggggatgt
     2041 cggcggggtg cttcacgtag gccttggagc cgtacatgaa ctgaggggac aggatgtccc
     2101 aggcgaaggg cagggggcca cccttggtca ccttcagctt ggcggtctgg gtgccctcgt
     2161 aggggcggcc ctcgccctcg ccctcgatct cgaactcgtg gccgttcacg gagccctcca
     2221 tgtgcacctt gaagcgcatg aactccttga tgatggccat gttatcctcc tcgcccttgc
     2281 tcaccccacc ccccttggat gcgctttcaa gactgaagat gtcagacctg ctttttgctt
     2341 tggacttccc ccatgcagcg ataccaccca cagctaccag gcagcacagg agcactgcag
     2401 tgatcagcat ccacatgcgg ccgtgattcc tggctggcag agcacgtcca ggagcagggg
     2461 cagtgggagc ctctgtgccg ttgtccgtgg gcaccgcggg gatcgcgctg gtcccgcagc
     2521 cccactcgtc ccgcccgtcg tcgcagtcgg ggtgtccgtc gcacagccag tcctgcgggt
     2581 aacactcccc gtgggcaccg ggcggctccg agcagcggaa ctgaccgggg gggcaacggg
     2641 acaaagaacc gttaccggac ccgttaccgg tcacgttacc gggcagcagc agcagcagca
     2701 gcgcgggcag cagccgcgcc atgtcggtac caccacatca agcttatcga taccgtcgac
     2761 tagcataact tcgtataaag tatcctatac gaagttattt gccttaaccc agaaattatc
     2821 actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct atacgaagtt
     2881 atgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
     2941 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     3001 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     3061 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     3121 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     3181 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     3241 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     3301 aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg cgcgggacgt
     3361 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     3421 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     3481 gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt ggcatccctg
     3541 tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc ccaccagcct
     3601 tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct ataatattat
     3661 ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct gtagggcctg
     3721 cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca ctgcaatctc
     3781 cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg gattccaggc
     3841 atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt tcaccatatt
     3901 ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc ctcccaaatt
     3961 gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt tgtaggtaac
     4021 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     4081 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4141 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4201 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4261 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4321 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4381 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4441 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     4501 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     4561 tcttgttcca aactggaaca acactcaacc ctatctcggg ctattctttt gatttataag
     4621 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     4681 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     4741 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     4801 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     4861 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     4921 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     4981 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     5041 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5101 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5161 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5221 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5281 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5341 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5401 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5461 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     5521 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     5581 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     5641 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     5701 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     5761 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     5821 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     5881 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     5941 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     6001 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6061 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6121 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6181 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6241 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     6301 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6361 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6421 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     6481 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     6541 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     6601 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     6661 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     6721 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
