LOCUS       sequence-403531-        6800 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Flp-dependent enzymatically inactive Cre and mRuby3 expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6800)
  AUTHORS   Roethler O, Zohar E, Cohen-Kashi Malina K, Bitan L, Gabel HW, 
            Spiegel I
  TITLE     Single genomic enhancers drive experience-dependent GABAergic 
            plasticity to maintain sensory processing in the adult cortex.
  JOURNAL   Neuron. 2023 Jun 20:S0896-6273(23)00399-9. doi: 
            10.1016/j.neuron.2023.05.026.
  PUBMED    37354902
REFERENCE   2  (bases 1 to 6800)
  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. 
            2023 Jun 20:S0896-6273(23)00399-9. doi: 
            10.1016/j.neuron.2023.05.026."
COMMENT     Sequence Label: pAAV-Ef1a-fDIO-mRuby3-2A-dCre
FEATURES             Location/Qualifiers
     source          1..6800
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    1440..1473
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(1932..1952)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1968..2021)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(2031..2741)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     CDS             complement(2751..2771)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    complement(2785..2818)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    2927..3515
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2980..3000)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3398..3409)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3547..4023
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3746..3765)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4063..4203
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4063..4192
                     /label=AAV2 ITR
     rep_origin      4278..4733
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4365..4384)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4575..4596
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4750..4769)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4869..4891
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4929..4947)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5015..5119
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5120..5188,5189..5980)
                     /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(5338..5357)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6151..6739
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6640..6659
                     /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 gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggag
     1441 aagttcctat tctctagaaa gtataggaac ttcgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttagaagttc ctattcttca aaaggtatag gaacttcggc
     1561 gcgccctact tacggattcg ccgcataacc agtgaaacag cattgctgtc acttggtcgt
     1621 ggcagcccgg accgacgatg aagcatgttt agctggccca aatgttgctg gatagttttt
     1681 actgccagac cgcgcgcctg aagatataga agataatcgc gaacatcttc aggttctgcg
     1741 ggaaaccatt tccggttatt caacttgcac catgccgccc acgaccggca aacggacaga
     1801 agcattttcc aggtatgctc agaaaacgcc tggcgatccc tgaacatgtc catcaggttc
     1861 ttgcgaacct catcactcgt tgcatcgacc ggtaatgcag gcaaattttg gtgtacggtc
     1921 agtaaattgg acaccttcct cttcttcttg ggggttctga aagcgcttgg gccaggattc
     1981 tcctcgacgt caccgcatgt tagcagactt cctctgccct ctccactgcc cttgtacagc
     2041 tcgtccatgc caccaccaag gttgctgtat ttggcaactg ccacttctct ttgcactacg
     2101 taggtttcat tgtcactctc ctcgatcctt tccaggcggt gatcaacggc atggacaccg
     2161 ggcatcttga tgttcccgac ggtctttttt gacctgtaag ttgtcacgaa gttgcagtgc
     2221 agatggccac caccatcaac tttcagtgcg atgtcagtgt atcctctcag accaccatct
     2281 gctggataca tcatctctgt attaggctcc caacccttgg tcttcttctg catcacggga
     2341 ccattggagg gaaagtttac ccctctgacc ttgacgttgt agacgagctc gccatcctca
     2401 aggctggtgt cctgcgtgac ggtgacgact ccaccatctt cgtatctcgt aactctttcc
     2461 caagtaaaac cctcaggaaa ggactgttta aagaaatcag ggatgtcggc cgggtacttg
     2521 ataaaggtac ggctgccata catgaacgac gtggcaagaa tgtcaaaggc aaatggcagg
     2581 ggtcctccct cgatgacttt gatcctcatg gtttgcactc cctcgtacgg tctgccttct
     2641 ccttcacctg tgcatttgaa ttggtggccg ttgaccgaac cttccatgac caccttcata
     2701 cgcatatttt ccttgatcag ctcttcgccc ttagacacca tagcggccgc caccttcctc
     2761 ttcttcttgg gcatggtggc tagcgaagtt cctatacttt ctagagaata ggaacttctt
     2821 gccttaaccc agaaattatc actgttattc tttagaatgg tgcaaagaga agttcctata
     2881 ccttttgaag aataggaact tcgaattcga tatcaagctt atcgataatc aacctctgga
     2941 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg
     3001 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt
     3061 ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag
     3121 gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc
     3181 caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga
     3241 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
     3301 ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac
     3361 ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct
     3421 tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca
     3481 gacgagtcgg atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag
     3541 atctacgggt ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc
     3601 actccagtgc ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag
     3661 gtgtccttct ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg
     3721 aagacaacct gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa
     3781 tcttggctca ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc
     3841 gagttgttgg gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag
     3901 agacggggtt tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac
     3961 ccaccttggc ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc
     4021 cttctgattt tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg
     4081 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     4141 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc
     4201 aggggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     4261 gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
     4321 tacgcgcagc gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt
     4381 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
     4441 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga
     4501 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
     4561 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg
     4621 ctattctttt gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct
     4681 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg
     4741 cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac
     4801 acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt
     4861 gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag
     4921 acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc
     4981 ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     5041 ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata
     5101 atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt
     5161 tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc
     5221 tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat
     5281 ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct
     5341 atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca
     5401 ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg
     5461 catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa
     5521 cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg
     5581 ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga
     5641 cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg
     5701 cgaactactt actctagctt cccggcaaca attaatagac tggatggagg cggataaagt
     5761 tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg
     5821 agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc
     5881 ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca
     5941 gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc
     6001 atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat
     6061 cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     6121 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     6181 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     6241 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     6301 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     6361 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     6421 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     6481 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     6541 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     6601 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     6661 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     6721 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     6781 ctggcctttt gctcacatgt
//
