LOCUS       sequence-403388-        6995 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Flp-dependent Cre and mScarlet-KASH expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6995)
  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 6995)
  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-Syn1-fDIO-mScarlet-KASH-2a-Cre
FEATURES             Location/Qualifiers
     source          1..6995
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        9..456
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    487..520
                     /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(613..1641)
                     /codon_start=1
                     /product="site-specific recombinase"
                     /label=Cre
                     /note="Cre is a site-specific recombinase from 
                     bacteriophage P1. Recombination occurs at loxP sequences."
                     /translation="SNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKMLL
                     SVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGLP
                     RPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAFL
                     GIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVER
                     WISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQR
                     YLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLED
                     GD"
     CDS             complement(1642..1662)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1678..1731)
                     /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(2005..2700)
                     /codon_start=1
                     /product="bright monomeric red fluorescent protein evolved 
                     from a synthetic template (Bindels et al., 2016)"
                     /label=mScarlet
                     /translation="MVSKGEAVIKEFMRFKVHMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFSWDILSPQFMYGSRAFTKHPADIPDYYKQSFPEGFKWERVMNFEDGG
                     AVTVTQDTSLEDGTLIYKVKLRGTNFPPDGPVMQKKTMGWEASTERLYPEDGVLKGDIK
                     MALRLKDGGRYLADFKTTYKAKKPVQMPGAYNVDRKLDITSHNEDYTVVEQYERSEGRH
                     STGGMDELYK"
     primer_bind     complement(2280..2299)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2549..2567
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2701..2730)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      2730..2739
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(2746..2779)
                     /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    2888..3476
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2941..2961)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3359..3370)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3508..3984
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3707..3726)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4024..4164
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4024..4153
                     /label=AAV2 ITR
     rep_origin      4239..4694
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4326..4345)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4536..4557
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4711..4730)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4830..4852
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4890..4908)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4976..5080
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5081..5149,5150..5941)
                     /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(5299..5318)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6112..6700
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6601..6620
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   6762..6891
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 tgcgtatgag tgcaagtggg ttttaggacc aggatgaggc ggggtggggg tgcctacctg
       61 acgaccgacc ccgacccact ggacaagcac ccaaccccca ttccccaaat tgcgcatccc
      121 ctatcagaga gggggagggg aaacaggatg cggcgaggcg cgtgcgcact gccagcttca
      181 gcaccgcgga cagtgccttc gcccccgcct ggcggcgcgc gccaccgccg cctcagcact
      241 gaaggcgcgc tgacgtcact cgccggtccc ccgcaaactc cccttcccgg ccaccttggt
      301 cgcgtccgcg ccgccgccgg cccagccgga ccgcaccacg cgaggcgcga gatagggggg
      361 cacgggcgcg accatctgcg ctgcggcgcc ggcgactcag cgctgcctca gtctgcggtg
      421 ggcagcggag gagtcgtgtc gtgcctgaga gcgcagggta ccggatcctc tagagtcgac
      481 tccggagaag ttcctattct ctagaaagta taggaacttc gcagaatggt agctggattg
      541 tagctgctat tagcaatatg aaacctctta gaagttccta ttcttcaaaa ggtataggaa
      601 cttcggcgcg ccctaatcgc catcttccag caggcgcacc attgcccctg tttcactatc
      661 caggttacgg atatagttca tgacaatatt tacattggtc cagccaccag cttgcatgat
      721 ctccggtatt gaaactccag cgcgggccat atctcgcgcg gctccgacac gggcactgtg
      781 tccagaccag gccaggtatc tctgaccaga gtcatcctta gcgccgtaaa tcaatcgatg
      841 agttgcttca aaaatccctt ccagggcgcg agttgatagc tggctggtgg cagatggcgc
      901 ggcaacacca ttttttctga cccggcaaaa caggtagtta ttcggatcat cagctacacc
      961 agagacggaa atccatcgct cgaccagttt agttaccccc aggctaagtg ccttctctac
     1021 acctgcggtg ctaaccagcg ttttcgttct gccaatatgg attaacattc tcccaccgtc
     1081 agtacgtgag atatctttaa ccctgatcct ggcaatttcg gctatacgta acagggtgtt
     1141 ataagcaatc cccagaaatg ccagattacg tatatcctgg cagcgatcgc tattttccat
     1201 gagtgaacga acctggtcga aatcagtgcg ttcgaacgct agagcctgtt ttgcacgttc
     1261 accggcatca acgttttctt ttcggatccg ccgcataacc agtgaaacag cattgctgtc
     1321 acttggtcgt ggcagcccgg accgacgatg aagcatgttt agctggccca aatgttgctg
     1381 gatagttttt actgccagac cgcgcgcctg aagatataga agataatcgc gaacatcttc
     1441 aggttctgcg ggaaaccatt tccggttatt caacttgcac catgccgccc acgaccggca
     1501 aacggacaga agcattttcc aggtatgctc agaaaacgcc tggcgatccc tgaacatgtc
     1561 catcaggttc ttgcgaacct catcactcgt tgcatcgacc ggtaatgcag gcaaattttg
     1621 gtgtacggtc agtaaattgg acaccttcct cttcttcttg ggggttctga aagcgcttgg
     1681 gccaggattc tcctcgacgt caccgcatgt tagcagactt cctctgccct ctccactgcc
     1741 ggtgggaggt ggcccgttgg tgtaccgcag catggggtag aaggatcggg caaagttgtt
     1801 ggcctgggtg cagctgtagt catcttcaga ggcaggtagc aggcaggcca ggagcagcag
     1861 cagcagcaga agcagctgca acggtagcgc tgccctgatc acccttgaga ggaaggagcg
     1921 tctcggctgg gagctgccgg ggctgtcgag gtggggcatc ctgctgtctg tctcctcctc
     1981 ctcctctcga gatctgagtc cggacttgta cagctcgtcc atgccgccgg tggagtggcg
     2041 gccctcggag cgttcgtact gttccaccac ggtgtagtcc tcgttgtggg aggtgatgtc
     2101 caacttgcgg tcgacgttgt aggcgccggg catctgcacg ggcttcttgg ccttgtaggt
     2161 ggtcttgaag tccgccaggt agcgtccgcc gtccttcagg cgcagggcca tcttaatgtc
     2221 gcccttcagc acgccgtcct cggggtacaa ccgctcggtg gacgcttccc agcccattgt
     2281 cttcttctgc attacggggc cgtcaggagg gaagttggtg ccgcggagct tcaccttgta
     2341 gatcagggtg ccgtcctcca gggaggtgtc ctgggtcacg gtcacggcgc cgccgtcctc
     2401 gaagttcatc acgcgctccc acttgaagcc ctcggggaag gactgcttat agtagtcggg
     2461 gatgtcggcg gggtgcttgg tgaaggccct ggagccgtac atgaactgag gggacaggat
     2521 gtcccaggag aagggcaggg ggccaccctt ggtcaccttc agcttggcgg tctgggtgcc
     2581 ctcgtagggg cggccctcgc cctcgccctc gatctcgaac tcgtggccgt tcatggagcc
     2641 ctccatgtgc accttgaacc gcatgaactc cttgatcact gcctcgccct tgctcaccat
     2701 cagatcttct tcgctaatca gtttctgttc catggtggcg ctagcgaagt tcctatactt
     2761 tctagagaat aggaacttct tgccttaacc cagaaattat cactgttatt ctttagaatg
     2821 gtgcaaagag aagttcctat accttttgaa gaataggaac ttcgaattcg atatcaagct
     2881 tatcgataat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     2941 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     3001 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     3061 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     3121 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     3181 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     3241 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg
     3301 gctgctcgcc tatgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     3361 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     3421 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg
     3481 ataccgagcg ctgctcgaga gatctacggg tggcatccct gtgacccctc cccagtgcct
     3541 ctcctggccc tggaagttgc cactccagtg cccaccagcc ttgtcctaat aaaattaagt
     3601 tgcatcattt tgtctgacta ggtgtccttc tataatatta tggggtggag gggggtggta
     3661 tggagcaagg ggcaagttgg gaagacaacc tgtagggcct gcggggtcta ttgggaacca
     3721 agctggagtg cagtggcaca atcttggctc actgcaatct ccgcctcctg ggttcaagcg
     3781 attctcctgc ctcagcctcc cgagttgttg ggattccagg catgcatgac caggctcagc
     3841 taatttttgt ttttttggta gagacggggt ttcaccatat tggccaggct ggtctccaac
     3901 tcctaatctc aggtgatcta cccaccttgg cctcccaaat tgctgggatt acaggcgtga
     3961 accactgctc ccttccctgt ccttctgatt ttgtaggtaa ccacgtgcgg accgagcggc
     4021 cgcaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc tcgctcactg
     4081 aggccgggcg accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg
     4141 agcgagcgcg cagctgcctg caggggcgcc tgatgcggta ttttctcctt acgcatctgt
     4201 gcggtatttc acaccgcata cgtcaaagca accatagtac gcgccctgta gcggcgcatt
     4261 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccttagc
     4321 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     4381 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     4441 caaaaaactt gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     4501 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     4561 aacactcaac tctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggt
     4621 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt
     4681 aacgtttaca attttatggt gcactctcag tacaatctgc tctgatgccg catagttaag
     4741 ccagccccga cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc
     4801 atccgcttac agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc
     4861 gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa
     4921 tgtcatgata ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg
     4981 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     5041 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     5101 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     5161 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     5221 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     5281 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     5341 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     5401 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     5461 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     5521 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     5581 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     5641 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     5701 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     5761 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     5821 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     5881 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     5941 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     6001 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     6061 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
     6121 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
     6181 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
     6241 gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc
     6301 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
     6361 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
     6421 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
     6481 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
     6541 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
     6601 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
     6661 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
     6721 tttacggttc ctggcctttt gctggccttt tgctcacatg tcctgcaggc agctgcgcgc
     6781 tcgctcgctc actgaggccg cccgggcgtc gggcgacctt tggtcgcccg gcctcagtga
     6841 gcgagcgagc gcgcagagag ggagtggcca actccatcac taggggttcc tgcggccgca
     6901 cgcgtaagct ttgcaaagat ggataaagtt ttaaacagag aggaatcttt gcagctaatg
     6961 gaccttctag gtcttgaaag gagtgggaat tggct
//
