LOCUS       sequence-204715-        7327 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of Chronos-tdTomato under the CAG promoter 
            in frt/reversed (Flp-dependent) manner..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7327)
  AUTHORS   Klapoetke NC, Murata Y, Kim SS, Pulver SR, Birdsey-Benson A, Cho YK,
            Morimoto TK, Chuong AS, Carpenter EJ, Tian Z, Wang J, Xie Y, Yan Z, 
            Zhang Y, Chow BY, Surek B, Melkonian M, Jayaraman V, 
            Constantine-Paton M, Wong GK, Boyden ES
  TITLE     Independent optical excitation of distinct neural populations.
  JOURNAL   Nat Methods. 2014 Mar;11(3):338-46. doi: 10.1038/nmeth.2836. Epub 
            2014 Feb 9.
  PUBMED    24509633
REFERENCE   2  (bases 1 to 7327)
  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.2836"; journalName: "Nat Methods"; date: "2014-03"; 
            volume: "11"; issue: "3"; pages: "338-46"
COMMENT     Sequence Label: AAV-CAG-FLPX-rc [Chronos-tdTomato]
FEATURES             Location/Qualifiers
     source          1..7327
                     /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        387..664
                     /label=chicken beta-actin promoter
     primer_bind     944..966
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1028..1047
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    1080..1113
                     /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)"
     regulatory      3624..3633
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3645..3678)
                     /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    3768..4356
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3821..3841)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4239..4250)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4364..4588
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4590..4730
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4590..4719
                     /label=AAV2 ITR
     rep_origin      4805..5260
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4892..4911)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5102..5123
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5277..5296)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5396..5418
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5456..5474)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5542..5646
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5647..5715,5716..6507)
                     /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(5865..5884)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6678..7266
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7167..7186
                     /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 gtcgacccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
      181 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
      241 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
      301 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
      361 cgtattagtc atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc
      421 catctccccc ccctccccac ccccaatttt gtatttattt attttttaat tattttgtgc
      481 agcgatgggg gcgggggggg ggggggggcg cgcgccaggc ggggcggggc ggggcgaggg
      541 gcggggcggg gcgaggcgga gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa
      601 gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg
      661 ggcgggagtc gctgcgacgc tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc
      721 gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg gacggccctt
      781 ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt ctgtggctgc
      841 gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc tcggggctgt
      901 ccgcgggggg acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt
      961 gtgaccggcg gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag
     1021 ctcctgggca acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attgaattcg
     1081 aagttcctat tctctagaaa gtataggaac ttcgcagaat ggtagctgga ttgtagctgc
     1141 tattagcaat atgaaacctc ttagaagttc ctattcttca aaaggtatag gaacttcgct
     1201 agctcattac ttatacagct catccatgcc gtacagaaac agatggtggc gtccttcaga
     1261 cctctcatac tgttccacaa tggtgtagtc ttcattgtgg gatgtgatat ccagcttggt
     1321 gtccacatag taatagccgg gcagctgcac tggtttcttg gccatgtaga tggttttgaa
     1381 ctccaccagg tagtggccgc cgtctttcag cttcagggcc tggtggattt cgcccttcag
     1441 cactccgtct ctaggataca gccgttcagt gctggcctcc caccccattg ttttcttctg
     1501 catcacgggc ccatcgggtg gaaaattggt gccgcgcatt ttgactttgt agatcagagt
     1561 gccatcctgc agagaggagt cctgtgtcac ggtcaccagg cctccatctt caaaattcat
     1621 cactctctcc cacttaaagc cctctgggaa agacagtttc ttgtaatcag gaatgtcagc
     1681 gggatgcttg acataggcct tacttccgta cataaactga ggagacagga tatcccaagc
     1741 gaaaggcagt ggtcctccct tggtcacttt cagcttggca gtctgtgtgc cctcgtaagg
     1801 cctcccttcg ccctctcctt caatctcaaa ttcgtgccca ttcatgctgc cctccatgcg
     1861 caccttaaac ctcatgaact ctttgatcac agccatattg ttatcctcgc ttgaggcggt
     1921 tccactagac cctgagccgg tacttccagt cccatggccc aggaacagat ggtgtctccc
     1981 ctcgctccgt tcatactgct ccacaatagt gtagtcctcg ttgtgtgaag tgatatccag
     2041 ttttgtgtcc acgtaatagt agcctggcag ctgcacaggt ttcttggcca tgtagattgt
     2101 cttgaactcc accaggtaat gtcccccatc tttcagcttc agagcctggt ggatttctcc
     2161 tttcagcacc ccgtcgcgtg gatacagcct ttcggtggag gcctcccatc ccatagtttt
     2221 cttctgcatc acaggtccat cagggggaaa atttgtccct ctcattttca ccttgtagat
     2281 cagggtccca tcctgcaggg agctgtcctg ggtcacagtc accaggcctc cgtcctcgaa
     2341 gttcatcact ctttcccact taaacccctc ggggaagctc agtttcttat aatcgggaat
     2401 gtcagctgga tgtttcacat aggccttgga gccgtacata aactgtgggc tcaggatgtc
     2461 ccaagcgaat ggcagaggcc cgccctttgt cactttcagc ttggctgtct gggtgccctc
     2521 gtagggccgt ccctctcctt ctccctcaat ttcgaactcg tggccgttca tgcttccttc
     2581 cattctcacc ttaaaccgca tgaactcttt gatcacttcc tcgcccttac tcactgctac
     2641 taccggtgcc gccactcctc cctcctcctc ctcgtcgacg aaggtctcga tctccatgtt
     2701 ctcgccggcc acgttgatgg tggttgtctt ccggatgtcg ccgtggatca ggatgtgctc
     2761 gtggatcttc actctcaggt ggtggcccag gaagccccaa gtgttcttgc tgatcagatc
     2821 caggatcagg tggccgattc cgctggtgta aggtgtgatc aggcccagtc cctcgtgtcc
     2881 ggcgatgaac atcacgggga acatcagcca agagcagaag aagacgtagg ccatgatctt
     2941 gcaaatcttc cggcagacgc ctttgggcag ggtgtggtag ctctcgatat acaccttggc
     3001 gatctggaag aagcaagtca cgccgtagaa caggccgatc atgaagaaca ggatcttcag
     3061 ggggcccttt gtaaaggcgg ctgtgatccc ccacacgatg ttgccgatgt cggtcaccag
     3121 gatggtcatg gtccgcttgc tgtactcttc gtgcagtccg gtcaggttgc tcaggtggat
     3181 caggatcacg gggcaagtca ggagccacat gctgtaccgc agccaaatca cggctcctcc
     3241 gttggtctgg tacactgtgg ctgggctgtc gacctcgtgg aacagctcga tgaagcactt
     3301 gacgagctcg atcacgcaga cgtagacctc ctcccagccg gttgtggcct tgaagctgtg
     3361 ccaaccgtag aagatgagct gcacgatagc cacgatcacg gtgaaccact ggagtcccac
     3421 tgcgatcttg tgctgcagct cggttccgtg gttgatgtgg gggaagcaac cgtgatcggc
     3481 tccggctgtt gtggcgttag aggtttctcc gtgagcgtct gctgctggtg tcaccactgc
     3541 tgcggcgctc agcaggcctc taattgtggc ttcggcgcta ggcacggctg agataaaggc
     3601 gtgggtcatt gtggcggctg tttccatggt ggcggcggcg cgccgaagtt cctatacttt
     3661 ctagagaata ggaacttctt gccttaaccc agaaattatc actgttattc tttagaatgg
     3721 tgcaaagaga agttcctata ccttttgaag aataggaact tctccggaat caacctctgg
     3781 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     3841 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     3901 tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca
     3961 ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg
     4021 ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg
     4081 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     4141 attccgtggt gttgtcgggg aagctgacgt cctttccatg gctgctcgcc tgtgttgcca
     4201 cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc
     4261 ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc
     4321 agacgagtcg gatctccctt tgggccgcct ccccgcctgg atcctgtgcc ttctagttgc
     4381 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     4441 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     4501 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     4561 catgctgggg atgcggtggg ctctatggca ggaaccccta gtgatggagt tggccactcc
     4621 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     4681 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     4741 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     4801 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     4861 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     4921 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     4981 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     5041 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     5101 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     5161 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     5221 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     5281 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     5341 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     5401 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     5461 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5521 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5581 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     5641 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     5701 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5761 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5821 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5881 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5941 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     6001 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     6061 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     6121 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     6181 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     6241 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     6301 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6361 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6421 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6481 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6541 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6601 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     6661 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6721 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6781 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6841 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6901 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6961 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     7021 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     7081 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     7141 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     7201 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     7261 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     7321 cacatgt
//
