LOCUS       sequence-221389-        7326 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of Chronos-tdTomato under the CAG promoter, 
            in floxed/reversed (Cre-dependent) manner. Using bGHpA signal. 
            tdTomato is a codon diversified version..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7326)
  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 7326)
  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: pAAV-CAG-FLEX-rc [Chronos-tdTomato]
FEATURES             Location/Qualifiers
     source          1..7326
                     /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     943..965
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1027..1046
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1079..1112)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(1163..1196)
                     /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)."
     regulatory      3623..3632
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3644..3677
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3728..3761
                     /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)."
     misc_feature    3767..4355
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3820..3840)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4238..4249)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4363..4587
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4589..4729
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4589..4718
                     /label=AAV2 ITR
     rep_origin      4804..5259
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4891..4910)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5101..5122
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5276..5295)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5395..5417
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5455..5473)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5541..5645
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5646..5714,5715..6506)
                     /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(5864..5883)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6677..7265
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7166..7185
                     /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 cgcgccgggg gggggggggg gggggggggg
      541 gggggggggg ggggggcggg gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa
      601 gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg
      661 ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg
      721 cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc
      781 tcctccgggc tgtaattagc gcttggttta atgacggctt gtttcttttc tgtggctgcg
      841 tgaaagcctt gaggggctcc gggagggccc tttgtgcggg gggagcggct cggggctgtc
      901 cgcgggggga cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg
      961 tgaccggcgg ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc
     1021 tcctgggcaa cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa ttgaattcat
     1081 aacttcgtat agcatacatt atacgaagtt attctttgca ccattctaaa gaataacagt
     1141 gataatttct gggttaaggc aaataacttc gtataggata ctttatacga agttatgcta
     1201 gctcattact tatacagctc atccatgccg tacagaaaca gatggtggcg tccttcagac
     1261 ctctcatact gttccacaat ggtgtagtct tcattgtggg atgtgatatc cagcttggtg
     1321 tccacatagt aatagccggg cagctgcact ggtttcttgg ccatgtagat ggttttgaac
     1381 tccaccaggt agtggccgcc gtctttcagc ttcagggcct ggtggatttc gcccttcagc
     1441 actccgtctc taggatacag ccgttcagtg ctggcctccc accccattgt tttcttctgc
     1501 atcacgggcc catcgggtgg aaaattggtg ccgcgcattt tgactttgta gatcagagtg
     1561 ccatcctgca gagaggagtc ctgtgtcacg gtcaccaggc ctccatcttc aaaattcatc
     1621 actctctccc acttaaagcc ctctgggaaa gacagtttct tgtaatcagg aatgtcagcg
     1681 ggatgcttga cataggcctt acttccgtac ataaactgag gagacaggat atcccaagcg
     1741 aaaggcagtg gtcctccctt ggtcactttc agcttggcag tctgtgtgcc ctcgtaaggc
     1801 ctcccttcgc cctctccttc aatctcaaat tcgtgcccat tcatgctgcc ctccatgcgc
     1861 accttaaacc tcatgaactc tttgatcaca gccatattgt tatcctcgct tgaggcggtt
     1921 ccactagacc ctgagccggt acttccagtc ccatggccca ggaacagatg gtgtctcccc
     1981 tcgctccgtt catactgctc cacaatagtg tagtcctcgt tgtgtgaagt gatatccagt
     2041 tttgtgtcca cgtaatagta gcctggcagc tgcacaggtt tcttggccat gtagattgtc
     2101 ttgaactcca ccaggtaatg tcccccatct ttcagcttca gagcctggtg gatttctcct
     2161 ttcagcaccc cgtcgcgtgg atacagcctt tcggtggagg cctcccatcc catagttttc
     2221 ttctgcatca caggtccatc agggggaaaa tttgtccctc tcattttcac cttgtagatc
     2281 agggtcccat cctgcaggga gctgtcctgg gtcacagtca ccaggcctcc gtcctcgaag
     2341 ttcatcactc tttcccactt aaacccctcg gggaagctca gtttcttata atcgggaatg
     2401 tcagctggat gtttcacata ggccttggag ccgtacataa actgtgggct caggatgtcc
     2461 caagcgaatg gcagaggccc gccctttgtc actttcagct tggctgtctg ggtgccctcg
     2521 tagggccgtc cctctccttc tccctcaatt tcgaactcgt ggccgttcat gcttccttcc
     2581 attctcacct taaaccgcat gaactctttg atcacttcct cgcccttact cactgctact
     2641 accggtgccg ccactcctcc ctcctcctcc tcgtcgacga aggtctcgat ctccatgttc
     2701 tcgccggcca cgttgatggt ggttgtcttc cggatgtcgc cgtggatcag gatgtgctcg
     2761 tggatcttca ctctcaggtg gtggcccagg aagccccaag tgttcttgct gatcagatcc
     2821 aggatcaggt ggccgattcc gctggtgtaa ggtgtgatca ggcccagtcc ctcgtgtccg
     2881 gcgatgaaca tcacggggaa catcagccaa gagcagaaga agacgtaggc catgatcttg
     2941 caaatcttcc ggcagacgcc tttgggcagg gtgtggtagc tctcgatata caccttggcg
     3001 atctggaaga agcaagtcac gccgtagaac aggccgatca tgaagaacag gatcttcagg
     3061 gggccctttg taaaggcggc tgtgatcccc cacacgatgt tgccgatgtc ggtcaccagg
     3121 atggtcatgg tccgcttgct gtactcttcg tgcagtccgg tcaggttgct caggtggatc
     3181 aggatcacgg ggcaagtcag gagccacatg ctgtaccgca gccaaatcac ggctcctccg
     3241 ttggtctggt acactgtggc tgggctgtcg acctcgtgga acagctcgat gaagcacttg
     3301 acgagctcga tcacgcagac gtagacctcc tcccagccgg ttgtggcctt gaagctgtgc
     3361 caaccgtaga agatgagctg cacgatagcc acgatcacgg tgaaccactg gagtcccact
     3421 gcgatcttgt gctgcagctc ggttccgtgg ttgatgtggg ggaagcaacc gtgatcggct
     3481 ccggctgttg tggcgttaga ggtttctccg tgagcgtctg ctgctggtgt caccactgct
     3541 gcggcgctca gcaggcctct aattgtggct tcggcgctag gcacggctga gataaaggcg
     3601 tgggtcattg tggcggctgt ttccatggtg gcggcggcgc gccataactt cgtataatgt
     3661 atgctatacg aagttattaa gaggtttcat attgctaata gcagctacaa tccagctacc
     3721 attctgcata acttcgtata aagtatccta tacgaagtta ttccggaatc aacctctgga
     3781 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg
     3841 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt
     3901 ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag
     3961 gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc
     4021 caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga
     4081 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
     4141 ttccgtggtg ttgtcgggga agctgacgtc ctttccatgg ctgctcgcct gtgttgccac
     4201 ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct
     4261 tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca
     4321 gacgagtcgg atctcccttt gggccgcctc cccgcctgga tcctgtgcct tctagttgcc
     4381 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     4441 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     4501 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     4561 atgctgggga tgcggtgggc tctatggcag gaacccctag tgatggagtt ggccactccc
     4621 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     4681 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     4741 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     4801 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     4861 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     4921 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     4981 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     5041 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     5101 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     5161 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     5221 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     5281 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     5341 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     5401 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     5461 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     5521 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     5581 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     5641 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     5701 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     5761 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     5821 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     5881 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     5941 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     6001 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     6061 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     6121 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     6181 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     6241 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     6301 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     6361 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     6421 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     6481 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     6541 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     6601 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     6661 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     6721 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     6781 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     6841 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     6901 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     6961 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     7021 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     7081 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     7141 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     7201 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     7261 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     7321 acatgt
//
