LOCUS       sequence-142414-        7372 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent viral expression of ChR2-tdTomato for neuronal 
            excitation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7372)
  AUTHORS   Atasoy D, Aponte Y, Su HH, Sternson SM
  TITLE     A FLEX switch targets Channelrhodopsin-2 to multiple cell types for 
            imaging and long-range circuit mapping.
  JOURNAL   J Neurosci. 2008 Jul 9. 28(28):7025-30.
  PUBMED    18614669
REFERENCE   2  (bases 1 to 7372)
  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: "J Neurosci.
            2008 Jul 9. 28(28):7025-30."
COMMENT     Sequence Label: AAV-FLEX-rev-ChR2-tdtomato
FEATURES             Location/Qualifiers
     source          1..7372
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1932..2387)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2069..2090)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2281..2300
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2459..2588
                     /label=AAV2 ITR
     primer_bind     3412..3434
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3496..3515
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    3550..3583
                     /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    3646..3679
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(3693..5120)
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGGL
                     VTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIHQ
                     ALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRHH
                     LFLGHGIGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYE
                     GTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMN
                     FEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVL
                     KGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYER
                     SEGRHHLFLYGMDELYK"
     CDS             complement(5133..6059)
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIHLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             complement(5133..6059)
                     /codon_start=1
                     /product="N-terminal fragment of channelrhodopsin-2 from 
                     the alga Chlamydomonas reinhardtii (Nagel et al., 2003)"
                     /label=ChR2
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIHLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      6056..6065
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(6095..6128)
                     /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(6199..6232)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    6274..6862
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6327..6347)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6745..6756)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    6905..7026
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6916..6935)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6970..6989
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   7188..7317
                     /label=AAV2 ITR
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     1981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
     2401 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     2461 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     2521 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     2581 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag
     2641 gaagagtacc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     2701 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     2761 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
     2821 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
     2881 tcatcgctat taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc
     2941 ccccctcccc acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg
     3001 gggcgggggg gggggggggg cgaggcggag aggtgcggcg gcagccaatc agagcggcgc
     3061 gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata aaaagcgaag
     3121 cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc
     3181 ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga gcgggcggga
     3241 cggcccttct cctccgggct gtaattagcg cttggtttaa tgacggcttg tttcttttct
     3301 gtggctgcgt gaaagccttg aggggctccg ggagggccct ttgtgcgggg ggagcggctc
     3361 ggggctgtcc gcggggggac ggctgccttc gggggggacg gggcagggcg gggttcggct
     3421 tctggcgtgt gaccggcggc tctagagcct ctgctaacca tgttcatgcc ttcttctttt
     3481 tcctacagct cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat
     3541 tggatcccca taacttcgta taaagtatcc tatacgaagt tatatcaaaa taggaagacc
     3601 aatgcttcac catcgacccg aattgccaag catcaccatc gacccataac ttcgtataat
     3661 gtatgctata cgaagttata ctagggatcc ccttacttgt acagctcgtc catgccgtac
     3721 aggaacaggt ggtggcggcc ctcggagcgc tcgtactgtt ccacgatggt gtagtcctcg
     3781 ttgtgggagg tgatgtccag cttggtgtcc acgtagtagt agccgggcag ttgcacgggc
     3841 ttcttggcca tgtagatggt cttgaactcc accaggtagt ggccgccgtc cttcagcttc
     3901 agggcctggt ggatctcgcc cttcagcacg ccgtcgcggg ggtacaggcg ctcggtggag
     3961 gcctcccagc ccatggtctt cttctgcatt acggggccgt cgggggggaa gttggtgccg
     4021 cgcatcttca ccttgtagat cagcgtgccg tcctgcaggg aggagtcctg ggtcacggtc
     4081 accagaccgc cgtcctcgaa gttcatcacg cgctcccact tgaagccctc ggggaaggac
     4141 agcttcttgt aatcggggat gtcggcgggg tgcttcacgt acgccttgga gccgtacatg
     4201 aactgggggg acaggatgtc ccaggcgaag ggcagggggc cgcccttggt caccttcagc
     4261 ttggcggtct gggtgccctc gtaggggcgg ccctcgccct cgccctcgat ctcgaactcg
     4321 tggccgttca tggagccctc catgcgcacc ttgaagcgca tgaactcttt gatgacggcc
     4381 atgttgttgt cctcggagga ggcggtgccg gagctgccgc tgccggtgct gccgatgcca
     4441 tgccccagga acaggtggtg gcggccctcg gagcgctcgt actgttccac gatggtgtag
     4501 tcctcgttgt gggaggtgat gtccagcttg gtgtccacgt agtagtagcc gggcagttgc
     4561 acgggcttct tggccatgta gatggtcttg aactccacca ggtagtggcc gccgtccttc
     4621 agcttcaggg cctggtggat ctcgcccttc agcacgccgt cgcgggggta caggcgctcg
     4681 gtggaggcct cccagcccat ggtcttcttc tgcattacgg ggccgtcggg ggggaagttg
     4741 gtgccgcgca tcttcacctt gtagatcagc gtgccgtcct gcagggagga gtcctgggtc
     4801 acggtcacca gaccgccgtc ctcgaagttc atcacgcgct cccacttgaa gccctcgggg
     4861 aaggacagct tcttgtaatc ggggatgtcg gcggggtgct tcacgtacgc cttggagccg
     4921 tacatgaact ggggggacag gatgtcccag gcgaagggca gggggccgcc cttggtcacc
     4981 ttcagcttgg cggtctgggt gccctcgtag gggcggccct cgccctcgcc ctcgatctcg
     5041 aactcgtggc cgttcatgga gccctccatg cgcaccttga agcgcatgaa ctctttgatg
     5101 acctcctcgc ccttgctcac ggccgcggct ggcacggctc cggcctcggc ttcgtcttcg
     5161 acgagagtct cgacctcgat ctccgttccg ccgatgttca gtttggtggt tttgcggata
     5221 tctccgtgaa tcaatatgtg ctcgtggatc aggacgcgca ggtagtgtcc caacaacccc
     5281 caacaatttt tactcatcag atcaataatc gtgtgaccta cggtggagcc atagacgctc
     5341 aggacgccaa aaccttcggg ccccaaaatg aagagaattg ggaacatacc ccagctcacg
     5401 aaaaacagcc atgccatgcc ggtcacgacc tggcggcacc gaccctttgg cacagtatga
     5461 taaccctcga tatatgcttt ggcggcgtga aaaaatgtgt tcgcgccata gcacaatcca
     5521 agacaaaaga agatgacttt aacatagccg gttgccatgg cgctggtagc cccccacacg
     5581 atagtcccga tgtctgagac aaggagtccc atggttctcc tgctgtagtc gttgctcagg
     5641 ccggtgaggt tgctcaggtg gataaggatg acaggacaag tgagcagcca ctctgcatag
     5701 cgcagccact gcacccggtg tcctgtggca aggtagagca tagagggatt cttaaactca
     5761 aaaaagaact cgagaatcac cttaaccatt tcaatggcgc acacatagat ctcctcccag
     5821 ccgcatgtag atttccaggt ttggtaggca tagaacatca gcagcaaaat gctgaatcct
     5881 gctgcaagcc actgcaggac atttgacgcg gtctgagcgc cgttcgtgcc gcgagattca
     5941 atccatccgg cacagtaaca ttgatcctca gggaccagga cggacccgtt caccaccaca
     6001 ggattagtaa cgaacaaaag ttcgcgtccg acggcagaca aagcgccgcc atagtccatg
     6061 gtggcgctag gggcccgcgg taccgaattc tagtataact tcgtatagga tactttatac
     6121 gaagttatca ttgggattct tcctattttg atccaagcat caccatcgac cctctagtcc
     6181 agatctcacc atcgacccat aacttcgtat agcatacatt atacgaagtt atgtccctcg
     6241 aagaggttcg aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     6301 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     6361 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     6421 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     6481 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     6541 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     6601 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     6661 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     6721 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     6781 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     6841 tccctttggg ccgcctcccc gcatcgatac cgtcgacccg ggcggccgct tcgagcagac
     6901 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
     6961 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     7021 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga gatgtgggag
     7081 gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tcttcctaga
     7141 gcatggctac gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt
     7201 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     7261 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     7321 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ct
//
