LOCUS       sequence-50211-e        8001 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ChETA-TdTomato in cells lacking Cre (Cre-Off), for 
            optogenetic excitation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8001)
  AUTHORS   Saunders A, Johnson CA, Sabatini BL
  TITLE     Novel recombinant adeno-associated viruses for Cre activated and 
            inactivated transgene expression in neurons.
  JOURNAL   Front Neural Circuits. 2012 Jul 27;6:47. doi: 
            10.3389/fncir.2012.00047. eCollection 2012.
  PUBMED    22866029
REFERENCE   2  (bases 1 to 8001)
  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: "Front 
            Neural Circuits."; date: "2012-07-27"; volume: "6:47. doi"; pages: "
            10.3389/fncir.2012.00047. eCollection 2012"
COMMENT     Sequence Label: pAAV-Ef1a-DO-ChETA-TdTomato-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..8001
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1450..1483)
                     /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(1534..1567)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      1579..1588
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1585..2511
                     /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
                     PSMLYLATGHRVQWLRYATWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     regulatory      2521..2530
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2527..3957
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     protein_bind    3986..4019
                     /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    4070..4103
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4128..4716
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4181..4201)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4599..4610)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4748..5224
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4947..4966)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5264..5404
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5264..5393
                     /label=AAV2 ITR
     rep_origin      5479..5934
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5566..5585)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5776..5797
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5951..5970)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6070..6092
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6130..6148)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6216..6320
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6321..6389,6390..7181)
                     /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(6539..6558)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7352..7940
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7841..7860
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc
     1441 gactccggaa taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga
     1501 ttgtagctgc tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg
     1561 aagttatggc gcgccctagc caccatggac tatggcggcg ctttgtctgc cgtcggacgc
     1621 gaacttttgt tcgttactaa tcctgtggtg gtgaacgggt ccgtcctggt ccctgaggat
     1681 caatgttact gtgccggatg gattgaatct cgcggcacga acggcgctca gaccgcgtca
     1741 aatgtcctgc agtggcttgc agcaggattc agcattttgc tgctgatgtt ctatgcctac
     1801 caaacctgga aatctacatg cggctgggag gagatctatg tgtgcgccat tgaaatggtt
     1861 aaggtgattc tcgagttctt ttttgagttt aagaatccct ctatgctcta ccttgccaca
     1921 ggacaccggg tgcagtggct gcgctatgca acctggctgc tcacttgtcc tgtcatcctt
     1981 atccgcctga gcaacctcac cggcctgagc aacgactaca gcaggagaac catgggactc
     2041 cttgtctcag acatcgggac tatcgtgtgg ggggctacca gcgccatggc aaccggctat
     2101 gttaaagtca tcttcttttg tcttggattg tgctatggcg cgaacacatt ttttcacgcc
     2161 gccaaagcat atatcgaggg ttatcatact gtgccaaagg gtcggtgccg ccaggtcgtg
     2221 accggcatgg catggctgtt tttcgtgagc tggggtatgt tcccaattct cttcattttg
     2281 gggcccgaag gttttggcgt cctgagcgtc tatggctcca ccgtaggtca cacgattatt
     2341 gatctgatga gtaaaaattg ttgggggttg ttgggacact acctgcgcgt cctgatccac
     2401 gagcacatat tgattcacgg agatatccgc aaaaccacca aactgaacat cggcggaacg
     2461 gagatcgagg tcgagactct cgtcgaagac gaagccgagg ccggagccgt gccagcggcc
     2521 gccaccatgg tgagcaaggg cgaggaggtc atcaaagagt tcatgcgctt caaggtgcgc
     2581 atggagggct ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc
     2641 tacgagggca cccagaccgc caagctgaag gtgaccaagg gcggccccct gcccttcgcc
     2701 tgggacatcc tgtcccccca gttcatgtac ggctccaagg cgtacgtgaa gcaccccgcc
     2761 gacatccccg attacaagaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg
     2821 aacttcgagg acggcggtct ggtgaccgtg acccaggact cctccctgca ggacggcacg
     2881 ctgatctaca aggtgaagat gcgcggcacc aacttccccc ccgacggccc cgtaatgcag
     2941 aagaagacca tgggctggga ggcctccacc gagcgcctgt acccccgcga cggcgtgctg
     3001 aagggcgaga tccaccaggc cctgaagctg aaggacggcg gccactacct ggtggagttc
     3061 aagaccatct acatggccaa gaagcccgtg caactgcccg gctactacta cgtggacacc
     3121 aagctggaca tcacctccca caacgaggac tacaccatcg tggaacagta cgagcgctcc
     3181 gagggccgcc accacctgtt cctggggcat ggcaccggca gcaccggcag cggcagctcc
     3241 ggcaccgcct cctccgagga caacaacatg gccgtcatca aagagttcat gcgcttcaag
     3301 gtgcgcatgg agggctccat gaacggccac gagttcgaga tcgagggcga gggcgagggc
     3361 cgcccctacg agggcaccca gaccgccaag ctgaaggtga ccaagggcgg ccccctgccc
     3421 ttcgcctggg acatcctgtc cccccagttc atgtacggct ccaaggcgta cgtgaagcac
     3481 cccgccgaca tccccgatta caagaagctg tccttccccg agggcttcaa gtgggagcgc
     3541 gtgatgaact tcgaggacgg cggtctggtg accgtgaccc aggactcctc cctgcaggac
     3601 ggcacgctga tctacaaggt gaagatgcgc ggcaccaact tcccccccga cggccccgta
     3661 atgcagaaga agaccatggg ctgggaggcc tccaccgagc gcctgtaccc ccgcgacggc
     3721 gtgctgaagg gcgagatcca ccaggccctg aagctgaagg acggcggcca ctacctggtg
     3781 gagttcaaga ccatctacat ggccaagaag cccgtgcaac tgcccggcta ctactacgtg
     3841 gacaccaagc tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgag
     3901 cgctccgagg gccgccacca cctgttcctg tacggcatgg acgagctgta caagtaaggt
     3961 ggcggtggcg accggtagcg ctagcataac ttcgtataaa gtatcctata cgaagttatt
     4021 tgccttaacc cagaaattat cactgttatt ctttagaatg gtgcaaagaa taacttcgta
     4081 taatgtatgc tatacgaagt tatgaattcg atatcaagct tatcgataat caacctctgg
     4141 attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat
     4201 gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt
     4261 tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca
     4321 ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg
     4381 ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg
     4441 aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca
     4501 attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tatgttgcca
     4561 cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc
     4621 ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc
     4681 agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgagcg ctgctcgaga
     4741 gatctacggg tggcatccct gtgacccctc cccagtgcct ctcctggccc tggaagttgc
     4801 cactccagtg cccaccagcc ttgtcctaat aaaattaagt tgcatcattt tgtctgacta
     4861 ggtgtccttc tataatatta tggggtggag gggggtggta tggagcaagg ggcaagttgg
     4921 gaagacaacc tgtagggcct gcggggtcta ttgggaacca agctggagtg cagtggcaca
     4981 atcttggctc actgcaatct ccgcctcctg ggttcaagcg attctcctgc ctcagcctcc
     5041 cgagttgttg ggattccagg catgcatgac caggctcagc taatttttgt ttttttggta
     5101 gagacggggt ttcaccatat tggccaggct ggtctccaac tcctaatctc aggtgatcta
     5161 cccaccttgg cctcccaaat tgctgggatt acaggcgtga accactgctc ccttccctgt
     5221 ccttctgatt ttgtaggtaa ccacgtgcgg accgagcggc cgcaggaacc cctagtgatg
     5281 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     5341 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     5401 caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     5461 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     5521 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     5581 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     5641 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     5701 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     5761 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     5821 gctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     5881 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     5941 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     6001 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     6061 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     6121 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     6181 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     6241 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     6301 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     6361 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     6421 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     6481 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     6541 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     6601 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     6661 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     6721 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     6781 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     6841 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     6901 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     6961 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     7021 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     7081 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     7141 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     7201 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     7261 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     7321 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     7381 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     7441 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc
     7501 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     7561 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     7621 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     7681 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     7741 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     7801 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     7861 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     7921 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     7981 gctggccttt tgctcacatg t
//
