LOCUS       sequence-46439-e        7260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent expression of hChR2(H134R) for optogenetic excitation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7260)
  AUTHORS   Kozorovitskiy Y, Saunders A, Johnson CA, Lowell BB, Sabatini BL
  TITLE     Recurrent network activity drives striatal synaptogenesis.
  JOURNAL   Nature. 2012 May 13;485(7400):646-50. doi: 10.1038/nature11052.
  PUBMED    22660328
REFERENCE   2  (bases 1 to 7260)
  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"; journalName: 
            "Nature"; date: "2012-05-13- 13"; volume: "485"; issue: "7400"; 
            pages: "646-50"
COMMENT     Sequence Label: pAAV-Ef1a-DO-hChR2(H134R)-mCherry-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..7260
                     /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      1578..1587
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1584..2510
                     /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
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             2523..3230
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2665..2683)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2933..2952
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     protein_bind    3245..3278
                     /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    3329..3362
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3387..3975
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3440..3460)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3858..3869)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4007..4483
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4206..4225)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4523..4663
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4523..4652
                     /label=AAV2 ITR
     rep_origin      4738..5193
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4825..4844)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5035..5056
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5210..5229)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5329..5351
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5389..5407)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5475..5579
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5580..5648,5649..6440)
                     /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(5798..5817)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6611..7199
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7100..7119
                     /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 gcgcctcgcc accatggact atggcggcgc tttgtctgcc gtcggacgcg
     1621 aacttttgtt cgttactaat cctgtggtgg tgaacgggtc cgtcctggtc cctgaggatc
     1681 aatgttactg tgccggatgg attgaatctc gcggcacgaa cggcgctcag accgcgtcaa
     1741 atgtcctgca gtggcttgca gcaggattca gcattttgct gctgatgttc tatgcctacc
     1801 aaacctggaa atctacatgc ggctgggagg agatctatgt gtgcgccatt gaaatggtta
     1861 aggtgattct cgagttcttt tttgagttta agaatccctc tatgctctac cttgccacag
     1921 gacaccgggt gcagtggctg cgctatgcag agtggctgct cacttgtcct gtcatcctta
     1981 tccgcctgag caacctcacc ggcctgagca acgactacag caggagaacc atgggactcc
     2041 ttgtctcaga catcgggact atcgtgtggg gggctaccag cgccatggca accggctatg
     2101 ttaaagtcat cttcttttgt cttggattgt gctatggcgc gaacacattt tttcacgccg
     2161 ccaaagcata tatcgagggt tatcatactg tgccaaaggg tcggtgccgc caggtcgtga
     2221 ccggcatggc atggctgttt ttcgtgagct ggggtatgtt cccaattctc ttcattttgg
     2281 ggcccgaagg ttttggcgtc ctgagcgtct atggctccac cgtaggtcac acgattattg
     2341 atctgatgag taaaaattgt tgggggttgt tgggacacta cctgcgcgtc ctgatccacg
     2401 agcacatatt gattcacgga gatatccgca aaaccaccaa actgaacatc ggcggaacgg
     2461 agatcgaggt cgagactctc gtcgaagacg aagccgaggc cggagccgtg ccagcgccgc
     2521 cggtgagcaa gggcgaggag gataacatgg ccatcatcaa ggagttcatg cgcttcaagg
     2581 tgcacatgga gggctccgtg aacggccacg agttcgagat cgagggcgag ggcgagggcc
     2641 gcccctacga gggcacccag accgccaagc tgaaggtgac caagggtggc cccctgccct
     2701 tcgcctggga catcctgtcc cctcagttca tgtacggctc caaggcctac gtgaagcacc
     2761 ccgccgacat ccccgactac ttgaagctgt ccttccccga gggcttcaag tgggagcgcg
     2821 tgatgaactt cgaggacggc ggcgtggtga ccgtgaccca ggactcctcc ctgcaggacg
     2881 gcgagttcat ctacaaggtg aagctgcgcg gcaccaactt cccctccgac ggccccgtaa
     2941 tgcagaagaa gaccatgggc tgggaggcct cctccgagcg gatgtacccc gaggacggcg
     3001 ccctgaaggg cgagatcaag cagaggctga agctgaagga cggcggccac tacgacgctg
     3061 aggtcaagac cacctacaag gccaagaagc ccgtgcagct gcccggcgcc tacaacgtca
     3121 acatcaagtt ggacatcacc tcccacaacg aggactacac catcgtggaa cagtacgaac
     3181 gcgccgaggg ccgccactcc accggcggca tggacgagct gtacaagtaa gcggccgcgc
     3241 tagcataact tcgtataaag tatcctatac gaagttattt gccttaaccc agaaattatc
     3301 actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct atacgaagtt
     3361 atgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
     3421 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     3481 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     3541 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     3601 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     3661 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     3721 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     3781 aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg cgcgggacgt
     3841 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     3901 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     3961 gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt ggcatccctg
     4021 tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc ccaccagcct
     4081 tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct ataatattat
     4141 ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct gtagggcctg
     4201 cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca ctgcaatctc
     4261 cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg gattccaggc
     4321 atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt tcaccatatt
     4381 ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc ctcccaaatt
     4441 gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt tgtaggtaac
     4501 cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
     4561 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
     4621 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
     4681 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
     4741 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4801 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4861 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     4921 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
     4981 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     5041 tcttgttcca aactggaaca acactcaacc ctatctcggg ctattctttt gatttataag
     5101 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5161 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     5221 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     5281 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     5341 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     5401 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     5461 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     5521 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     5581 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     5641 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     5701 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     5761 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     5821 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     5881 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     5941 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     6001 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     6061 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     6121 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     6181 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     6241 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     6301 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     6361 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     6421 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     6481 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     6541 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     6601 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     6661 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     6721 taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag ccgtagttag
     6781 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     6841 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     6901 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     6961 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     7021 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     7081 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     7141 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     7201 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
