LOCUS       sequence-46522-e        7082 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses humanized ChR2(H134R)-mCherry in cells lacking Cre 
            (Cre-Off, flanked by alternative lox FAS sites), for optogenetic 
            excitation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7082)
  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 7082)
  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-FAS-hChR2(H134R)-mCherry-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..7082
                     /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"
     regulatory      1492..1501
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1498..2424
                     /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             2437..3144
                     /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(2579..2597)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2847..2866
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    3209..3797
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3262..3282)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3680..3691)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3829..4305
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4028..4047)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4345..4485
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4345..4474
                     /label=AAV2 ITR
     rep_origin      4560..5015
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4647..4666)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4857..4878
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5032..5051)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5151..5173
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5211..5229)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5297..5401
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5402..5470,5471..6262)
                     /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(5620..5639)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6433..7021
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6922..6941
                     /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 caacttcgta tatacctttc tatacgaagt tgtggcgcgc cgccaccatg
     1501 gactatggcg gcgctttgtc tgccgtcgga cgcgaacttt tgttcgttac taatcctgtg
     1561 gtggtgaacg ggtccgtcct ggtccctgag gatcaatgtt actgtgccgg atggattgaa
     1621 tctcgcggca cgaacggcgc tcagaccgcg tcaaatgtcc tgcagtggct tgcagcagga
     1681 ttcagcattt tgctgctgat gttctatgcc taccaaacct ggaaatctac atgcggctgg
     1741 gaggagatct atgtgtgcgc cattgaaatg gttaaggtga ttctcgagtt cttttttgag
     1801 tttaagaatc cctctatgct ctaccttgcc acaggacacc gggtgcagtg gctgcgctat
     1861 gcagagtggc tgctcacttg tcctgtcatc cttatccgcc tgagcaacct caccggcctg
     1921 agcaacgact acagcaggag aaccatggga ctccttgtct cagacatcgg gactatcgtg
     1981 tggggggcta ccagcgccat ggcaaccggc tatgttaaag tcatcttctt ttgtcttgga
     2041 ttgtgctatg gcgcgaacac attttttcac gccgccaaag catatatcga gggttatcat
     2101 actgtgccaa agggtcggtg ccgccaggtc gtgaccggca tggcatggct gtttttcgtg
     2161 agctggggta tgttcccaat tctcttcatt ttggggcccg aaggttttgg cgtcctgagc
     2221 gtctatggct ccaccgtagg tcacacgatt attgatctga tgagtaaaaa ttgttggggg
     2281 ttgttgggac actacctgcg cgtcctgatc cacgagcaca tattgattca cggagatatc
     2341 cgcaaaacca ccaaactgaa catcggcgga acggagatcg aggtcgagac tctcgtcgaa
     2401 gacgaagccg aggccggagc cgtgccagcg gccgccgtga gcaagggcga ggaggataac
     2461 atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc cgtgaacggc
     2521 cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac ccagaccgcc
     2581 aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct gtcccctcag
     2641 ttcatgtacg gctccaaggc ctacgtgaag caccccgccg acatccccga ctacttgaag
     2701 ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga cggcggcgtg
     2761 gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa ggtgaagctg
     2821 cgcggcacca acttcccctc cgacggcccc gtaatgcaga agaagaccat gggctgggag
     2881 gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat caagcagagg
     2941 ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta caaggccaag
     3001 aagcccgtgc agctgcccgg cgcctacaac gtcaacatca agttggacat cacctcccac
     3061 aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca ctccaccggc
     3121 ggcatggacg agctgtacaa gtaagctagc acaacttcgt atataccttt ctatacgaag
     3181 ttgtgaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     3241 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     3301 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     3361 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     3421 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     3481 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     3541 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     3601 gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc tgcgcgggac
     3661 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     3721 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     3781 ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctacgg gtggcatccc
     3841 tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt gcccaccagc
     3901 cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt ctataatatt
     3961 atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac ctgtagggcc
     4021 tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct cactgcaatc
     4081 tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt gggattccag
     4141 gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg tttcaccata
     4201 ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg gcctcccaaa
     4261 ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat tttgtaggta
     4321 accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc
     4381 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg
     4441 cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt
     4501 attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta
     4561 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     4621 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     4681 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4741 tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc
     4801 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4861 actcttgttc caaactggaa caacactcaa ccctatctcg ggctattctt ttgatttata
     4921 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4981 cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg
     5041 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     5101 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     5161 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     5221 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     5281 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     5341 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     5401 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     5461 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     5521 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5581 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5641 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5701 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5761 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5821 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5881 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5941 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     6001 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     6061 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     6121 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     6181 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     6241 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     6301 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     6361 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     6421 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     6481 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6541 ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt agccgtagtt
     6601 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6661 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6721 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6781 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6841 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6901 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6961 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     7021 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     7081 gt
//
