LOCUS       sequence-252565-        7642 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-dependent fusion protein of Gaussia luciferase variant slow 
            burn, Volvox channelrhodopsin-1, and enhanced yellow fluorescent 
            protein (luminopsin LMO3) for bioluminescent optogenetics.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7642)
  AUTHORS   Hochgeschwender, U
  TITLE     luciferase opsin fusion proteins for bioluminescent optogenetics
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 7642)
  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: 
            "unpublished"
COMMENT     Sequence Label: pAAV-EFIa-DIO-LMO3 (sbGLuc-VChR1-EYFP)
FEATURES             Location/Qualifiers
     source          1..7642
                     /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        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1440..1473)
                     /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(1524..1557)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(join(1605..2315,2316..2318))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1647..1668)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1996..2015
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2255..2276
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3284..3293
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(3333..3887)
                     /codon_start=1
                     /product="enhanced variant of secreted Gaussia luciferase"
                     /label=Gaussia-Dura Luc
                     /note="human codon-optimized"
                     /translation="MGVKVLFALICIAVAEAKPTENNEDFNIVAVASNFATTDLDADRG
                     KLPGKKLPLEVLKELEANARKAGCTRGCLICLSHIKCTPKMKKFIPGRCHTYEGDKESA
                     QGGIGEAIVDIPEIPGFKDLEPLEQFIAQVDLCVDCTTGCLKGLANVQCSDLLKKWLPQ
                     RCATFASKIQGQVDKIKGAGGD"
     CDS             complement(3333..3887)
                     /codon_start=1
                     /product="secreted Gaussia luciferase"
                     /label=hGLuc
                     /note="human codon-optimized"
                     /translation="MGVKVLFALICIAVAEAKPTENNEDFNIVAVASNFATTDLDADRG
                     KLPGKKLPLEVLKELEANARKAGCTRGCLICLSHIKCTPKMKKFIPGRCHTYEGDKESA
                     QGGIGEAIVDIPEIPGFKDLEPLEQFIAQVDLCVDCTTGCLKGLANVQCSDLLKKWLPQ
                     RCATFASKIQGQVDKIKGAGGD"
     regulatory      3884..3893
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3912..3945
                     /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    3996..4029
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4054..4642
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4107..4127)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4525..4536)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4676..4787
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4905..5045
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4905..5034
                     /label=AAV2 ITR
     rep_origin      5120..5575
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5207..5226)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5417..5438
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5592..5611)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5711..5733
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5771..5789)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5857..5961
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5962..6030,6031..6822)
                     /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(6180..6199)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6993..7581
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7482..7501
                     /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 gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa
     1441 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
     1561 gcgccgctag ctaattctca ttacacctcg ttctcgtagc agaacttgta cagctcgtcc
     1621 atgccgagag tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc
     1681 tcgttggggt ctttgctcag ggcggactgg tagctcaggt agtggttgtc gggcagcagc
     1741 acggggccgt cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgctgccg
     1801 tcctcgatgt tgtggcggat cttgaagttc accttgatgc cgttcttctg cttgtcggcc
     1861 atgatataga cgttgtggct gttgtagttg tactccagct tgtgccccag gatgttgccg
     1921 tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac
     1981 ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg
     2041 acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag
     2101 cgggcgaagc actgcaggcc gtagccgaag gtggtcacga gggtgggcca gggcacgggc
     2161 agcttgccgg tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg
     2221 ccctcgccgg acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg
     2281 ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca cgttgatgtc gatctggtcc
     2341 agggggatgt actcgccctc gctggtgatc ctgctcttgg cggccgcgtc ctcctcttca
     2401 gccaccagtg tctcaacctc catctcttgg ccggcgatcg taattttctg cttctttctg
     2461 atgtcgccat acaacaggat gtgctcgtgg attttgacgc gcaggtaatt ccccagcaca
     2521 ccccacatgt tctttgcaat caggtcgagg atggagtgcc caatggcaga tccataaggg
     2581 ctgatatgac cgaatccttc cgtgcccagc agaaacagga ctggaaacat tccccaagcc
     2641 acaaagaatg tccaggccat cactcggacc agttctcgac aaatcccttt tggaactgtg
     2701 tggaatgctt cgatataaac tttagcagca tgaaaatagg tatacatacc atagctcaat
     2761 gagatgagga agaacaggat cttcgtccac cccgtacaca tggcgctggt tgcgccccac
     2821 acgatgcatc ccacatcaga aaccaacagg cccatagtcc ttttgctata atcgtccttc
     2881 aggcctgtga ggttgctcag atggatcaga aggaccgggc aggtgaggag ccattcgcca
     2941 taccgcatcc aaactacgcc gttccctgaa gacagccaca gtgttgcagg gctgtcgaac
     3001 tcgtggaagg cctcgatgat gcttttcatc atttctatca aggctacgta aacctcctcc
     3061 catccgcagg tagctcgcca cgcttgatac gcataccacc ccaggcaagc cactgacaaa
     3121 gcaaagacta cccactgaag agtaattgct attgtctttt caatggaagt gccgcgactc
     3181 cgaagccaac cttcacagta acactgtcct cttggcatgc agacggtgcc gtttccgagg
     3241 tcggttgggt agcgaactat gaggcttcta gcaacagggt aatccatggt ggcggatccg
     3301 gcaattccac cacactggac tagtgggtcg ccgtcaccac cggccccctt gatcttgtcc
     3361 acctggccct ggatcttgct ggcaaaggtc gcacagcgtt gcggcagcca cttcttgagc
     3421 aggtcagaac actgcacgtt ggcaagccct ttgaggcagc cagttgtgca gtccacacac
     3481 agatcgacct gtgcgatgaa ctgctccagg ggctccaagt ccttgaaccc aggaatctca
     3541 ggaatgtcga cgatcgcctc gcctatgccg ccctgtgcgg actctttgtc gccttcgtag
     3601 gtgtggcagc gtcctgggat gaacttcttc atcttgggcg tgcacttgat gtgggacagg
     3661 cagatcagac agcccctggt gcagccagct ttccgggcat tggcttccag ctctttgagc
     3721 acctccagcg gcagcttctt gccgggcaac ttcccgcggt cagcatcgag atccgtggtc
     3781 gcgaagttgc tggccacggc cacgatgttg aagtcttcgt tgttctcggt gggcttggcc
     3841 tcggccacag cgatgcagat cagggcaaac agaactttga ctcccatggt ggcaagctta
     3901 ccggtgctag cataacttcg tataaagtat cctatacgaa gttatttgcc ttaacccaga
     3961 aattatcact gttattcttt agaatggtgc aaagaataac ttcgtataat gtatgctata
     4021 cgaagttatg aattcgatat caagcttatc gataatcaac ctctggatta caaaatttgt
     4081 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     4141 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     4201 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     4261 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     4321 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     4381 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     4441 tcggggaaat catcgtcctt tccttggctg ctcgcctatg ttgccacctg gattctgcgc
     4501 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     4561 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     4621 tccctttggg ccgcctcccc gcatcgatac cgagcgctgc tcgagagatc ttcgactgtg
     4681 ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa
     4741 ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt
     4801 aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa
     4861 gacaatagca ggcatgagat ctcacgtgcg gaccgagcgg ccgcaggaac ccctagtgat
     4921 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     4981 cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct
     5041 gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     5101 acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     5161 gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc tttcgctttc
     5221 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     5281 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt
     5341 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     5401 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ctctatctcg
     5461 ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt aaaaaatgag
     5521 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg
     5581 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca
     5641 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     5701 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     5761 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
     5821 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     5881 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     5941 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     6001 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     6061 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     6121 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     6181 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     6241 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     6301 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     6361 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     6421 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     6481 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
     6541 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     6601 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     6661 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     6721 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     6781 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     6841 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     6901 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     6961 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     7021 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     7081 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     7141 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     7201 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     7261 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     7321 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     7381 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     7441 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     7501 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     7561 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     7621 tgctggcctt ttgctcacat gt
//
