LOCUS       sequence-481801-        7430 bp    DNA     circular SYN 05-AUG-2025
DEFINITION  The transfer plasmid for packaging AAV expressing Cre-dependent 
            chicken OPN5 was constructed by inserting the OPN5 coding sequence 
            into the multiple cloning site under the control of EF1-alpha 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7430)
  AUTHORS   Dai R, Yu T, Weng D, Li H, Cui Y, Wu Z, Guo Q, Zou H, Wu W, Gao X, 
            Qi Z, Ren Y, Wang S, Li Y, Luo M
  TITLE     A neuropsin-based optogenetic tool for precise control of Gq 
            signaling.
  JOURNAL   Sci China Life Sci. 2022 Jul;65(7):1271-1284. doi: 
            10.1007/s11427-022-2122-0. Epub 2022 May 12.
  PUBMED    35579776
REFERENCE   2  (bases 1 to 7430)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 5, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1007/s11427-022-2122-0"; journalName: "Sci China Life Sci"; 
            date: "2022-07"; volume: "65"; issue: "7"; pages: "1271-1284"
COMMENT     Sequence Label: pAAV-EF1-alpha-DIO-cOpn5-T2A-mcherry
FEATURES             Location/Qualifiers
     source          1..7430
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        881..2059
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1112..2050
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2007..2027
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(2090..2123)
                     /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(2174..2207)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(2219..2929)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2497..2516)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     2766..2784
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(2930..2983)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     protein_bind    4065..4098
                     /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    4149..4182
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4207..4795
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4260..4280)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4678..4689)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4827..5303
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(5026..5045)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5343..5483
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5343..5472
                     /label=AAV2 ITR
     rep_origin      5558..6013
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5645..5664)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5855..5876
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6030..6049)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6149..6171
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6209..6227)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6295..6399
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6400..6468,6469..7260)
                     /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(6618..6637)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggccgcac gcgtaagctt tgcaaagatg gataaagttt taaacagaga ggaatctttg
      841 cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg cccgtcagtg
      901 ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg gcaattgaac
      961 cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg
     1021 cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg tgaacgttct
     1081 ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt cccgcgggcc
     1141 tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct ggctgcagta
     1201 cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga ggccttgcgc
     1261 ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct ggggccgccg
     1321 cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt ctctagccat
     1381 ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc ttgtaaatgc
     1441 gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg acggggcccg
     1501 tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac cgagaatcgg
     1561 acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc cgccgtgtat
     1621 cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag cggaaagatg
     1681 gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct cgggagagcg
     1741 ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg tcgcttcatg
     1801 tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga gcttttggag
     1861 tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc acactgagtg
     1921 ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg aatttgccct
     1981 ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa gtttttttct
     2041 tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa taacttcgta
     2101 taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc tattagcaat
     2161 atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc gcgccttatt
     2221 acttgtacag ctcgtccatg ccgccggtgg agtggcggcc ctcggcgcgt tcgtactgtt
     2281 ccacgatggt gtagtcctcg ttgtgggagg tgatgtccaa cttgatgttg acgttgtagg
     2341 cgccgggcag ctgcacgggc ttcttggcct tgtaggtggt cttgacctca gcgtcgtagt
     2401 ggccgccgtc cttcagcttc agcctctgct tgatctcgcc cttcagggcg ccgtcctcgg
     2461 ggtacatccg ctcggaggag gcctcccagc ccatggtctt cttctgcatt acggggccgt
     2521 cggaggggaa gttggtgccg cgcagcttca ccttgtagat gaactcgccg tcctgcaggg
     2581 aggagtcctg ggtcacggtc accacgccgc cgtcctcgaa gttcatcacg cgctcccact
     2641 tgaagccctc ggggaaggac agcttcaagt agtcggggat gtcggcgggg tgcttcacgt
     2701 aggccttgga gccgtacatg aactgagggg acaggatgtc ccaggcgaag ggcagggggc
     2761 cacccttggt caccttcagc ttggcggtct gggtgccctc gtaggggcgg ccctcgccct
     2821 cgccctcgat ctcgaactcg tggccgttca cggagccctc catgtgcacc ttgaagcgca
     2881 tgaactcctt gatgatggcc atgttatcct cctcgccctt gctcaccatt gggccaggat
     2941 tctcctcgac gtcaccgcat gttagcagac ttcctctgcc ctcgacttcc agttgggttc
     3001 cgctcagtgc cgctgaatct ctgtgtgaag ataaggtgta catcctcgat tccttcaaag
     3061 agctcttctt ctgcagtgtc ttcggcccac ctgaccggca gcaggcaaat ttgcaatcaa
     3121 tgacctggta aataattggg ttgtacatgg cagccgactt ggcaagcaga gttggtacca
     3181 ccgaaaattg aatgggaacg gaatccggct gtccaaaggc tgaccacaca gagaccacgg
     3241 cgtaggggat ccaagcaatg aggaatccgg cacagatcag cattgccacc ttggtcagct
     3301 tcatttcaag tatgtggctg ttctggatcc tggtgtcata gtgagcaact tctttggtgg
     3361 acgacttgac ttttaaaata attttaacat aggagaacac aatcactgca gttggaaaca
     3421 ggaggcaaaa gaaaagaatg ctgagaacaa aagcctgccc agccactgaa gcctgcgcca
     3481 gccaccagtc cagggtgcac gaggtcccaa acggctcagg agcgtagctc cccacaccag
     3541 caaaaggcac tgtagcccag aacgtcgcat aggcccagat tagtgccagg cagataaagg
     3601 catggtgtct cttaagccag gtaccataag ccaagtgaca gatttttaag tatctatcca
     3661 ggctgacggc tgtcatggta ataaggctcc cacagccaaa gaagaagcca gcccatccgt
     3721 accagcggca gcccatccat ccaaagatcc agcggtgaga aaagaaggaa atgatactga
     3781 agggttttcc tacaactgag atgcccagat cacacaccgc taaattaaca gtcattatct
     3841 cagcgggtct taacttcttt ttccgctttg aggacataaa gataacatac ccgtttccca
     3901 gagttgaaag aatcccaatt actgtcaaat aaaaccccgc tatgatgtca gcttctctag
     3961 acagctttga tgcaaagggg tcttcctgct gcacgtagtg tgggaggtat tcttcctgag
     4021 aggaggagtt gcagtccgat gccatcccac tcatggtggc tagcataact tcgtataaag
     4081 tatcctatac gaagttattt gccttaaccc agaaattatc actgttattc tttagaatgg
     4141 tgcaaagaat aacttcgtat aatgtatgct atacgaagtt atgaattcga tatcaagctt
     4201 atcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     4261 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     4321 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     4381 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     4441 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     4501 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     4561 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg
     4621 ctgctcgcct atgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     4681 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     4741 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcatcga
     4801 taccgagcgc tgctcgagag atctacgggt ggcatccctg tgacccctcc ccagtgcctc
     4861 tcctggccct ggaagttgcc actccagtgc ccaccagcct tgtcctaata aaattaagtt
     4921 gcatcatttt gtctgactag gtgtccttct ataatattat ggggtggagg ggggtggtat
     4981 ggagcaaggg gcaagttggg aagacaacct gtagggcctg cggggtctat tgggaaccaa
     5041 gctggagtgc agtggcacaa tcttggctca ctgcaatctc cgcctcctgg gttcaagcga
     5101 ttctcctgcc tcagcctccc gagttgttgg gattccaggc atgcatgacc aggctcagct
     5161 aatttttgtt tttttggtag agacggggtt tcaccatatt ggccaggctg gtctccaact
     5221 cctaatctca ggtgatctac ccaccttggc ctcccaaatt gctgggatta caggcgtgaa
     5281 ccactgctcc cttccctgtc cttctgattt tgtaggtaac cacgtgcgga ccgagcggcc
     5341 gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga
     5401 ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga
     5461 gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg
     5521 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     5581 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg
     5641 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     5701 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     5761 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     5821 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     5881 acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc
     5941 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     6001 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     6061 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     6121 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     6181 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     6241 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     6301 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     6361 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     6421 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     6481 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     6541 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     6601 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     6661 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     6721 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     6781 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     6841 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     6901 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     6961 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     7021 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     7081 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     7141 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     7201 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     7261 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     7321 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     7381 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
//
