LOCUS       sequence-67998-e        7593 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An AAV vector that expresses engineered halorhodopsin 3.0 (fused to 
            iRFP) in a Cre-dependent manner (DIO, double floxed inverted open 
            reading frame)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7593)
  AUTHORS   Richie CT, Whitaker LR, Whitaker KW, Necarsulmer J, Baldwin HA, 
            Zhang Y, Fortuno L, Hinkle JJ, Koivula P, Henderson MJ, Sun W, Wang 
            K, Smith JC, Pickel J, Ji N, Hope BT, Harvey BK
  TITLE     Near-infrared fluorescent protein iRFP713 as a reporter protein for 
            optogenetic vectors, a transgenic Cre-reporter rat, and other 
            neuronal studies.
  JOURNAL   J Neurosci Methods. 2017 Jun 1;284:1-14. doi: 
            10.1016/j.jneumeth.2017.03.020. Epub 2017 Apr 2.
  PUBMED    28380331
REFERENCE   2  (bases 1 to 7593)
  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: "J Neurosci 
            Methods."; date: "2017-06-1"; volume: "284:1-14. doi"; pages: " 
            10.1016/j.jneumeth.2017.03.020. Epub 2017 Apr 2"
COMMENT     Sequence Label: pOTTC469 - pAAV EF1a DIO eNpHR3.0-iRFP
FEATURES             Location/Qualifiers
     source          1..7593
                     /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)."
     CDS             complement(1632..2579)
                     /codon_start=1
                     /product="phytochrome-based near-infrared fluorescent 
                     protein, originally termed iRFP (Filonov et al., 2011)"
                     /label=iRFP713
                     /note="name was changed to iRFP713 (Shcherbakova and 
                     Verkhusha, 2013)"
                     /translation="MAEGSVARQPDLLTCDDEPIHIPGAIQPHGLLLALAADMTIVAGS
                     DNLPELTGLAIGALIGRSAADVFDSETHNRLTIALAEPGAAVGAPITVGFTMRKDAGFI
                     GSWHRHDQLIFLELEPPQRDVAEPQAFFRRTNSAIRRLQAAETLESACAAAAQEVRKIT
                     GFDRVMIYRFASDFSGEVIAEDRCAEVESKLGLHYPASTVPAQARRLYTINPVRIIPDI
                     NYRPVPVTPDLNPVTGRPIDLSFAILRSVSPVHLEFMRNIGMHGTMSISILRGERLWGL
                     IVCHHRTPYYVDLDGRQACELVAQVLAWQIGVMEE"
     protein_bind    3578..3611
                     /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    3662..3695
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3720..4308
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3773..3793)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4191..4202)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4340..4816
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4539..4558)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4856..4996
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4856..4985
                     /label=AAV2 ITR
     rep_origin      5071..5526
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5158..5177)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5368..5389
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5543..5562)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5662..5684
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5722..5740)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5808..5912
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5913..5981,5982..6773)
                     /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(6131..6150)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6944..7532
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7433..7452
                     /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 gcgccataag cttgatatcg aattctcatt acacctcgtt ctcgtagcag
     1621 aacttgtaca gctcttccat cacgccgatc tgccaggcca gaacctgggc gactagctcg
     1681 caggcttggc ggccatcgag atcgacgtag tacggcgttc ggtgatggca aacgatcaat
     1741 ccccacagtc gctcgccgcg caaaatcgag atcgacatcg tgccgtgcat gcctatgttg
     1801 cgcatgaatt ccagatggac gggcgagacg ctgcgcagga tggcgaagct aagatcaatc
     1861 ggccgcccgg tgaccggatt gaggtctggg gtgaccggca ccggccgata attgatatcg
     1921 ggaatgatcc gtaccgggtt gatggtatag agccgacggg cctgcgccgg cacggttgag
     1981 gcaggatagt gcaggcctag ttttgactcg acctcggcgc accgatcctc tgcgatcact
     2041 tcgccgctga agtcggaggc gaagcgatag atcatcaccc gatcgaagcc ggtaatcttc
     2101 cgcacctctt gcgccgcggc ggcgcaggcg ctttccaagg tttcggcggc ctgcaggcgg
     2161 cggatggcgc tgttggtgcg gcggaagaac gcctgcggct cggcgacgtc ccgctgggga
     2221 ggctcgagct cgaggaagat gagctgatca tggcgatgcc aggagccgat gaagcctgcg
     2281 tcctttcgca tcgtgaagcc gacagtgatc ggtgctccga cggccgcccc gggctcggcc
     2341 aaggcgatcg tcagacggtt gtgcgtctcc gagtcgaaga catcggccgc agagcggccg
     2401 atcagggcgc cgatcgccag tccggtgagt tcgggaaggt tgtcgctgcc ggcaacgatc
     2461 gtcatgtcgg cggcgagggc gagcagcagt ccatgcggtt ggatggcacc ggggatatgg
     2521 atcggctcat cgtcgcaggt caagaggtca ggctgcctgg cgacggatcc ttcagccata
     2581 ctagtcacca cgttgatgtc gatctggtcc agggggatgt actcgccctc gctggtgatc
     2641 ctgctcttgg cggccgcatc atcagccggg gtcccagaag cagatggaac atccaaaatg
     2701 ctgccggaca cgacagattc gtttgacgtc agatagttca ggagcaagaa tgcgaaaatg
     2761 tactttgcca cgatatccag gaaagaatat ccccagctcg tcaccccaac gggcaacacc
     2821 gcgataccct cgactccaag agcccacaca attggatacc ccagccacat cactactgtc
     2881 aacagcttca gggtattgaa catgtcagcg gttcccgcgg ctttggcgtc ctgtgcccac
     2941 tccaccagca ggatatacaa gaccaccaga aagcatgcac aactgatagc gtaccagaac
     3001 catctcatga ggtgggagga ggtagtcagg gccgcggcaa ggccagtcac gcacatagcg
     3061 atatcgaaag tgatagctgt gaagagcttt gtagcattgc ttccggccag gagacccaga
     3121 gcgaggagaa tcatgggcgt ggaaagtgcc caggtgagat accgtcccca catggtgact
     3181 acaccatcta cctcttctcc tcccagcatc actgagctgc cttctgcaaa atggcccgct
     3241 ggcatttcaa gaacggaaat tgtcaggccg ctcgccaatc cagtgtagga ggcaatgctg
     3301 acgacaggca caaggatggt tgacacagca ataagttttg cccgtggatc atcgagtcct
     3361 cgggtcataa aaacgaacag cagtatactc agtcctgcaa gtgcgatgtt gatatagaga
     3421 ctgcttgcaa gcaaagggtc gttcagcacg aactcgaaca actccctttg ggtaacctcg
     3481 gcttgaaggg ccacggcact ctcggtcacg ggaggcaggg tctctgtcat ggtggcctac
     3541 cggtacccgg ggatccggta ccgctctaga gctagcgata acttcgtata aagtatccta
     3601 tacgaagtta tttgccttaa cccagaaatt atcactgtta ttctttagaa tggtgcaaag
     3661 aataacttcg tataatgtat gctatacgaa gttatgaatt cgatatcaag cttatcgata
     3721 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     3781 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     3841 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     3901 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     3961 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     4021 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     4081 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     4141 cctatgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     4201 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     4261 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgag
     4321 cgctgctcga gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc
     4381 cctggaagtt gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat
     4441 tttgtctgac taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa
     4501 ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag
     4561 tgcagtggca caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct
     4621 gcctcagcct cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt
     4681 gtttttttgg tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc
     4741 tcaggtgatc tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc
     4801 tcccttccct gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa
     4861 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     4921 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     4981 cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     5041 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg
     5101 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     5161 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5221 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5281 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     5341 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5401 accctatctc gggctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     5461 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     5521 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     5581 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     5641 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     5701 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     5761 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     5821 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     5881 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     5941 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     6001 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     6061 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     6121 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     6181 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     6241 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     6301 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     6361 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6421 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     6481 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     6541 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6601 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6661 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6721 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     6781 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     6841 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     6901 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     6961 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     7021 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     7081 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     7141 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     7201 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     7261 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     7321 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7381 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7441 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7501 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7561 tcctggcctt ttgctggcct tttgctcaca tgt
//
