LOCUS       sequence-257202-        7013 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses JRGECO1a under control of eukaryotic Ef1a promoter after 
            Flp-dependent recombination..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7013)
  TITLE     AAV constructs for Flp-dependent expression of fluorescent proteins 
            and calcium indicators
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7013)
  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            "Unpublished"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-Ef1a-fDIO-JRGECO1a
FEATURES             Location/Qualifiers
     source          1..7013
                     /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    1440..1473
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(2036..2055)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             complement(2751..2807)
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKAGHAVRAIGRLSS"
     CDS             complement(join(2835..2849,2850..2858))
                     /codon_start=1
                     /product="Xpress(TM) epitope tag, including an enterokinase
                     recognition and cleavage site"
                     /label=Xpress(TM) tag
                     /translation="DLYDDDDK"
     CDS             complement(2862..2894)
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             complement(2898..2915)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     protein_bind    complement(2998..3031)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    3140..3728
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3193..3213)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3611..3622)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3760..4236
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3959..3978)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4276..4416
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4276..4405
                     /label=AAV2 ITR
     rep_origin      4491..4946
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4578..4597)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4788..4809
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4963..4982)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5082..5104
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5142..5160)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5228..5332
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5333..5401,5402..6193)
                     /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(5551..5570)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6364..6952
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6853..6872
                     /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 gactccggag
     1441 aagttcctat tctctagaaa gtataggaac ttcgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttagaagttc ctattcttca aaaggtatag gaacttcggc
     1561 gcggccgcct acttcgctgt catcatttgt acaaactctt cgtagtttac ctgaccatcc
     1621 ccatcgatgt ctgctaccct gatcatttca tcaacctcct catctgttaa cttctctcca
     1681 aggtctgtca tcacgtggcg aagctctgct gcgccgatgt agccattgcc gtccttatca
     1741 aacacgcgga acgcttcgcg aatttcctct tcactgtctg tgtcattcat ttttcttgcc
     1801 atcatcgtca ggaactcagg gaagtcgaat gtgccgtcac cgtcggcatc tacttcattg
     1861 atcatgtcct gcagctctgc ttctgtgggg ttctgcccca gcgatcggaa caccgtccca
     1921 agctccttgg ttgttatcgt cccatccccg tccttgtcaa atagggagaa agcctcttta
     1981 aattctgcga tctgctcttc agtcaggtcg tcacgcgtag cctcccagcc catggtcttc
     2041 ttctgcatta cggggccgtc gggggggaag ttggtgccgc gcagcttcac cttgtagatg
     2101 aatacgccgt cctgcaggga ggagtcctgg ttcacgtgaa taatgccgcc gtcctcgaag
     2161 ttcatcacgc gctcccacct gaagccctcg gggaaggaca gcttgaagta gtcggggatg
     2221 tcggctgggt gcttaatgta ggccttggag ccgtacatga actgagggga caggatgtcc
     2281 caggcgaagg gcagggggcc acccttggtc accttcagct tagcggtctg aaaggcctcg
     2341 taggggcggc cctcgccctc gccctcgatc tcgaactcgt ggccgttcac ggagccctcc
     2401 atgtgcacct tgaagcgcat gaactccttg atgatggcca tgttatcctc ctcgcccttg
     2461 ctcaccagac tcccgcctgt acctcccttg tacagctcgt ccatgccgcc ggtggagtgg
     2521 cggccctcgg cgcgttcgca ctgttccacg atggtgtagt cctcgttgtg ggacacgatg
     2581 tccaacttga tgtccacgat gtaggcgccg ggcagctgca cgggcttctt ggccttgtag
     2641 gtggtcttga cctcggcggc gtagtggccg ccgtccttca gcctcagccc cttcttgatc
     2701 tcgctcttca gggcgccgtc ctcggggtac atccgctcgg aaaccacggg tgagctcagc
     2761 cgacctatag ctctgactgc gtgacctgcc ttattccact tacgacgtga tgagtcgacc
     2821 attgttgcga gatccttatc gtcatcgtcg tacagatccc gacccatttg ctgtccacca
     2881 gtcatgctag ccataccatg atgatgatga tgatgagaac ctcctccagt cttgttaata
     2941 tcaagtccag ccaacttaag agcaagctcg ttctgcagca tggtggcgag atctagcgaa
     3001 gttcctatac tttctagaga ataggaactt cttgccttaa cccagaaatt atcactgtta
     3061 ttctttagaa tggtgcaaag agaagttcct ataccttttg aagaatagga acttcgaatt
     3121 cgatatcaag cttatcgata atcaacctct ggattacaaa atttgtgaaa gattgactgg
     3181 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     3241 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttgct
     3301 gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt
     3361 tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc tttccgggac
     3421 tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg
     3481 ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg ggaaatcatc
     3541 gtcctttcct tggctgctcg cctatgttgc cacctggatt ctgcgcggga cgtccttctg
     3601 ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc tgccggctct
     3661 gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc tttgggccgc
     3721 ctccccgcat cgataccgag cgctgctcga gagatctacg ggtggcatcc ctgtgacccc
     3781 tccccagtgc ctctcctggc cctggaagtt gccactccag tgcccaccag ccttgtccta
     3841 ataaaattaa gttgcatcat tttgtctgac taggtgtcct tctataatat tatggggtgg
     3901 aggggggtgg tatggagcaa ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc
     3961 tattgggaac caagctggag tgcagtggca caatcttggc tcactgcaat ctccgcctcc
     4021 tgggttcaag cgattctcct gcctcagcct cccgagttgt tgggattcca ggcatgcatg
     4081 accaggctca gctaattttt gtttttttgg tagagacggg gtttcaccat attggccagg
     4141 ctggtctcca actcctaatc tcaggtgatc tacccacctt ggcctcccaa attgctggga
     4201 ttacaggcgt gaaccactgc tcccttccct gtccttctga ttttgtaggt aaccacgtgc
     4261 ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc
     4321 gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg
     4381 cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc
     4441 ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg
     4501 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
     4561 cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
     4621 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
     4681 gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg
     4741 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
     4801 ccaaactgga acaacactca actctatctc gggctattct tttgatttat aagggatttt
     4861 gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt
     4921 taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc
     4981 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     5041 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     5101 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     5161 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     5221 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     5281 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     5341 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     5401 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     5461 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     5521 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     5581 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     5641 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     5701 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     5761 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     5821 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     5881 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     5941 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     6001 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     6061 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     6121 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     6181 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     6241 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     6301 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     6361 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     6421 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     6481 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     6541 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     6601 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     6661 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     6721 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     6781 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     6841 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     6901 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     6961 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgt
//
