LOCUS       sequence-446762-        6706 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid with Ple388 (PITX3 MiniPromoter) driving expression of 
            EmGFP. Contains WPRE..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6706)
  AUTHORS   Fornes O, Av-Shalom TV, Korecki AJ, Farkas RA, Arenillas DJ, 
            Mathelier A, Simpson EM, Wasserman WW
  TITLE     OnTarget: in silico design of MiniPromoters for targeted delivery of
            expression.
  JOURNAL   Nucleic Acids Res. 2023 Jul 5;51(W1):W379-W386. doi: 
            10.1093/nar/gkad375.
  PUBMED    37166953
REFERENCE   2  (bases 1 to 6706)
  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: "Nucleic 
            Acids Res."; date: "2023-07-5"; volume: "51(W1):W379-W386. doi"; 
            pages: " 10.1093/nar/gkad375"
COMMENT     Sequence Label: pEMS2386
FEATURES             Location/Qualifiers
     source          1..6706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   12..141
                     /label=AAV2 ITR
     intron          1956..2088
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2071..2090
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      2109..2118
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2115..2117,2118..2120,2121..2834)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2160..2181)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2421..2440)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2768..2789
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2900..2920)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3318..3329)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3450..3571
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3461..3480)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3515..3534
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3767..3896
                     /label=AAV2 ITR
     primer_bind     complement(3915..3932)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3915..3931)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3924..3946)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4073..4528
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4160..4179)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4370..4391
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4554..4658
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4659..4727,4728..5519)
                     /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(4877..4896)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5690..6278
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6179..6198
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6566..6587
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6602..6607,6608..6625,6626..6632)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6640..6656
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     6645..6667
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6664..6680
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6664..6680
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ccttaattag gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc ttgtagttaa tgattaaccc gccatgctac ttatctacgt
      181 agccatgctc taggaagatc ggaattccct aggccggcct aagcgcgccc ccttccgggg
      241 gtgcaggaca taacagcttc atccccagtg cgggtgggac cccggctggg cagccccggt
      301 gggagccagc gagtggctta ggaggtcccc gagccggaga atatgcgctg gcttgggcgc
      361 tctgtgacct gcccccaccc tggcccccag tcagtcccaa ctccctagct tcctgacctc
      421 cagcagtttg ggggagctta atcttcagcc agcctgtctc cagctcagac aacagcttgg
      481 cgcccccagc ctttgcctga ccagcccggg gttaaagttg aggcagcaaa tttatcccca
      541 gagatatatg tctagcacaa agctgagatt cacaagattc tccctgagtt ttccttacct
      601 ctaaatgctg gaataactga tgctcactga attccgatat ttgtatgaat aaaaatgaag
      661 ggatttagca aatcttgttc ctgcacacag ttgtcaaaaa aaatctgacc tttacaacag
      721 gaaaaagagg ggggatttaa agaatgtgag ttccacaaac gggcaagtaa ttgctcaata
      781 agcgtggaac aaggaaaggt cagttgtgat ctgtgcacat gtggcagccc ttcacaactc
      841 gacctacagt ggtaacagtg gtaacagaat gctgatgtta cagaagagac atcggattct
      901 ttggggatag agaaaataca taatatatat tactataaat gcaaaactgt actaaaagct
      961 aactcaaatg tcaaagatta cagtttgaat aaatttaaat ataaaagcta gccggagcgc
     1021 ggtctctgcc acgtggggag gggcgcggcc gagttgctga agagcgcttc tgattggcca
     1081 gagggcgggg ttcttggcgt ctcgccggcc agacccctcc ctcaaaggcg gggcctggag
     1141 atccacagct ggaaagggcg gagccccagc agggcagctg gaaaggggcg gggcctgacg
     1201 cgcgcggctc gccgcggcgg gctgggggcg ccctggtctg ccataaagtg aatgggcgcc
     1261 ggctgggggt ggcagtacgc ggtgaggctc actccctccg agagtccagg agcgcccgag
     1321 cggagaggcg gcccgggagc aggggggcgg cccccactcc ggccgggtgc ccggcccctg
     1381 gcccctgcct gccctctaga tcgccgccgc agccgccgct actgggagtc tgcctgttgc
     1441 aggacgcact gtgagtgttt gaccattcta aggacagggg ctcatggggc cgaggagcta
     1501 ttgctggacc ttggaatccc taaaacgaga cctggggcag ctgaactccg gtgccttggg
     1561 aacaggtcga gaatgggagg gcggactgca gacctggggc caggacttgg gaaaagggaa
     1621 actcagggta agaaccaggg gatagggaag ggggagactc agaggatccg gaacgaagcc
     1681 ggagacttga aaccaggata ggggcttcgc gaggcaggga gtagggagtc tggaaaccat
     1741 taccagaaag cgccctggaa tcccagggca cggggcaggc ggccagggag cgaaaggggt
     1801 tacagaggcc ctggccagcg ggggagctgg cggggcggcc cccagtgtgg gggcagttga
     1861 cggcgcccga tgcccctcgg cctaggctca gactcctggg gctgggagcc ggggtgcgcc
     1921 tggcgcgccg aagttggtcg tgaggcactg ggcaggtaag tatcaaggtt acaagacagg
     1981 tttaaggaga ccaatagaaa ctgggcttgt cgagacagag aagactcttg cgtttctgat
     2041 aggcacctat tggtcttact gacatccact ttgcctttct ctccacaggt gtccactccc
     2101 aggcggccgc caccatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
     2161 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
     2221 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
     2281 tgccctggcc caccctcgtg accaccttca cctacggcgt gcagtgcttc gcccgctacc
     2341 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     2401 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
     2461 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     2521 acatcctggg gcacaagctg gagtacaact acaacagcca caaggtctat atcaccgccg
     2581 acaagcagaa gaacggcatc aaggtgaact tcaagacccg ccacaacatc gaggacggca
     2641 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     2701 tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc aacgagaagc
     2761 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     2821 agctgtacaa gtaagcggcc gcgatcgctc aacctctgga ttacaaaatt tgtgaaagat
     2881 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     2941 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     3001 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     3061 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     3121 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     3181 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     3241 aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
     3301 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     3361 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     3421 gggccgcctc cccgcgatcg cagacatgat aagatacatt gatgagtttg gacaaaccac
     3481 aactagaatg cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt
     3541 tgtaaccatt ataagctgca ataaacaagt taacaacaac aattgcattc attttatgtt
     3601 tcaggttcag ggggagatgt gggaggtttt ttaaagcaag taaaacctct acaaatgtgg
     3661 tagtcgaaat tcccgataag gatcttcgtc gacgaattcc cgataaggat cttcctagag
     3721 catggctacg tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg
     3781 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
     3841 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagcctt
     3901 aattaaccta attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     3961 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     4021 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc
     4081 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     4141 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4201 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4261 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     4321 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     4381 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     4441 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     4501 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
     4561 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     4621 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     4681 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     4741 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     4801 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4861 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     4921 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4981 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     5041 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     5101 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     5161 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     5221 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     5281 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     5341 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     5401 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     5461 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     5521 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     5581 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     5641 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     5701 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     5761 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     5821 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5881 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     5941 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     6001 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     6061 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     6121 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     6181 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     6241 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     6301 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     6361 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     6421 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc
     6481 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     6541 aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg
     6601 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     6661 acacaggaaa cagctatgac catgattacg ccagatttaa ttaagg
//
