LOCUS       sequence-309096-        7488 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid with Ple253 (PITX3 MiniPromoter) driving expression of 
            EmGFP. Contains WPRE..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7488)
  AUTHORS   Korecki AJ, Cueva-Vargas JL, Fornes O, Agostinone J, Farkas RA, 
            Hickmott JW, Lam SL, Mathelier A, Zhou M, Wasserman WW, Di Polo A, 
            Simpson EM
  TITLE     Human MiniPromoters for ocular-rAAV expression in ON bipolar, cone, 
            corneal, endothelial, Muller glial, and PAX6 cells.
  JOURNAL   Gene Ther. 2021 Feb 2. pii: 10.1038/s41434-021-00227-z. doi: 
            10.1038/s41434-021-00227-z.
  PUBMED    33531684
REFERENCE   2  (bases 1 to 7488)
  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: "Gene Ther. 
            2021 Feb 2. pii: 10.1038/s41434-021-00227-z. doi: 
            10.1038/s41434-021-00227-z."
COMMENT     Sequence Label: pEMS2230
FEATURES             Location/Qualifiers
     source          1..7488
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     intron          2727..2859
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2842..2861
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      2880..2889
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2886..2888,2889..2891,2892..3605)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2931..2952)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3192..3211)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3539..3560
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3671..3691)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4089..4100)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4221..4342
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4232..4251)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4286..4305
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4538..4667
                     /label=AAV2 ITR
     primer_bind     complement(4686..4703)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4686..4702)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4695..4717)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4844..5299
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4931..4950)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5141..5162
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5325..5429
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5430..5498,5499..6290)
                     /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(5648..5667)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6461..7049
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6950..6969
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    7337..7358
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7373..7378,7379..7396,7397..7403)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7411..7427
                     /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     7416..7438
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7435..7451
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7435..7451
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcg gaattcccta ggccggccac ctggccacga tgctctagtg taaagcaccc
      241 atcagatatc agacgtgaag aacccctcag taacttccta cacctgcaca ttgggcttct
      301 ggtcggtggg tccagctgag gatctagcct ctatcaccca aagagagtca gggaaagagg
      361 atataccatc atagcacctc tttccacatt acctcaaagc cattttcctc ttcatttgaa
      421 aactgagagt tggtgtcctg ggcctgtctc agattgagag gtcctggaaa gccgattgga
      481 aaatcctcta agtgggccaa aaatttctgc tcgccaatat ctacttctac atacctccag
      541 aaattcatca ttctaaaagc atgtgattgt gcaatagaga tggggaggag agagatgaca
      601 attccctagt ccctcctaac cactaatcat catcttacca ggcagaacat cttcctcata
      661 gatttcagtg tgacatttat tgagattcta cattatgcct gggaataaca gacatggtcc
      721 ttaccctcct aaattttatt atcatagcag acaacacata agctgatccc ttcaatatgg
      781 tgggtggtgt aatagaaggt atacactgag ggtcatgaga gcaaggggct aaactgagag
      841 ttcagaaaag gcttcccaga gaaaataaga cttgagtgga acattggagg agtaggaatt
      901 gacaatataa agatgggtta ctttcacagg aagggccaga ggtgtgcagg acactactag
      961 gaaattggag atgaagacaa ggtcaggtcc tgaaggtctt gtatgccagg ctagggagct
     1021 tgacctttat cctaagggca atgggagtca ctgaacagtc aagacatgac tgctcagatt
     1081 tgccttaggg gcatattgtg tgactgggct taggctcttt tttttttttt tgagacagag
     1141 tcttgctctg tcgcccagtc tggagtgcag tggcgcgatc tcagctcact gcaagctctg
     1201 cctcctgggt tcacgccatt cttcttcctc agcctcccca gtagctgaga ctacaggcac
     1261 ccgccaccaa gcccggctaa ttttttgtat tttttagtag agatggggtt tcaccgtgtt
     1321 agccagggtg gtctcaatct cctgaccttg tgatccgccc gcctcggcct cccaaagtgc
     1381 tgggattaca ggcatgagcc acagtgtctg gcctgggctt aggctcttcg gagcacccaa
     1441 gcaacatagc ctaactggga gaagagaagg agcctgtttc ctggaaattg aggcctaagt
     1501 cagagagggc cgaagtcaca gtctgccagg atgttccttg gtcatctgtc ctgtcccttg
     1561 caaattccta accagcaggc accaaggcca caccttgccc acaggactta acctggtacc
     1621 accagcgaga aaccagccct gctgaaggag ggagagcaca gcctaagcaa gacaagggta
     1681 cagactgaaa ggcaatgcct ttggagtgcc cagcccaagc cctgagatcc agcaggaacc
     1741 tcctcttaat tgttttgcgc ggggagggag atgatagtta ctcgtccaag ttcactcagt
     1801 gatggtgcag ttcgtggcag cgttggaata ctactttggt ttgtatcatt ccaaaccctg
     1861 ctgtctccac gaagtcacct tccttggaga agaaggggga aatgaagaag aggaaagata
     1921 tgaggaaaga ggagcagaga gaaagagaaa tgagtctttt aaagtggcag aggaaggaga
     1981 atggtaaaca gcaggagcga agcggctgag gagaaagaag aggaaagaaa ggcgagacgt
     2041 gggaggattg gaacagagac aaaagggagg agagacggac agcgacaagt ggagaaaatc
     2101 ggcgaaactt gagtggcaga gaagtctgag cgctgagacc cggcggcccc gtgcgccttc
     2161 ccacctggcg ccgatccact ttcctcgggg tagcggccca acccacttcg ctgccagccg
     2221 atccctttta cccgtggcta ccgggaccac tctactctcg cccacttggc tctgcctaag
     2281 cgtcctagcc ggagcgcggt ctctgccacg tggggagggg cgcggccgag ttgctgaaga
     2341 gcgcttctga ttggccagag ggcggggttc ttggcgtctc gccggccaga cccctccctc
     2401 aaaggcgggg cctggagatc cacagctgga aagggcggag ccccagcagg acagctggaa
     2461 aggggcgggg cctgacgcgc gcggctcgcc gcggcgggct gggggcgccc tggtctgcca
     2521 taaagtgaat gggcgccggc tgggggtggc agtacgcggt gaggctcact ccctccgaga
     2581 gtccaggagc gcccgagcgg agaggcggcc cgggagcagg ggggcggccc ccactccggc
     2641 cgggtgcccg gcccctggcc cctgcctgcc ctctagatcg ccgccgcagc cgggcgcgcc
     2701 gaagttggtc gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag
     2761 accaatagaa actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta
     2821 ttggtcttac tgacatccac tttgcctttc tctccacagg tgtccactcc caggcggccg
     2881 ccaccatggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2941 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     3001 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     3061 ccaccctcgt gaccaccttc acctacggcg tgcagtgctt cgcccgctac cccgaccaca
     3121 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     3181 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     3241 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     3301 ggcacaagct ggagtacaac tacaacagcc acaaggtcta tatcaccgcc gacaagcaga
     3361 agaacggcat caaggtgaac ttcaagaccc gccacaacat cgaggacggc agcgtgcagc
     3421 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3481 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     3541 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3601 agtaagcggc cgcgatcgct caacctctgg attacaaaat ttgtgaaaga ttgactggta
     3661 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     3721 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     3781 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     3841 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     3901 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     3961 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt
     4021 cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     4081 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     4141 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct
     4201 ccccgcgatc gcagacatga taagatacat tgatgagttt ggacaaacca caactagaat
     4261 gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat ttgtaaccat
     4321 tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt ttcaggttca
     4381 gggggagatg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg gtagtcgaaa
     4441 ttcccgataa ggatcttcgt cgacgaattc ccgataagga tcttcctaga gcatggctac
     4501 gtagataagt agcatggcgg gttaatcatt aactacaagg aacccctagt gatggagttg
     4561 gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga
     4621 cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagcct taattaacct
     4681 aattcactgg ccgtcgtttt acaacgtcgt gactgggaaa accctggcgt tacccaactt
     4741 aatcgccttg cagcacatcc ccctttcgcc agctggcgta atagcgaaga ggcccgcacc
     4801 gatcgccctt cccaacagtt gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc
     4861 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     4921 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     4981 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     5041 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     5101 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     5161 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     5221 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     5281 atattaacgc ttacaattta ggtggcactt ttcggggaaa tgtgcgcgga acccctattt
     5341 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     5401 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     5461 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5521 taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg gatctcaaca
     5581 gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg agcactttta
     5641 aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag caactcggtc
     5701 gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca gaaaagcatc
     5761 ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg agtgataaca
     5821 ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc gcttttttgc
     5881 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     5941 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac
     6001 tattaactgg cgaactactt actctagctt cccggcaaca attaatagac tggatggagg
     6061 cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg tttattgctg
     6121 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     6181 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     6241 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacc
     6301 aagtttactc atatatactt tagattgatt taaaacttca tttttaattt aaaaggatct
     6361 aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc
     6421 actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc
     6481 gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg
     6541 atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa
     6601 atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc
     6661 ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt
     6721 gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa
     6781 cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc
     6841 tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc
     6901 cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct
     6961 ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat
     7021 gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc
     7081 tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg
     7141 ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc
     7201 gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg
     7261 cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca
     7321 gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact
     7381 ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa
     7441 acagctatga ccatgattac gccagattta attaaggcct taattagg
//
