LOCUS       sequence-310109-        6985 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV plasmid with Ple331 (PAX6 MiniPromoter) driving expression of 
            EmGFP. Contains WPRE..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6985)
  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 6985)
  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: pEMS2260
FEATURES             Location/Qualifiers
     source          1..6985
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     intron          2224..2356
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2339..2358
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      2377..2386
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2383..2385,2386..2388,2389..3102)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2428..2449)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2689..2708)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3036..3057
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3168..3188)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3586..3597)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3718..3839
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3729..3748)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3783..3802
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4035..4164
                     /label=AAV2 ITR
     primer_bind     complement(4183..4200)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4183..4199)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4192..4214)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4341..4796
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4428..4447)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4638..4659
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4822..4926
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4927..4995,4996..5787)
                     /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(5145..5164)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5958..6546
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6447..6466
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6834..6855
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6870..6875,6876..6893,6894..6900)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6908..6924
                     /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     6913..6935
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6932..6948
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6932..6948
                     /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 ggccggcctc ccttttgcta caggcatagt ttaagtcttg
      241 tgtcacaaat aaaaggatcc tccctagcat atggagtagt tgccatgtcc tcccagggat
      301 ggtctgtagc tatgttcacc tcaggctcat caagaagagg taaactgtca agtataattt
      361 cagtcatagg gtggtttggt tgagacagat tagtggtatg caatcatgat agatagttta
      421 tcccagctcc aaactgccgc ttgggaaatc ttgaactcaa atgaacagag actcagtcat
      481 tattagtgtc aaaaaggtat acagaggcaa aatttggttg gcagcataaa agatgaatca
      541 aagcaaaaaa gtgaccaatt gcatgttcaa aatcgaaatt gagatgcata ctacagcttg
      601 gaaatacttt aaataccaga aaagtcacac attattaatt gcgcattgat taatatttaa
      661 aacatgtttc tgtaatgtat tttcaaagag acaataaaat tacatgtatt ttcttatttc
      721 aaaaccttta ttctcaaggt gatgcatagt aaggagtacc cttttagata caggagtaag
      781 atagcaaaat gttttttaaa aatttgttta atttttagta tctttccttc agctgcatat
      841 ttccagaaat aagacttttt agaagaagaa tttagtttta tgggactatt ttaatattca
      901 aatgtcttac agtttgatga ctttctcagg aagaaatcag gttataataa aagcatgaat
      961 ttaaaatgtg gaagaaccta atctaaataa ctttccaaat attttctaaa cactgaaaaa
     1021 tatatattta taaattgtaa aatacattat tctattagca aacacaaact ttggtatctt
     1081 taaatcagac tcccccccca accaaatatc cacaaaaaga atgtcctcaa gtgtcaatgc
     1141 atgaagtcat gcgtgaagat aaatctaaca gctctttctg tgtcattcct aatgcacttg
     1201 tcactcagta ttatccatcc tcattaatga caatgggaaa gtttatcttg ggaacacgtt
     1261 ttaaatcatc tactctttac aatgggacca tgaaatctct tacaaaacat gaggcgggaa
     1321 ctactctgac aacaaaaccc cttcctggca gctttattct taattcctgt tcaaggagta
     1381 atctgtttat tataaaatca tacagaacaa tatattaaac agaatttgaa acaagaatag
     1441 ctagtgtgat ttccaagctt acaagtattt ttacaacatt tttattaggc tgaaaacact
     1501 acattgagtt ctttttgtgt gatttcatgc aatcttctat tctacatcgt aaacaccttt
     1561 gaaaagaagt tttgttttac ataatttaaa aataattttc aaatggtaag gacactggtg
     1621 tttatataat tattttaaaa tacaataact tacattagct taaataaaag cagttattaa
     1681 gtaaaaatgt gctcatgttt aaatttccta accaaaatgc caaaatattc caactcttgg
     1741 ggatcggagg aggggacagg gtgattaccc agagaggtag ctggccagcc taagggcaga
     1801 gatcttgggg ccctagtgcc cgaaggtgcg gaggagcgca ctcggcaaga ctagtttcct
     1861 ggggatcgac tctacgccat acaggacggc ggcccaggct ggaccgggcc gggctagagc
     1921 agtcacaggc cgggccaagg aaggccaaag caggggttgg agccggccgg accctgggtg
     1981 gggagaagca ggctcccgcc cggccggaaa actagtcggc gcagagctgt gcccaactct
     2041 agccgccatg acgtcacgcg ggccgggcag ccaatgagga cggcgctggc gtggatatta
     2101 aggaaagtta gcgcctgcct gagcaccctc ttttcttatc attgacattt aaactctggg
     2161 gcaggtcctc gcgtagaacg cggctgtcag gcgcgccgaa gttggtcgtg aggcactggg
     2221 caggtaagta tcaaggttac aagacaggtt taaggagacc aatagaaact gggcttgtcg
     2281 agacagagaa gactcttgcg tttctgatag gcacctattg gtcttactga catccacttt
     2341 gcctttctct ccacaggtgt ccactcccag gcggccgcca ccatggtgag caagggcgag
     2401 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
     2461 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
     2521 ttcatctgca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccttcacc
     2581 tacggcgtgc agtgcttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag
     2641 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     2701 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     2761 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     2821 aacagccaca aggtctatat caccgccgac aagcagaaga acggcatcaa ggtgaacttc
     2881 aagacccgcc acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac
     2941 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     3001 gccctgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     3061 gccgccggga tcactctcgg catggacgag ctgtacaagt aagcggccgc gatcgctcaa
     3121 cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt tgctcctttt
     3181 acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc ccgtatggct
     3241 ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga gttgtggccc
     3301 gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc cactggttgg
     3361 ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct ccctattgcc
     3421 acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg gctgttgggc
     3481 actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct gctcgcctgt
     3541 gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc cctcaatcca
     3601 gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg tcttcgcctt
     3661 cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcgatcgca gacatgataa
     3721 gatacattga tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt
     3781 gtgaaatttg tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta
     3841 acaacaacaa ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt
     3901 aaagcaagta aaacctctac aaatgtggta gtcgaaattc ccgataagga tcttcgtcga
     3961 cgaattcccg ataaggatct tcctagagca tggctacgta gataagtagc atggcgggtt
     4021 aatcattaac tacaaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg
     4081 ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc
     4141 ctcagtgagc gagcgagcgc gcagccttaa ttaacctaat tcactggccg tcgttttaca
     4201 acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc
     4261 tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg
     4321 cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     4381 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     4441 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     4501 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     4561 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     4621 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     4681 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     4741 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt
     4801 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     4861 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     4921 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     4981 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     5041 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     5101 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     5161 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     5221 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     5281 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     5341 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     5401 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     5461 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     5521 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     5581 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     5641 gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     5701 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     5761 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     5821 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     5881 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     5941 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     6001 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     6061 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     6121 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     6181 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     6241 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     6301 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     6361 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     6421 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     6481 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     6541 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     6601 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     6661 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     6721 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc
     6781 agctggcacg acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg
     6841 agttagctca ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg
     6901 tgtggaattg tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc
     6961 agatttaatt aaggccttaa ttagg
//
