LOCUS       sequence-216052-        6833 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ssAAV genome with Ple321 (TUBB3 MiniPromoter) driving an emerald GFP
            (EmGFP) reporter. Contains WPRE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6833)
  AUTHORS   Simpson E, Korecki AJ, Fornes O, McGill TJ, Cueva-Vargas JL, 
            Agostinone J, Farkas RA, Hickmott JW, Lam SL, Mathelier A, Renner 
            LM, Stoddard J, Zhou M, Di Polo A, Neuringer M, Wasserman WW
  TITLE     New MiniPromoter Ple345 (NEFL) Drives Strong and Specific Expression
            in Retinal Ganglion Cells of Mouse and Primate Retina.
  JOURNAL   Hum Gene Ther. 2018 Jul 31. doi: 10.1089/hum.2018.118.
  PUBMED    30062914
REFERENCE   2  (bases 1 to 6833)
  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: "Hum Gene 
            Ther. 2018 Jul 31. doi: 10.1089/hum.2018.118."
COMMENT     Sequence Label: pEMS2242
FEATURES             Location/Qualifiers
     source          1..6833
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     intron          2072..2204
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     2187..2206
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      2225..2234
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2231..2233,2234..2236,2237..2950)
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2276..2297)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2537..2556)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2884..2905
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3016..3036)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3434..3445)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3566..3687
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3577..3596)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3631..3650
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   3883..4012
                     /label=AAV2 ITR
     primer_bind     complement(4031..4048)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4031..4047)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4040..4062)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      4189..4644
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4276..4295)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4486..4507
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4670..4774
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4775..4843,4844..5635)
                     /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(4993..5012)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5806..6394
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6295..6314
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6682..6703
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6718..6723,6724..6741,6742..6748)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6756..6772
                     /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     6761..6783
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6780..6796
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6780..6796
                     /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 ggccggccag gggacgtcgt gtcccctggt cctgggaggc
      241 tgtccctggc ccaaggtcac acagcaggtg cgagagagct gggctggccc ttggttctca
      301 tcccctgcag gagccaacct ggggctcggg cggggcgggt tgggggcgct gcttgccaaa
      361 ggatctgccc tccgaacagt gttggggaac aggggatcgg ggggggaggc tggctttgtc
      421 aggaccaagg ccagggaccc agctcagggc tggtgcagcg ctgattctgg gaaggagggg
      481 gtgcaggaga cggtcctgga ggggaagggg ccagagaaag gcagcctccg gaggctgttg
      541 ccatggaaac caggcaggaa caaaaagcta attacaacag ggccagccac gtggctgaca
      601 tgtaaattgc aactttggct ccgggtggag gcgcctctga gggtgggagg cccagtagtg
      661 gggagactct aatcacccaa ggagacagct gtgcctgcca aggagagcct gggggagggg
      721 agacctgggg gacagtggcc tcaaatcaaa ccaggactcc cggagcagcc tctgacagct
      781 gccttggcct gatgggaggg gctgagggct cattaatgtg gtcactgggg ccaagcctcc
      841 cctgagcatc tgctcctggg agggtcaggg gtcgctgggg ggagggttcg cagctgctca
      901 ttttgtgaag tgaatgctga tttgggggcc gggtggggtt cgtctgtaca tcgtgcactg
      961 tcagaccctt cctgaaggat ttttgttact gaagtatcag aaggcccttg ttctaaggtg
     1021 gtggcactgt aatatctggc ccaggaggag gtcagtgccg tgccggcgag tcctttgctc
     1081 ccttccttct ctgaccagta aacgcatctc cagtcgctcc cccgaggccc ccctgcacgc
     1141 agtctccgac ctcagagagt gagtcagata gaagccgctg cccaccctcc cccaccctgg
     1201 ccgggttctt gcctggcttg tgcgtccttg tgacaattcc tgacacggat gcgcaccggg
     1261 cagtggcacg tccaggtgct gtgcgcgccc gggcccacaa agctcctttg gacgctcatg
     1321 gcagcccctt ggtggaggag gtcgggtgcc caccccctcc ccgcccactg cggaagccgg
     1381 cgacccacgg agctcgctct cggccgccgc cacccctctg tgttcgcgcc cttccgagct
     1441 ctgatccgac gctttgtttc ttctcagtgg gttcagggcc tgggccagcc tttacctacc
     1501 tcccccaccc aaaaccggca aaagctcaga gcaccttgtc tgccaaaaga cagggagctg
     1561 ggatggtgcg ggttggtctc taaaccggcg tggggaaaaa agaccctccg tacaaagccg
     1621 cagggtgggg ctgtcgcaag ggcggaaccg agagggtagc tgggggcggg gttcccaggg
     1681 ccaagagggg ccattgtcct ccctggagcc cggcgccccc acagccagct cctctgggag
     1741 acagcccctc ctttcgaatg cgcggggccc tcagaccgcg cccggcccag cgctggggga
     1801 tccttggctg cgggaggggc gccgcattgc gcgcggcggg cggggacgcg cggtgcggag
     1861 cctgcgggcc gggcggggct ctgcggcggc gcctcccgat tggccacccg cggtgacatc
     1921 agccgatgcg aagggcgggg ccgcggctat aagagcgcgc ggccgcggtc cccgaccctc
     1981 agcagccagc ccggcccgcc cgcgcccgtc cgcagccgcc cgccagacgc gcccagtggc
     2041 gcgccgaagt tggtcgtgag gcactgggca ggtaagtatc aaggttacaa gacaggttta
     2101 aggagaccaa tagaaactgg gcttgtcgag acagagaaga ctcttgcgtt tctgataggc
     2161 acctattggt cttactgaca tccactttgc ctttctctcc acaggtgtcc actcccaggc
     2221 ggccgccacc atggtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt
     2281 cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga
     2341 tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc
     2401 ctggcccacc ctcgtgacca ccttcaccta cggcgtgcag tgcttcgccc gctaccccga
     2461 ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg
     2521 caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg
     2581 cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat
     2641 cctggggcac aagctggagt acaactacaa cagccacaag gtctatatca ccgccgacaa
     2701 gcagaagaac ggcatcaagg tgaacttcaa gacccgccac aacatcgagg acggcagcgt
     2761 gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc
     2821 cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga
     2881 tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct
     2941 gtacaagtaa gcggccgcga tcgctcaacc tctggattac aaaatttgtg aaagattgac
     3001 tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt
     3061 gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt
     3121 gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt
     3181 gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg
     3241 gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg
     3301 ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc
     3361 atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt
     3421 ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc
     3481 tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc
     3541 cgcctccccg cgatcgcaga catgataaga tacattgatg agtttggaca aaccacaact
     3601 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
     3661 accattataa gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag
     3721 gttcaggggg agatgtggga ggttttttaa agcaagtaaa acctctacaa atgtggtagt
     3781 cgaaattccc gataaggatc ttcgtcgacg aattcccgat aaggatcttc ctagagcatg
     3841 gctacgtaga taagtagcat ggcgggttaa tcattaacta caaggaaccc ctagtgatgg
     3901 agttggccac tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg
     3961 cccgacgccc gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agccttaatt
     4021 aacctaattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
     4081 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     4141 gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatgggac gcgccctgta
     4201 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     4261 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     4321 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     4381 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     4441 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     4501 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     4561 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     4621 acaaaatatt aacgcttaca atttaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     4681 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     4741 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     4801 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     4861 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     4921 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     4981 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     5041 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     5101 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     5161 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     5221 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     5281 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     5341 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     5401 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     5461 tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc
     5521 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     5581 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     5641 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     5701 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     5761 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     5821 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     5881 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     5941 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     6001 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     6061 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     6121 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     6181 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     6241 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     6301 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     6361 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     6421 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     6481 tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac
     6541 cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct
     6601 ccccgcgcgt tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     6661 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
     6721 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
     6781 aggaaacagc tatgaccatg attacgccag atttaattaa ggccttaatt agg
//
