LOCUS       Exported                5794 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  ssAAV genome with Ple344 (TUBB3 MiniPromoter) driving an emerald GFP
            (EmGFP) reporter. Contains WPRE.
ACCESSION   .
VERSION     .
KEYWORDS    pEMS2274
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5794)
  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 5794)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5794
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..130
                     /label=AAV2 ITR
     intron          1044..1176
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     1159..1178
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     regulatory      1197..1206
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1203..1922
                     /codon_start=1
                     /product="Emerald GFP"
                     /label=EmGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK
                     VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1248..1269)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1509..1528)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1856..1877
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(1988..2008)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2406..2417)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2538..2659
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2549..2568)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2603..2622
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2855..2984
                     /label=AAV2 ITR
     primer_bind     complement(3003..3020)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3003..3019)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3012..3034)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3161..3616
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3248..3267)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3458..3479
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3642..3746
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3747..4607
                     /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(3965..3984)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4778..5366
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5267..5286
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5520..5537
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5654..5675
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5690..5720
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5728..5744
                     /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     5733..5755
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5752..5768
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5752..5768
                     /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 gcagctgctc attttgtgaa gtgaatgctg
      241 atttgggggc cgggtggggt tcgtctgtac atcgtgcact gtcagaccct tcctgaagga
      301 tttttgttac tgaagtatca gaaggccctt gttctaaggt ggtggcactg taatatctgg
      361 cccaggagga ggtcagtgcc gtgccggcga gtcctttgct cccttccttc tctgaccagt
      421 aaacgcatct ccagtcgctc ccccgaggcc cccctgcacg cagtctccga cctcagagag
      481 tgagtcagat agaagccgct gcccaccctc ccccaccctg gccgggttct tgcctggctt
      541 gtgcgtcctt gtgacaattc ctgacacgga tgcgcaccgg gcagtggcac gtccaggtgc
      601 tgtgcgcgcc cgggcccaca aagctccttt ggacgctcat ggcagcccct tggtggagga
      661 ggtcgggtgc ccaccccctc cccgcccact gcggaagccg ctcctctggg agacagcccc
      721 tcctttcgaa tgcgcggggc cctcagaccg cgcccggccc agcgctgggg gatccttggc
      781 tgcgggaggg gcgccgcatt gcgcgcggcg ggcggggacg cgcggtgcgg agcctgcggg
      841 ccgggcgggg ctctgcggcg gcgcctcccg attggccacc cgcggtgaca tcagccgatg
      901 cgaagggcgg ggccgcggct ataagagcgc gcggccgcgg tccccgaccc tcagcagcca
      961 gcccggcccg cccgcgcccg tccgcagccg cccgccagac gcgcccagtg gcgcgccgaa
     1021 gttggtcgtg aggcactggg caggtaagta tcaaggttac aagacaggtt taaggagacc
     1081 aatagaaact gggcttgtcg agacagagaa gactcttgcg tttctgatag gcacctattg
     1141 gtcttactga catccacttt gcctttctct ccacaggtgt ccactcccag gcggccgcca
     1201 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     1261 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     1321 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     1381 ccctcgtgac caccttcacc tacggcgtgc agtgcttcgc ccgctacccc gaccacatga
     1441 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     1501 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     1561 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     1621 acaagctgga gtacaactac aacagccaca aggtctatat caccgccgac aagcagaaga
     1681 acggcatcaa ggtgaacttc aagacccgcc acaacatcga ggacggcagc gtgcagctcg
     1741 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     1801 actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     1861 tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagt
     1921 aagcggccgc gatcgctcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     1981 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     2041 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc
     2101 tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg
     2161 acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg
     2221 ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga
     2281 caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct
     2341 ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg
     2401 tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc
     2461 ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc
     2521 cgcgatcgca gacatgataa gatacattga tgagtttgga caaaccacaa ctagaatgca
     2581 gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg taaccattat
     2641 aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc aggttcaggg
     2701 ggagatgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta gtcgaaattc
     2761 ccgataagga tcttcgtcga cgaattcccg ataaggatct tcctagagca tggctacgta
     2821 gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat ggagttggcc
     2881 actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc
     2941 ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagccttaa ttaacctaat
     3001 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     3061 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     3121 cgcccttccc aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca
     3181 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta
     3241 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     3301 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     3361 cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     3421 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     3481 acaacactca accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg
     3541 gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     3601 ttaacgctta caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     3661 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     3721 ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc
     3781 ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa
     3841 aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg
     3901 gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag
     3961 ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc
     4021 gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta
     4081 cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg
     4141 cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca
     4201 acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac
     4261 caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat
     4321 taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg
     4381 ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata
     4441 aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta
     4501 agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa
     4561 atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag
     4621 tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg
     4681 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     4741 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     4801 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     4861 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     4921 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     4981 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     5041 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     5101 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     5161 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     5221 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     5281 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     5341 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     5401 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     5461 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     5521 gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg caaaccgcct ctccccgcgc
     5581 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     5641 agcgcaacgc aattaatgtg agttagctca ctcattaggc accccaggct ttacacttta
     5701 tgcttccggc tcgtatgttg tgtggaattg tgagcggata acaatttcac acaggaaaca
     5761 gctatgacca tgattacgcc agatttaatt aagg
//
