LOCUS Exported 6961 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION ssAAV genome with Ple320 (POU4F1 MiniPromoter) driving an emerald GFP (EmGFP) reporter. Contains WPRE. ACCESSION . VERSION . KEYWORDS pEMS2241 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6961) 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 6961) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6961 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR CDS 1565..1576 /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" intron 2211..2343 /label=chimeric intron /note="chimera between introns from human beta-globin and immunoglobulin heavy chain genes" primer_bind 2326..2345 /label=pMT2-F /note="Synthetic intron, forward primer" regulatory 2364..2373 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2370..3089 /codon_start=1 /product="Emerald GFP" /label=EmGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2415..2436) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2676..2695) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3023..3044 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3155..3175) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3573..3584) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3705..3826 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3716..3735) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3770..3789 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 4022..4151 /label=AAV2 ITR primer_bind complement(4170..4187) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(4170..4186) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4179..4201) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4328..4783 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4415..4434) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4625..4646 /label=F1ori-F /note="F1 origin, forward primer" promoter 4809..4913 /gene="bla" /label=AmpR promoter CDS 4914..5774 /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(5132..5151) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5945..6533 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6434..6453 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6687..6704 /label=L4440 /note="L4440 vector, forward primer" protein_bind 6821..6842 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6857..6887 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6895..6911 /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 6900..6922 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 6919..6935 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 6919..6935 /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 ggccggccga aatacataca ctagatttct cccccttttg 241 tgcctttgga tggtatttgt agagagaatc aaactgagat taactgttta tgggatgtac 301 tttattagat tagaagggtt tcccagagcc tcataagaag ttctttcatt cccatttgtg 361 ggagtcctac aacacagctt tggcctatca cgatgtaacc tgatcctttt aataatactt 421 ggcagagaaa accttttaag caatgatctg agtgcacagt gaacatttat tgcctaattc 481 tcggaacccc agtcagctag gaagcatctg tgaagatgca gatttcttta cagaacccac 541 ctggagtgca agttagggcg atcttcagtt ttatatgact gagacagtct agaatgcctt 601 gagttgcagc tccacttcag aaacaaggca gattcttagg cctgctatgt ctttcctgta 661 cccttcaggg tccctaaaga ccagcgcttc cttgagaggg gaggggcagg ttccaggccc 721 tcaccctctg tgggggcctt ttctttcccg tcaagagaga gcgcagggaa cggccaccag 781 gggtgtcagg aaggactagg ttaagcccgg actcccggtg tgatgactac cgtaggactc 841 actctttccc tctagcgacg tcctgctcat cgtccactcc ccgttgagat aaagttgcat 901 ttagcgctcc caggggactg cgtaatggat ggaaccagct ccttctctca ccagcaagtg 961 aacatccttc tcacttccct ctggagctag aaggaaggga gggggtgggg gggaacatga 1021 aaagaaagaa agaaaaagcc atggtgacgc ctgtgggctc ttcttggaat ctgagtaggt 1081 gagtgggtat gctaaacaca catgcatttg tgcagaggaa agataaaatc cacgggaacg 1141 caaatttagg tttatagaaa tgagatctgc tctttggtgg aaatacagac ttccttggtg 1201 tgggagagaa cagaaagatt atctgatttc tgctttccat aagcagcccc ttgctgctct 1261 ccagctctct taccgcagat gtgataacat ctgggctgtt aggttattat caccgcgagc 1321 aagtgctcct tccctgggga ctttggcacc tgaactgtgg ctcccctggg aggctcttga 1381 tctgctacca cataagggcc atataacagg ttggctaatt gatttattct ttttaggtct 1441 taatttcaca ataatttttt aaaaatttaa aaactggacc agagcttgtc cccgttcttt 1501 caaattggct tgatttaata tttttgaatc aagctttcct gtaagttagt ttgatctgtc 1561 acacatagag gggcgtgcgc gcatctcgtc ttttttattc taaaaggtgc ccacccttag 1621 acaatagctg gaccttctga tctttccaga ccacggtggg aaaggtgagg cgggcagtgc 1681 gcatgcccgg gtagctacag gtgcatcacc tgcgctgtcc ttgctggtgg tcactataag 1741 agcggctccc acgcgctggt cgcggccgga cgagcgcgcg cgcgcgcaga cgcacggggc 1801 cgccgagaag gagagcgcgc gtcagacaga cggagacagc tgatgcctgt cagcgcggtg 1861 gggccgcgag ctctcgcgag agctccccgg gacggccgcg agaagtgggg ctcaggtgtt 1921 agaggagtgc atcccgtcgg cgcgcggggc tagagctgtc ggagaagcgg gaccgcgagg 1981 ccggcgcgcg gcgctctgcg cggtcagagg gagcgcctgg cagcagcagg agcagcagca 2041 gcagcccgcg gcggggccgc cgccagccgc cgcgaccgcc gcggctgcag cctccgaagg 2101 gaggccgggt gagccggcgt acgcactttc ccgcggactt tcggagtgtt tgtggatata 2161 catgccaagc cgccacggcg cgccgaagtt ggtcgtgagg cactgggcag gtaagtatca 2221 aggttacaag acaggtttaa ggagaccaat agaaactggg cttgtcgaga cagagaagac 2281 tcttgcgttt ctgataggca cctattggtc ttactgacat ccactttgcc tttctctcca 2341 caggtgtcca ctcccaggcg gccgccacca tggtgagcaa gggcgaggag ctgttcaccg 2401 gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt 2461 ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca 2521 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cttcacctac ggcgtgcagt 2581 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg 2641 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg 2701 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact 2761 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaagg 2821 tctatatcac cgccgacaag cagaagaacg gcatcaaggt gaacttcaag acccgccaca 2881 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg 2941 acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag 3001 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca 3061 ctctcggcat ggacgagctg tacaagtaag cggccgcgat cgctcaacct ctggattaca 3121 aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat 3181 acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct 3241 ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac 3301 gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca 3361 cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca 3421 tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg 3481 tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga 3541 ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt 3601 cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga 3661 gtcggatctc cctttgggcc gcctccccgc gatcgcagac atgataagat acattgatga 3721 gtttggacaa accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga 3781 tgctattgct ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg 3841 cattcatttt atgtttcagg ttcaggggga gatgtgggag gttttttaaa gcaagtaaaa 3901 cctctacaaa tgtggtagtc gaaattcccg ataaggatct tcgtcgacga attcccgata 3961 aggatcttcc tagagcatgg ctacgtagat aagtagcatg gcgggttaat cattaactac 4021 aaggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag 4081 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag 4141 cgagcgcgca gccttaatta acctaattca ctggccgtcg ttttacaacg tcgtgactgg 4201 gaaaaccctg gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg 4261 cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc 4321 gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc 4381 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt 4441 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc 4501 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt 4561 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt 4621 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt 4681 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa 4741 aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg 4801 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc 4861 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta 4921 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg 4981 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg 5041 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac 5101 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg 5161 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt 5221 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg 5281 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac 5341 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt 5401 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag 5461 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc 5521 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc 5581 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta 5641 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg 5701 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga 5761 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac 5821 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa 5881 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat 5941 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc 6001 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg 6061 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc 6121 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg 6181 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg 6241 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa 6301 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg 6361 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga 6421 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct 6481 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca 6541 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc 6601 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg 6661 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc 6721 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca 6781 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc 6841 attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga 6901 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaga tttaattaag 6961 g //