LOCUS Exported 6996 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION ssAAV genome with Ple342 (TUBB3 MiniPromoter) driving an emerald GFP (EmGFP) reporter. Contains WPRE. ACCESSION . VERSION . KEYWORDS pEMS2272 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 6996) 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 6996) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6996 /organism="synthetic DNA construct" /mol_type="other DNA" repeat_region 1..130 /label=AAV2 ITR intron 2235..2367 /label=chimeric intron /note="chimera between introns from human beta-globin and immunoglobulin heavy chain genes" primer_bind 2350..2369 /label=pMT2-F /note="Synthetic intron, forward primer" regulatory 2388..2397 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2394..3113 /codon_start=1 /product="Emerald GFP" /label=EmGFP /note="mammalian codon-optimized" /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL KFICTTGKLPVPWPTLVTTFTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHKVYITADKQKNGIK VNFKTRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL EFVTAAGITLGMDELYK" primer_bind complement(2439..2460) /label=EGFP-N /note="EGFP, reverse primer" primer_bind complement(2700..2719) /label=EXFP-R /note="For distinguishing EGFP variants, reverse primer" primer_bind 3047..3068 /label=EGFP-C /note="EGFP, forward primer" primer_bind complement(3179..3199) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(3597..3608) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" polyA_signal 3729..3850 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(3740..3759) /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind 3794..3813 /label=SV40pA-R /note="SV40 polyA, reverse primer" repeat_region 4046..4175 /label=AAV2 ITR primer_bind complement(4194..4211) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(4194..4210) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(4203..4225) /label=M13/pUC Forward /note="In lacZ gene" rep_origin 4352..4807 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(4439..4458) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 4649..4670 /label=F1ori-F /note="F1 origin, forward primer" promoter 4833..4937 /gene="bla" /label=AmpR promoter CDS 4938..5798 /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(5156..5175) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 5969..6557 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 6458..6477 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 6711..6728 /label=L4440 /note="L4440 vector, forward primer" protein_bind 6845..6866 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 6881..6911 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 6919..6935 /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 6924..6946 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 6943..6959 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 6943..6959 /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 ggccggcccc tgcgctggga gccgagcccc cggcagctcg 241 cggggatgca gtaagccggg cgcagcccgc cgggaggatg agaaaacccg aagcccggaa 301 agccgagcgg ccgcctgcag tcaccccggg gggggcaggg gcgggccggg ctatgcagaa 361 acaccggggc ccgcgggaca caggacgctg ccccactgcc gcagagctga atctggaccc 421 ccgggctttg ctttcctggg gcctgtgctc ctccctcccc ggggcggggt tcgcctgcag 481 cggcggaggg gggagccttt gtcctccgcg cgggctgcgg caccgcctcc tcgcggctgc 541 ggggcggggc tgggggcggg gcctggccgt ctcagccaat gggaggcgga gtccctggct 601 cgccccgccc tctcgcctga aagacctgcg gagccgcggg ccgggcggga accgcattcg 661 ggtcctggag gggcttggag gccccagtct cgcagcccct tcatcgggga cccccgctcc 721 cacctgcctt ggccactgcc ctcaaagccc ggccccgctt tccccaggac cccaagcgct 781 gcgcgaggtc agcaccaagg acagcgcccg ggccgagcgc ctggcctcga acgccgggtt 841 ctgctttgtg tttccaggtt tttgtttttg gttatgagag gaggcctggc tctgtcgccc 901 gggctggcgt ccagaggcgc gatctcggct cactgcagcc tcggcctccc gagtagctgg 961 gaccgcaggc gctcaccacc acgcctggcc acaattttcc ttttttctta ttttctttta 1021 gatggagttt ggctcttgtt gcccaggctg tcgcagctgc tcattttgtg aagtgaatgc 1081 tgatttgggg gccgggtggg gttcgtctgt acatcgtgca ctgtcagacc cttcctgaag 1141 gatttttgtt actgaagtat cagaaggccc ttgttctaag gtggtggcac tgtaatatct 1201 ggcccaggag gaggtcagtg ccgtgccggc gagtcctttg ctcccttcct tctctgacca 1261 gtaaacgcat ctccagtcgc tcccccgagg cccccctgca cgcagtctcc gacctcagag 1321 agtgagtcag atagaagccg ctgcccaccc tcccccaccc tggccgggtt cttgcctggc 1381 ttgtgcgtcc ttgtgacaat tcctgacacg gatgcgcacc gggcagtggc acgtccaggt 1441 gctgtgcgcg cccgggccca caaagctcct ttggacgctc atggcagccc cttggtggag 1501 gaggtcgggt gcccaccccc tccccgccca ctgcggaagc cggcgaccca cggagctcgc 1561 tctcggccgc cgccacccct ctgtgttcgc gcccttccga gctctgatcc gacgctttgt 1621 ttcttctcag tgggttcagg gcctgggcca gcctttacct acctccccca cccaaaaccg 1681 gcaaaagctc agagcacctt gtctgccaaa agacagggag ctgggatggt gcgggttggt 1741 ctctaaaccg gcgtggggaa aaaagaccct ccgtacaaag ccgcagggtg gggctgtcgc 1801 aagggcggaa ccgagagggt agctgggggc ggggttccca gggccaagag gggccattgt 1861 cctccctgga gcccggcgcc cccacagcca gctcctctgg gagacagccc ctcctttcga 1921 atgcgcgggg ccctcagacc gcgcccggcc cagcgctggg ggatccttgg ctgcgggagg 1981 ggcgccgcat tgcgcgcggc gggcggggac gcgcggtgcg gagcctgcgg gccgggcggg 2041 gctctgcggc ggcgcctccc gattggccac ccgcggtgac atcagccgat gcgaagggcg 2101 gggccgcggc tataagagcg cgcggccgcg gtccccgacc ctcagcagcc agcccggccc 2161 gcccgcgccc gtccgcagcc gcccgccaga cgcgcccagt ggcgcgccga agttggtcgt 2221 gaggcactgg gcaggtaagt atcaaggtta caagacaggt ttaaggagac caatagaaac 2281 tgggcttgtc gagacagaga agactcttgc gtttctgata ggcacctatt ggtcttactg 2341 acatccactt tgcctttctc tccacaggtg tccactccca ggcggccgcc accatggtga 2401 gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg 2461 taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc 2521 tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga 2581 ccaccttcac ctacggcgtg cagtgcttcg cccgctaccc cgaccacatg aagcagcacg 2641 acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg 2701 acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc 2761 gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg 2821 agtacaacta caacagccac aaggtctata tcaccgccga caagcagaag aacggcatca 2881 aggtgaactt caagacccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact 2941 accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga 3001 gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg 3061 agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag taagcggccg 3121 cgatcgctca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg 3181 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt 3241 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg 3301 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc 3361 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc 3421 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc 3481 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc 3541 tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg 3601 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc 3661 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcgatcgc 3721 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa 3781 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa 3841 taaacaagtt aacaacaaca attgcattca ttttatgttt caggttcagg gggagatgtg 3901 ggaggttttt taaagcaagt aaaacctcta caaatgtggt agtcgaaatt cccgataagg 3961 atcttcgtcg acgaattccc gataaggatc ttcctagagc atggctacgt agataagtag 4021 catggcgggt taatcattaa ctacaaggaa cccctagtga tggagttggc cactccctct 4081 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt 4141 gcccgggcgg cctcagtgag cgagcgagcg cgcagcctta attaacctaa ttcactggcc 4201 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca 4261 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc 4321 caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg 4381 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct 4441 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta 4501 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa 4561 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct 4621 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc 4681 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg 4741 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt 4801 acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct 4861 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat 4921 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg 4981 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg 5041 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc 5101 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat 5161 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact 5221 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca 5281 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact 5341 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg 5401 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg 5461 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg 5521 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg 5581 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag 5641 ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc 5701 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga 5761 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat 5821 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc 5881 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag 5941 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct 6001 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac 6061 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc 6121 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg 6181 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt 6241 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt 6301 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc 6361 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca 6421 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata 6481 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg 6541 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct 6601 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta 6661 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag 6721 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga 6781 ttcattaatg cagctggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg 6841 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg 6901 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc 6961 atgattacgc cagatttaat taaggcctta attagg //