LOCUS Exported 6985 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION AAV plasmid with Ple331 (PAX6 MiniPromoter) driving expression of EmGFP. Contains WPRE.. ACCESSION . VERSION . KEYWORDS pEMS2260 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 May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..6985 /organism="synthetic DNA construct" /mol_type="other DNA" 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 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 2383..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 4927..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" primer_bind 6700..6717 /label=L4440 /note="L4440 vector, 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 6870..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 //