LOCUS       Exported                7425 bp DNA     circular SYN 03-AUG-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7425)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 16, 2022 from SnapGene 6.0.5
            https://www.snapgene.com
COMMENT     LOCUS       pAAV-EF1a-EYFP-W        6067 bp    DNA     circular     
            20-MAY-2010

            SOURCE      

              ORGANISM  

            COMMENT     VNTDATE|538328968|

            COMMENT     VNTDBDATE|538763081|

            COMMENT     VNTNAME|pAAV-EF1a-EYFP-WPRE|

            COMMENT     VNTAUTHORNAME|William Hendriks|

            COMMENT     VNTAUTHOREML|hendriks.william@mgh.harvard.edu|

            COMMENT     VNTAUTHORNAME|Feng Zhang|

            COMMENT     VNTAUTHOREML|zhang72@fas.harvard.edu|
FEATURES             Location/Qualifiers
     source          1..7425
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     misc_feature    150..155
                     /label=MluI
                     /note="MluI"
     promoter        201..412
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     misc_feature    218..238
                     /label=EF1a-R
                     /note="EF1a-R"
     misc_feature    393..419
                     /label=pSC028_F2
                     /note="pSC028_F2"
     misc_feature    393..416
                     /label=pSC028_F1
                     /note="pSC028_F1"
     primer_bind     393..413
                     /label=GGTTTGCCGCCAGAACACAGG
     misc_feature    402..449
                     /label=pGW07-F1
                     /note="pGW07-F1"
     promoter        414..426
                     /label=EFS
                     /note="EFS"
     misc_feature    426
                     /label=BamHI site
                     /note="BamHI site"
     CDS             434..481
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             482..4582
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     misc_feature    1614..1633
                     /label=GtC9-F2
                     /note="GtC9-F2"
     misc_feature    1943..1962
                     /label=GTC9-R2
                     /note="GTC9-R2"
     misc_feature    2498..2517
                     /label=GtC9-F1
                     /note="GtC9-F1"
     misc_feature    3140..3159
                     /label=GtC9-F3
                     /note="GtC9-F3"
     misc_feature    3318..3337
                     /label=GtC9-R1
                     /note="GtC9-R1"
     misc_feature    3788..3807
                     /label=pGW07-F2
                     /note="pGW07-F2"
     misc_feature    3894..3913
                     /label=GtC9-R3
                     /note="GtC9-R3"
     CDS             4586..4606
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    4623..4671
                     /note="synthetic polyadenylation signal"
     repeat_region   4688..4828
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      4903..5358
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     exon            4920..5226
                     /label=f1 Origin
     promoter        5640..5744
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5745..6605
                     /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"
     exon            5745..6602
                     /label=AmpR
     rep_origin      6753..7420
                     /direction=RIGHT
                     /label=pUC ori
     rep_origin      6776..7364
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgttctag gtcttgaaag gagtgggaat
      181 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      241 gggaggggtc ggcaattgat ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      301 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      361 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtgtcgtg
      421 acgcggatcc atgaaaaggc cggcggccac gaaaaaggcc ggccaggcaa aaaagaaaaa
      481 ggacaagaag tacagcatcg gcctggccat cggcaccaac tctgtgggct gggccgtgat
      541 caccgacgag tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca
      601 cagcatcaag aagaacctga tcggagccct gctgttcgac agcggcgaaa cagccgaggc
      661 cacccggctg aagagaaccg ccagaagaag atacaccaga cggaagaacc ggatctgcta
      721 tctgcaagag atcttcagca acgagatggc caaggtggac gacagcttct tccacagact
      781 ggaagagtcc ttcctggtgg aagaggataa gaagcacgag cggcacccca tcttcggcaa
      841 catcgtggac gaggtggcct accacgagaa gtaccccacc atctaccacc tgagaaagaa
      901 actggtggac agcaccgaca aggccgacct gcggctgatc tatctggccc tggcccacat
      961 gatcaagttc cggggccact tcctgatcga gggcgacctg aaccccgaca acagcgacgt
     1021 ggacaagctg ttcatccagc tggtgcagac ctacaaccag ctgttcgagg aaaaccccat
     1081 caacgccagc ggcgtggacg ccaaggccat cctgtctgcc agactgagca agagcagacg
     1141 gctggaaaat ctgatcgccc agctgcccgg cgagaagaag aatggcctgt tcggcaacct
     1201 gattgccctg agcctgggcc tgacccccaa cttcaagagc aacttcgacc tggccgagga
     1261 tgccaaactg cagctgagca aggacaccta cgacgacgac ctggacaacc tgctggccca
     1321 gatcggcgac cagtacgccg acctgtttct ggccgccaag aacctgtccg acgccatcct
     1381 gctgagcgac atcctgagag tgaacaccga gatcaccaag gcccccctga gcgcctctat
     1441 gatcaagaga tacgacgagc accaccagga cctgaccctg ctgaaagctc tcgtgcggca
     1501 gcagctgcct gagaagtaca aagagatttt cttcgaccag agcaagaacg gctacgccgg
     1561 ctacattgac ggcggagcca gccaggaaga gttctacaag ttcatcaagc ccatcctgga
     1621 aaagatggac ggcaccgagg aactgctcgt gaagctgaac agagaggacc tgctgcggaa
     1681 gcagcggacc ttcgacaacg gcagcatccc ccaccagatc cacctgggag agctgcacgc
     1741 cattctgcgg cggcaggaag atttttaccc attcctgaag gacaaccggg aaaagatcga
     1801 gaagatcctg accttccgca tcccctacta cgtgggccct ctggccaggg gaaacagcag
     1861 attcgcctgg atgaccagaa agagcgagga aaccatcacc ccctggaact tcgaggaagt
     1921 ggtggacaag ggcgcttccg cccagagctt catcgagcgg atgaccaact tcgataagaa
     1981 cctgcccaac gagaaggtgc tgcccaagca cagcctgctg tacgagtact tcaccgtgta
     2041 taacgagctg accaaagtga aatacgtgac cgagggaatg agaaagcccg ccttcctgag
     2101 cggcgagcag aaaaaggcca tcgtggacct gctgttcaag accaaccgga aagtgaccgt
     2161 gaagcagctg aaagaggact acttcaagaa aatcgagtgc ttcgactccg tggaaatctc
     2221 cggcgtggaa gatcggttca acgcctccct gggcacatac cacgatctgc tgaaaattat
     2281 caaggacaag gacttcctgg acaatgagga aaacgaggac attctggaag atatcgtgct
     2341 gaccctgaca ctgtttgagg acagagagat gatcgaggaa cggctgaaaa cctatgccca
     2401 cctgttcgac gacaaagtga tgaagcagct gaagcggcgg agatacaccg gctggggcag
     2461 gctgagccgg aagctgatca acggcatccg ggacaagcag tccggcaaga caatcctgga
     2521 tttcctgaag tccgacggct tcgccaacag aaacttcatg cagctgatcc acgacgacag
     2581 cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg atagcctgca
     2641 cgagcacatt gccaatctgg ccggcagccc cgccattaag aagggcatcc tgcagacagt
     2701 gaaggtggtg gacgagctcg tgaaagtgat gggccggcac aagcccgaga acatcgtgat
     2761 cgaaatggcc agagagaacc agaccaccca gaagggacag aagaacagcc gcgagagaat
     2821 gaagcggatc gaagagggca tcaaagagct gggcagccag atcctgaaag aacaccccgt
     2881 ggaaaacacc cagctgcaga acgagaagct gtacctgtac tacctgcaga atgggcggga
     2941 tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc gactacgatg tggaccacat
     3001 cgtgcctcag agctttctga aggacgactc catcgacaac aaggtgctga ccagaagcga
     3061 caaggcccgg ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa
     3121 ctactggcgg cagctgctga acgccaagct gattacccag agaaagttcg acaatctgac
     3181 caaggccgag agaggcggcc tgagcgaact ggataaggcc ggcttcatca agagacagct
     3241 ggtggaaacc cggcagatca caaagcacgt ggcacagatc ctggactccc ggatgaacac
     3301 taagtacgac gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa
     3361 gctggtgtcc gatttccgga aggatttcca gttttacaaa gtgcgcgaga tcaacaacta
     3421 ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga accgccctga tcaaaaagta
     3481 ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat
     3541 gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc aagtacttct tctacagcaa
     3601 catcatgaac tttttcaaga ccgagattac cctggccaac ggcgagatcc ggaagcggcc
     3661 tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg gataagggcc gggattttgc
     3721 caccgtgcgg aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca
     3781 gacaggcggc ttcagcaaag agtctatcct gcccaagagg aacagcgata agctgatcgc
     3841 cagaaagaag gactgggacc ctaagaagta cggcggcttc gacagcccca ccgtggccta
     3901 ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa
     3961 agagctgctg gggatcacca tcatggaaag aagcagcttc gagaagaatc ccatcgactt
     4021 tctggaagcc aagggctaca aagaagtgaa aaaggacctg atcatcaagc tgcctaagta
     4081 ctccctgttc gagctggaaa acggccggaa gagaatgctg gcctctgccg gcgaactgca
     4141 gaagggaaac gaactggccc tgccctccaa atatgtgaac ttcctgtacc tggccagcca
     4201 ctatgagaag ctgaagggct cccccgagga taatgagcag aaacagctgt ttgtggaaca
     4261 gcacaagcac tacctggacg agatcatcga gcagatcagc gagttctcca agagagtgat
     4321 cctggccgac gctaatctgg acaaagtgct gtccgcctac aacaagcacc gggataagcc
     4381 catcagagag caggccgaga atatcatcca cctgtttacc ctgaccaatc tgggagcccc
     4441 tgccgccttc aagtactttg acaccaccat cgaccggaag aggtacacca gcaccaaaga
     4501 ggtgctggac gccaccctga tccaccagag catcaccggc ctgtacgaga cacggatcga
     4561 cctgtctcag ctgggaggcg acagccctaa gaaaaagagg aaggtgagct gagaattcaa
     4621 taaataaaag atctttattt tcattagatc tgtgtgttgg ttttttgtgt gcggaccgag
     4681 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     4741 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     4801 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     4861 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     4921 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
     4981 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     5041 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     5101 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     5161 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     5221 gaacaacact caaccctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     5281 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     5341 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     5401 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     5461 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     5521 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     5581 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     5641 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     5701 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     5761 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     5821 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     5881 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     5941 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     6001 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     6061 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     6121 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     6181 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     6241 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     6301 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     6361 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     6421 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     6481 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     6541 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     6601 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     6661 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     6721 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     6781 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     6841 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     6901 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     6961 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     7021 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     7081 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     7141 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     7201 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     7261 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     7321 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     7381 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
