LOCUS       Exported                7210 bp ds-DNA     circular SYN 17-JUN-2024
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7210)
  AUTHORS   Addgene
  TITLE     Direct Submission
  JOURNAL   Exported Monday, Jun 17, 2024 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7210
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     promoter        197..408
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      416..425
                     /regulatory_class="other"
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             425..490
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             500..520
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             524..544
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3725..3772
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4025..4045
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    4055..4103
                     /note="synthetic polyadenylation signal"
     promoter        4115..4355
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        4384..4459
                     /label=Sa gRNA scaffold
                     /note="guide RNA scaffold for the Staphylococcus aureus 
                     CRISPR/Cas9 system"
     repeat_region   4473..4613
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      4688..5143
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        5425..5529
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5530..6390
                     /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"
     rep_origin      6561..7149
                     /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 tgcggcctct agctaggtct tgaaaggagt gggaattggc
      181 tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga
      241 ggggtcggca attgatccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat
      301 gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta
      361 gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacagga ccggtgccac
      421 catggactac aaagaccatg acggtgatta taaagatcat gacatagatt acaaggatga
      481 cgatgacaag ggtggatcgc caaagaaaaa gaggaaagtc gccccaaaga agaagcggaa
      541 ggtcggtatc cacggagtcc cagcagccaa gcggaactac atcctgggcc tggccatcgg
      601 catcaccagc gtgggctacg gcatcatcga ctacgagaca cgggacgtga tcgatgccgg
      661 cgtgcggctg ttcaaagagg ccaacgtgga aaacaacgag ggcaggcgga gcaagagagg
      721 cgccagaagg ctgaagcggc ggaggcggca tagaatccag agagtgaaga agctgctgtt
      781 cgactacaac ctgctgaccg accacagcga gctgagcggc atcaacccct acgaggccag
      841 agtgaagggc ctgagccaga agctgagcga ggaagagttc tctgccgccc tgctgcacct
      901 ggccaagaga agaggcgtgc acaacgtgaa cgaggtggaa gaggacaccg gcaacgagct
      961 gtccaccaaa gagcagatca gccggaacag caaggccctg gaagagaaat acgtggccga
     1021 actgcagctg gaacggctga agaaagacgg cgaagtgcgg ggcagcatca acagattcaa
     1081 gaccagcgac tacgtgaaag aagccaaaca gctgctgaag gtgcagaagg cctaccacca
     1141 gctggaccag agcttcatcg acacctacat cgacctgctg gaaacccggc ggacctacta
     1201 tgagggacct ggcgagggca gccccttcgg ctggaaggac atcaaagaat ggtacgagat
     1261 gctgatgggc cactgcacct acttccccga ggaactgcgg agcgtgaagt acgcctacaa
     1321 cgccgacctg tacaacgccc tgaacgacct gaacaatctc gtgatcacca gggacgagaa
     1381 cgagaagctg gaatattacg agaagttcca gatcatcgag aacgtgttca agcagaagaa
     1441 gaagcccacc ctgaagcaga tcgccaaaga aatcctcgtg aacgaagagg atattaaggg
     1501 ctacagagtg accagcaccg gcaagcccga gttcaccaac ctgaaggtgt accacgacat
     1561 caaggacatt accgcccgga aagagattat tgagaacgcc gagctgctgg atcagattgc
     1621 caagatcctg accatctacc agagcagcga ggacatccag gaagaactga ccaatctgaa
     1681 ctccgagctg acccaggaag agatcgagca gatctctaat ctgaagggct ataccggcac
     1741 ccacaacctg agcctgaagg ccatcaacct gatcctggac gagctgtggc acaccaacga
     1801 caaccagatc gctatcttca accggctgaa gctggtgccc aagaaggtgg acctgtccca
     1861 gcagaaagag atccccacca ccctggtgga cgacttcatc ctgagccccg tcgtgaagag
     1921 aagcttcatc cagagcatca aagtgatcaa cgccatcatc aagaagtacg gcctgcccaa
     1981 cgacatcatt atcgagctgg cccgcgagaa gaactccaag gacgcccaga aaatgatcaa
     2041 cgagatgcag aagcggaacc ggcagaccaa cgagcggatc gaggaaatca tccggaccac
     2101 cggcaaagag aacgccaagt acctgatcga gaagatcaag ctgcacgaca tgcaggaagg
     2161 caagtgcctg tacagcctgg aagccatccc tctggaagat ctgctgaaca accccttcaa
     2221 ctatgaggtg gaccacatca tccccagaag cgtgtccttc gacaacagct tcaacaacaa
     2281 ggtgctcgtg aagcaggaag aagccagcaa gaagggcaac cggaccccat tccagtacct
     2341 gagcagcagc gacagcaaga tcagctacga aaccttcaag aagcacatcc tgaatctggc
     2401 caagggcaag ggcagaatca gcaagaccaa gaaagagtat ctgctggaag aacgggacat
     2461 caacaggttc tccgtgcaga aagacttcat caaccggaac ctggtggata ccagatacgc
     2521 caccagaggc ctgatgaacc tgctgcggag ctacttcaga gtgaacaacc tggacgtgaa
     2581 agtgaagtcc atcaatggcg gcttcaccag ctttctgcgg cggaagtgga agtttaagaa
     2641 agagcggaac aaggggtaca agcaccacgc cgaggacgcc ctgatcattg ccaacgccga
     2701 tttcatcttc aaagagtgga agaaactgga caaggccaaa aaagtgatgg aaaaccagat
     2761 gttcgaggaa aagcaggccg agagcatgcc cgagatcgaa accgagcagg agtacaaaga
     2821 gatcttcatc accccccacc agatcaagca cattaaggac ttcaaggact acaagtacag
     2881 ccaccgggtg gacaagaagc ctaatagaga gctgattaac gacaccctgt actccacccg
     2941 gaaggacgac aagggcaaca ccctgatcgt gaacaatctg aacggcctgt acgacaagga
     3001 caatgacaag ctgaaaaagc tgatcaacaa gagccccgaa aagctgctga tgtaccacca
     3061 cgacccccag acctaccaga aactgaagct gattatggaa cagtacggcg acgagaagaa
     3121 tcccctgtac aagtactacg aggaaaccgg gaactacctg accaagtact ccaaaaagga
     3181 caacggcccc gtgatcaaga agattaagta ttacggcaac aaactgaacg cccatctgga
     3241 catcaccgac gactacccca acagcagaaa caaggtcgtg aagctgtccc tgaagcccta
     3301 cagattcgac gtgtacctgg acaatggcgt gtacaagttc gtgaccgtga agaatctgga
     3361 tgtgatcaaa aaagaaaact actacgaagt gaatagcaag tgctatgagg aagctaagaa
     3421 gctgaagaag atcagcaacc aggccgagtt tatcgcctcc ttctacaaca acgatctgat
     3481 caagatcaac ggcgagctgt atagagtgat cggcgtgaac aacgacctgc tgaaccggat
     3541 cgaagtgaac atgatcgaca tcacctaccg cgagtacctg gaaaacatga acgacaagag
     3601 gccccccagg atcattaaga caatcgcctc caagacccag agcattaaga agtacagcac
     3661 agacattctg ggcaacctgt atgaagtgaa atctaagaag caccctcaga tcatcaaaaa
     3721 gggcaaaagg ccggcggcca cgaaaaaggc cggccaggca aaaaagaaaa agggatccgg
     3781 tggaggaagt ggcgggtcag ggtcgggtgg cactagtcga ggctttgctc gaggtttgga
     3841 gccggagcgg attattggag ctacagactc cagtggagag ctcatgttcc tgatgaaatg
     3901 gaaaaactct gatgaggctg acctggtccc tgccaaggaa gccaatgtca agtgcccaca
     3961 ggttgtcata tccttctatg aggaaaggct gacgtggcat tcctacgcta gcggaggtgg
     4021 ttcgccaaag aagaaacgga aggtgggatc ctgaaataaa agatctttgt tttcattaga
     4081 tctgtgtgtt ggttttttgt gtgcggccgg taccgagggc ctatttccca tgattccttc
     4141 atatttgcat atacgataca aggctgttag agagataatt ggaattaatt tgactgtaaa
     4201 cacaaagata ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc
     4261 agttttaaaa ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt
     4321 cgatttcttg gctttatata tcttgtggaa aggacgaaac accggagacc acggcaggtc
     4381 tcagttttag tactctggaa acagaatcta ctaaaacaag gcaaaatgcc gtgtttatct
     4441 cgtcaacttg ttggcgagat ttttgcggcc gcaggaaccc ctagtgatgg agttggccac
     4501 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     4561 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct
     4621 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa
     4681 ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     4741 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     4801 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     4861 cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt
     4921 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     4981 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggg ctattctttt
     5041 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     5101 aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt
     5161 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     5221 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     5281 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     5341 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     5401 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     5461 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     5521 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     5581 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     5641 ttgggtgcac gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt
     5701 tttcgccccg aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg
     5761 gtattatccc gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag
     5821 aatgacttgg ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta
     5881 agagaattat gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     5941 acaacgatcg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     6001 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     6061 accacgatgc ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt
     6121 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     6181 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     6241 cgtggaagcc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     6301 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     6361 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     6421 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     6481 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     6541 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     6601 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     6661 tttccgaagg taactggctt cagcagagcg cagataccaa atactgtcct tctagtgtag
     6721 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     6781 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     6841 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     6901 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     6961 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     7021 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     7081 gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     7141 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     7201 gctcacatgt
//
