LOCUS       Exported                7339 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 7339)
  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..7339
                     /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             4154..4174
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    4184..4232
                     /note="synthetic polyadenylation signal"
     promoter        4244..4484
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        4513..4588
                     /label=Sa gRNA scaffold
                     /note="guide RNA scaffold for the Staphylococcus aureus 
                     CRISPR/Cas9 system"
     repeat_region   4602..4742
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      4817..5272
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        5554..5658
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5659..6519
                     /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      6690..7278
                     /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 cactagtaag atgaaggagg gtgaaaataa
     3841 taaacccagg gagaagtcag aaagtaacaa gaggaaatcc aatttctcaa acagtgccga
     3901 tgacatcaaa tctaaaaaaa agagagagca gagcaatgat atcgctcggg gctttgagag
     3961 aggactggaa ccagaaaaga tcattggggc aacagattcc tgtggtgatt taatgttcct
     4021 aatgaaatgg aaagacacag atgaagctga cctggttctt gcaaaagaag ctaatgtgaa
     4081 atgtccacaa attgtgatag cattttatga agagagactg acatggcatg catatgctag
     4141 cggaggtggt tcgccaaaga agaaacggaa ggtgggatcc tgaaataaaa gatctttgtt
     4201 ttcattagat ctgtgtgttg gttttttgtg tgcggccggt accgagggcc tatttcccat
     4261 gattccttca tatttgcata tacgatacaa ggctgttaga gagataattg gaattaattt
     4321 gactgtaaac acaaagatat tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg
     4381 gtagtttgca gttttaaaat tatgttttaa aatggactat catatgctta ccgtaacttg
     4441 aaagtatttc gatttcttgg ctttatatat cttgtggaaa ggacgaaaca ccggagacca
     4501 cggcaggtct cagttttagt actctggaaa cagaatctac taaaacaagg caaaatgccg
     4561 tgtttatctc gtcaacttgt tggcgagatt tttgcggccg caggaacccc tagtgatgga
     4621 gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc
     4681 ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca
     4741 ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg
     4801 tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     4861 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     4921 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     4981 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat
     5041 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     5101 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc
     5161 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     5221 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc
     5281 actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca
     5341 cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg
     5401 accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga
     5461 cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct
     5521 tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     5581 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     5641 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     5701 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     5761 gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc
     5821 cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta
     5881 tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac
     5941 tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc
     6001 atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac
     6061 ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg
     6121 gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac
     6181 gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc
     6241 gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt
     6301 gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga
     6361 gccggtgagc gtggaagccg cggtatcatt gcagcactgg ggccagatgg taagccctcc
     6421 cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag
     6481 atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca
     6541 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     6601 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     6661 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     6721 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     6781 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt
     6841 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     6901 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     6961 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     7021 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     7081 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     7141 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     7201 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     7261 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     7321 tggccttttg ctcacatgt
//
