LOCUS       62056_6025        8246 bp DNA     circular SYN 01-JAN-1980
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8246)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8246
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             944..1009
                     /codon_start=1
                     /label=3xFLAG
                     /note="three tandem FLAG(R) epitope tags, followed by an 
                     enterokinase cleavage site"
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1016..1036
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     CDS             1061..3718
                     /codon_start=1
                     /label=NgAgo
                     /note="Argonaute protein from Natronobacterium gregoryi
                     (Gao et al., 2016)"
                     /translation="TVIDLDSTTTADELTSGHTYDISVTLTGVYDNTDEQHPRMSLAFE
                     QDNGERRYITLWKNTTPKDVFTYDYATGSTYIFTNIDYEVKDGYENLTATYQTTVENAT
                     AQEVGTTDEDETFAGGEPLDHHLDDALNETPDDAETESDSGHVMTSFASRDQLPEWTLH
                     TYTLTATDGAKTDTEYARRTLAYTVRQELYTDHDAAPVATDGLMLLTPEPLGETPLDLD
                     CGVRVEADETRTLDYTTAKDRLLARELVEEGLKRSLWDDYLVRGIDEVLSKEPVLTCDE
                     FDLHERYDLSVEVGHSGRAYLHINFRHRFVPKLTLADIDDDNIYPGLRVKTTYRPRRGH
                     IVWGLRDECATDSLNTLGNQSVVAYHRNNQTPINTDLLDAIEAADRRVVETRRQGHGDD
                     AVSFPQELLAVEPNTHQIKQFASDGFHQQARSKTRLSASRCSEKAQAFAERLDPVRLNG
                     STVEFSSEFFTGNNEQQLRLLYENGESVLTFRDGARGAHPDETFSKGIVNPPESFEVAV
                     VLPEQQADTCKAQWDTMADLLNQAGAPPTRSETVQYDAFSSPESISLNVAGAIDPSEVD
                     AAFVVLPPDQEGFADLASPTETYDELKKALANMGIYSQMAYFDRFRDAKIFYTRNVALG
                     LLAAAGGVAFTTEHAMPGDADMFIGIDVSRSYPEDGASGQINIAATATAVYKDGTILGH
                     SSTRPQLGEKLQSTDVRDIMKNAILGYQQVTGESPTHIVIHRDGFMNEDLDPATEFLNE
                     QGVEYDIVEIRKQPQTRLLAVSDVQYDTPVKSIAAINQNEPRATVATFGAPEYLATRDG
                     GGLPRPIQIERVAGETDIETLTRQVYLLSQSHIQVHNSTARLPITTAYADQASTHATKG
                     YLVQTGAFESNVGFL"
     CDS             3719..3766
                     /codon_start=1
                     /label=nucleoplasmin NLS
                     /note="bipartite nuclear localization signal from
                     nucleoplasmin"
                     /translation="KRPAATKKAGQAKKKK"
     polyA_signal    3846..4070
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4116..4544
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        4558..4887
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             4954..5745
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    5922..6055
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6092..6108)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(6116..6132)
                     /label=lac operator
                     /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)."
     promoter        complement(6140..6170)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(6185..6206)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(6494..7079)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7253..8110)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(8111..8215)
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccgccacc atggactata aggaccacga
      961 cggagactac aaggatcatg atattgatta caaagacgat gacgataaga tggccccaaa
     1021 gaagaagcgg aaggtcggta tccacggagt cccagcagcc accgtgatcg acctggacag
     1081 caccaccacc gccgacgagc tgaccagcgg ccacacctac gacatcagcg tgaccctgac
     1141 cggcgtgtac gacaacaccg acgagcagca cccccgcatg agcctggcct tcgagcagga
     1201 caacggcgag cgccgctaca tcaccctgtg gaagaacacc acccccaagg acgtgttcac
     1261 ctacgactac gccaccggca gcacctacat cttcaccaac atcgactacg aggtgaagga
     1321 cggctacgag aacctgaccg ccacctacca gaccaccgtg gagaacgcca ccgcccagga
     1381 ggtgggcacc accgacgagg acgagacctt cgccggcggc gagcccctgg accaccacct
     1441 ggacgacgcc ctgaacgaga cccccgacga cgccgagacc gagagcgaca gcggccacgt
     1501 gatgaccagc ttcgccagcc gcgaccagct gcccgagtgg accctgcaca cctacaccct
     1561 gaccgccacc gacggcgcca agaccgacac cgagtacgcc cgccgcaccc tggcctacac
     1621 cgtgcgccag gagctgtaca ccgaccacga cgccgccccc gtggccaccg acggcctgat
     1681 gctgctgacc cccgagcccc tgggcgagac ccccctggac ctggactgcg gcgtgcgcgt
     1741 ggaggccgac gagacccgca ccctggacta caccaccgcc aaggaccgcc tgctggcccg
     1801 cgagctggtg gaggagggcc tgaagcgcag cctgtgggac gactacctgg tgcgcggcat
     1861 cgacgaggtg ctgagcaagg agcccgtgct gacctgcgac gagttcgacc tgcacgagcg
     1921 ctacgacctg agcgtggagg tgggccacag cggccgcgcc tacctgcaca tcaacttccg
     1981 ccaccgcttc gtgcccaagc tgaccctggc cgacatcgac gacgacaaca tctaccccgg
     2041 cctgcgcgtg aagaccacct accgcccccg ccgcggccac atcgtgtggg gcctgcgcga
     2101 cgagtgcgcc accgacagcc tgaacaccct gggcaaccag agcgtggtgg cctaccaccg
     2161 caacaaccag acccccatca acaccgacct gctggacgcc atcgaggccg ccgaccgccg
     2221 cgtggtggag acccgccgcc agggccacgg cgacgacgcc gtgagcttcc cccaggagct
     2281 gctggccgtg gagcccaaca cccaccagat caagcagttc gccagcgacg gcttccacca
     2341 gcaggcccgc agcaagaccc gcctgagcgc cagccgctgc agcgagaagg cccaggcctt
     2401 cgccgagcgc ctggaccccg tgcgcctgaa cggcagcacc gtggagttca gcagcgagtt
     2461 cttcaccggc aacaacgagc agcagctgcg cctgctgtac gagaacggcg agagcgtgct
     2521 gaccttccgc gacggcgccc gcggcgccca ccccgacgag accttcagca agggcatcgt
     2581 gaaccccccc gagagcttcg aggtggccgt ggtgctgccc gagcagcagg ccgacacctg
     2641 caaggcccag tgggacacca tggccgacct gctgaaccag gccggcgccc cccccacccg
     2701 cagcgagacc gtgcagtacg acgccttcag cagccccgag agcatcagcc tgaacgtggc
     2761 cggcgccatc gaccccagcg aggtggacgc cgccttcgtg gtgctgcccc ccgaccagga
     2821 gggcttcgcc gacctggcca gccccaccga gacctacgac gagctgaaga aggccctggc
     2881 caacatgggc atctacagcc agatggccta cttcgaccgc ttccgcgacg ccaagatctt
     2941 ctacacccgc aacgtggccc tgggcctgct ggccgccgcc ggcggcgtgg ccttcaccac
     3001 cgagcacgcc atgcccggcg acgccgacat gttcatcggc atcgacgtga gccgcagcta
     3061 ccccgaggac ggcgccagcg gccagatcaa catcgccgcc accgccaccg ccgtgtacaa
     3121 ggacggcacc atcctgggcc acagcagcac ccgcccccag ctgggcgaga agctgcagag
     3181 caccgacgtg cgcgacatca tgaagaacgc catcctgggc taccagcagg tgaccggcga
     3241 gagccccacc cacatcgtga tccaccgcga cggcttcatg aacgaggacc tggaccccgc
     3301 caccgagttc ctgaacgagc agggcgtgga gtacgacatc gtggagatcc gcaagcagcc
     3361 ccagacccgc ctgctggccg tgagcgacgt gcagtacgac acccccgtga agagcatcgc
     3421 cgccatcaac cagaacgagc cccgcgccac cgtggccacc ttcggcgccc ccgagtacct
     3481 ggccacccgc gacggcggcg gcctgccccg ccccatccag atcgagcgcg tggccggcga
     3541 gaccgacatc gagaccctga cccgccaggt gtacctgctg agccagagcc acatccaggt
     3601 gcacaacagc accgcccgcc tgcccatcac caccgcctac gccgaccagg ccagcaccca
     3661 cgccaccaag ggctacctgg tgcagaccgg cgccttcgag agcaacgtgg gcttcctgaa
     3721 aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaagtgag aattctgcag
     3781 atatccagca cagtggcggc cgctcgagtc tagagggccc gtttaaaccc gctgatcagc
     3841 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     3901 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     3961 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     4021 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
     4081 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
     4141 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     4201 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     4261 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     4321 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     4381 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     4441 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     4501 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
     4561 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     4621 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     4681 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     4741 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     4801 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
     4861 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
     4921 tctgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     4981 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     5041 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     5101 aagaccgacc tgtccggtgc cctgaatgaa ctgcaggacg aggcagcgcg gctatcgtgg
     5161 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     5221 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     5281 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     5341 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     5401 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     5461 ctgttcgcca ggctcaaggc gcgcatgccc gacggcgagg atctcgtcgt gacccatggc
     5521 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     5581 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     5641 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     5701 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     5761 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     5821 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     5881 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     5941 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     6001 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     6061 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     6121 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     6181 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     6241 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     6301 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     6361 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     6421 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     6481 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     6541 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     6601 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     6661 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     6721 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     6781 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     6841 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     6901 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     6961 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     7021 ctggtagcgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     7081 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag
     7141 ggattttggt catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat
     7201 gaagttttaa atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct
     7261 taatcagtga ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac
     7321 tccccgtcgt gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa
     7381 tgataccgcg agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg
     7441 gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt
     7501 gttgccggga agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca
     7561 ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt
     7621 cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct
     7681 tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg
     7741 cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg
     7801 agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg
     7861 cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa
     7921 aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt
     7981 aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt
     8041 gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt
     8101 gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca
     8161 tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat
     8221 ttccccgaaa agtgccacct gacgtc
//
