LOCUS       62056_22900        7354 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 7354)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7354
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     393..409
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        417..435
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     3'UTR           complement(541..666)
                     /label=Xenopus globin 3'-UTR
                     /note="3'-UTR of the major beta-globin gene of Xenopus
                     laevis"
     CDS             complement(750..4853)
                     /codon_start=1
                     /label=Cas9
                     /note="Cas9 (Csn1) endonuclease from the Streptococcus
                     pyogenes Type II CRISPR/Cas system"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(4878..4898)
                     /codon_start=1
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /translation="PKKKRKV"
     5'UTR           complement(4923..4965)
                     /label=Xenopus globin 5'-UTR
                     /note="translational enhancer from the 5'-UTR of the major 
                     beta-globin gene of Xenopus laevis"
     promoter        complement(4978..4996)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5015..5031)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(5039..5055)
                     /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(5063..5093)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(5108..5129)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5417..6005)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(6179..7036)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(7037..7141)
                     /label=AmpR promoter
ORIGIN
        1 tagcgtatca gagtctcgtc cgtcggtcgt atgacggtga aacctctgac acatgcagct
       61 cccggagacg gtcacagctt gtctgtaagc ggatgccggg agcagacaag cccgtcaggg
      121 cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac tatgcggcat cagagcagat
      181 tgtactgaga gtgcaccata tgcggtgtga aataccgcac agatgcgtaa ggagaaaata
      241 ccgcatcagg cgccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
      301 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
      361 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattggattt
      421 aggtgacact atagaatacg aattaattcg agctcgcccg gggatcctct agagtcgacc
      481 tgcagggggg gggggggggg gttttttttt tttttttttt tttttttttt tttttagaat
      541 gtgaagaaac tttcttttta ttaggagcag atacgaatgg ctacattttg ggggacaaca
      601 ttttgtaaag tgtaagttgg tattatgtag cttagagact ccattcgggt gttcttgagg
      661 ctggtttagt ggtaaccaga tcctagtcag tcactagatt atttcttttt cttagcttga
      721 ccagctttct tagtagcagc aggacgcttg tctccaccga gctgagagag gtcgattctt
      781 gtttcataga gccccgtaat tgactgatga atcagtgtgg cgtccaggac ctcctttgta
      841 gaggtgtacc gctttctgtc tatggtggtg tcgaagtact tgaaggctgc aggcgcgccc
      901 aagttggtca gagtaaacaa gtggataatg ttttctgcct gctccctgat gggcttatcc
      961 ctgtgcttat tgtaagcaga aagcacctta tcgaggttag cgtcggcgag gatcactctt
     1021 ttggagaatt cgcttatttg ctcgatgatc tcatcaaggt agtgtttgtg ttgttccacg
     1081 aacagctgct tctgctcatt atcttcggga gaccctttga gcttttcata gtggctggcc
     1141 agatacaaga aattaacgta tttagagggc agtgccagct cgttaccttt ctgcagctcg
     1201 cccgcactag cgagcattcg tttccggccg ttttcaagct caaagagaga gtacttggga
     1261 agcttaatga tgaggtcttt tttgacctct ttatatcctt tcgcctcgag aaagtcgatg
     1321 gggttttttt cgaagcttga tcgctccatg attgtgatgc ccagcagttc cttgacgctt
     1381 ttgagttttt tagacttccc tttctccact ttggccacaa ccagtacact gtaagcgact
     1441 gtaggagaat cgaatccgcc gtatttcttg gggtcccaat cttttttgcg tgcgatcagc
     1501 ttgtcgctgt tccttttcgg gaggatactt tccttggaga agcctccggt ctgtacttcg
     1561 gtctttttaa cgatgttcac ctgcggcatg gacaggacct tccggactgt cgcgaaatcc
     1621 ctacccttgt cccacacgat ttctcctgtt tctccgtttg tttcgataag tggtcgcttc
     1681 cgaatctctc cattggccag tgtaatctcg gtcttgaaaa aattcataat attgctgtaa
     1741 aagaagtact tagcggtggc cttgcctatt tcctgctcag actttgcgat cattttccta
     1801 acatcgtaca ctttatagtc tccgtaaaca aattcagatt caagcttggg atattttttg
     1861 ataagtgcag tgcctaccac tgcattcagg taggcatcat gcgcatggtg gtaattgttg
     1921 atctctctca ccttataaaa ctgaaagtcc tttctgaaat ctgagaccag cttagacttc
     1981 agagtaataa ctttcacctc tcgaatcagt ttgtcatttt catcgtactt ggtgttcatg
     2041 cgtgaatcga gaatttgggc cacgtgcttg gtgatctggc gtgtctcaac aagctgcctt
     2101 ttgatgaagc ctgctttatc caactcagac aggccacctc gttcagcctt agtcagatta
     2161 tcgaacttcc gttgtgtgat cagtttggcg ttcagcagct gccgccaata atttttcatt
     2221 ttcttgacaa cttcttctga ggggacgtta tcactcttcc ctctattttt atcggatctt
     2281 gtcaacactt tattatcaat agaatcatct ttgagaaaag actggggcac gatatgatcc
     2341 acgtcgtagt cggagagccg attgatgtcc agttcctgat ccacgtacat gtccctgccg
     2401 ttctgcaggt agtacaggta gagcttctca ttctgaagct gggtgttttc aactgggtgt
     2461 tccttaagga tttgggaccc cagttctttt ataccctctt caatcctctt catcctttcc
     2521 ctactgttct tctgtccctt ctgggtagtt tggttctctc gggccatctc gataacgata
     2581 ttctcgggct tatgccttcc cattactttg acgagttcat ccacgacctt aacggtctgc
     2641 agtattccct ttttgatagc tgggctacct gcaagattag cgatgtgctc gtgaagactg
     2701 tccccctggc cagaaacttg tgctttctgg atgtcctcct taaaggtgag agagtcatca
     2761 tggatcaact gcatgaagtt ccggttggca aatccatcgg acttaagaaa atccaggatt
     2821 gtctttccac tctgcttgtc tcggatccca ttgatcagtt ttcttgacag ccgcccccat
     2881 cctgtatatc ggcgcctctt gagctgtttc atgactttgt cgtcgaagag atgagcgtaa
     2941 gttttcaagc gttcttcaat catctcccta tcttcaaaca acgtaagggt gaggacaatg
     3001 tcctcaagaa tgtcctcgtt ctcctcattg tccaggaagt ccttgtcttt aatgattttc
     3061 aggagatcgt gatacgttcc cagggatgcg ttgaagcgat cctccactcc gctgatttca
     3121 acagagtcga aacattcaat ctttttgaaa tagtcttctt tgagctgttt cacggtaact
     3181 ttccggttcg tcttgaagag gaggtccacg atagctttct tctgctctcc agacaggaat
     3241 gctggctttc tcatcccttc tgtgacgtat ttgaccttgg tgagctcgtt ataaactgtg
     3301 aagtactcgt acagcagaga gtgtttagga agcacctttt cgttaggcag atttttatca
     3361 aagttagtca tcctttcgat gaaggactgg gcagaggccc ccttatccac gacttcctcg
     3421 aagttccagg gagtgatggt ctcttctgat ttgcgagtca tccacgcgaa tctggaattt
     3481 ccccgggcga gggggcctac atagtagggt atccgaaatg tgaggatttt ctcaatcttt
     3541 tccctgttat ctttcaaaaa ggggtagaaa tcctcttgcc gcctgagtat agcgtgcagt
     3601 tcgcccaggt gaatctggtg ggggatgctt ccattgtcga aagtgcgctg tttgcgcaac
     3661 agatcttctc tgttaagctt taccagcagc tcctcggtgc cgtccatttt ttccaagatg
     3721 ggcttaataa atttgtaaaa ttcctcctgg cttgctccgc cgtcaatgta tccggcgtag
     3781 ccatttttag actgatcgaa gaaaatttcc ttgtacttct caggcagttg ctgtctgaca
     3841 agggccttca gcaaagtcaa gtcttggtgg tgctcatcat agcgcttgat catactagcg
     3901 ctcagcggag ctttggtgat ctccgtgttc actcgcagaa tatcactcag cagaatggcg
     3961 tctgacaggt tctttgccgc caaaaaaagg tctgcgtact ggtcgccgat ctgggccagc
     4021 agattgtcga gatcatcatc gtaggtgtct ttgctcagtt gaagcttggc atcttcggcc
     4081 aggtcgaagt tagatttaaa gttgggggtc agcccgagtg acagggcgat aagattacca
     4141 aacaggccgt tcttcttctc cccagggagc tgtgcgatga ggttttcgag ccgccgggat
     4201 ttggacagcc tagcgctcag gattgctttg gcgtcaactc cggatgcgtt gatcgggttc
     4261 tcttcgaaaa gctgattgta agtctgaacc agttggataa agagtttgtc gacatcgctg
     4321 ttgtctgggt tcaggtcccc ctcgatgagg aagtgtcccc gaaatttgat catatgcgcc
     4381 agcgcgagat agatcaaccg caagtcagcc ttatcagtac tgtctacaag cttcttcctc
     4441 agatgatata tggttgggta cttttcatgg tacgccacct cgtccacgat attgccaaag
     4501 attgggtggc gctcgtgctt tttatcctcc tccaccaaaa aggactcctc cagcctatgg
     4561 aagaaagagt catccacctt agccatctca ttactaaaga tctcctgcag gtagcagatc
     4621 cgattctttc tgcgggtata tctgcgccgt gctgttcttt tgagccgcgt ggcttcggcc
     4681 gtctccccgg agtcgaacag gagggcgcca atgaggttct tctttatgct gtggcgatcg
     4741 gtattgccca gaactttgaa ttttttgctc ggcaccttgt actcgtccgt aatgacggcc
     4801 cagccgacgc tgtttgtgcc gatatcgagc ccaatggagt acttcttgtc catggctgct
     4861 gggacaccgt ggataccgac cttccgcttc ttctttgggg ccatggtggc ggtaccgatc
     4921 tgccaaagtt gagcgtttat tctgagcttc tgcaaaaaga acaagcaagc ttgtctccct
     4981 atagtgagtc gtattagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
     5041 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
     5101 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
     5161 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     5221 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     5281 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     5341 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5401 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5461 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5521 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5581 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     5641 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     5701 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     5761 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     5821 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     5881 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     5941 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6001 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6061 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6121 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6181 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6241 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     6301 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6361 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6421 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6481 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6541 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     6601 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     6661 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     6721 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     6781 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6841 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6901 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6961 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     7021 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     7081 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     7141 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     7201 cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg
     7261 aaaacctctg acacatgcag ctcccggaga cggtcacagc ttgtctgtaa gcggatgccg
     7321 ggagcagaca agcccgtcag ggcgcgtcag cggt
//
