LOCUS       sequence-201889-        7059 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cas9 coding gene template with optimized sequence for human codon 
            usage.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7059)
  AUTHORS   Cho N, Seo HN, Ryu T, Kwon E, Huh S, Noh J, Yeom H, Hwang B, Ha H, 
            Lee JH, Kwon S, Bang D
  TITLE     High-throughput construction of multiple cas9 gene variants via 
            assembly of high-depth tiled and sequence-verified oligonucleotides.
  JOURNAL   Nucleic Acids Res. 2018 May 18;46(9):e55. doi: 10.1093/nar/gky112.
  PUBMED    29529247
REFERENCE   2  (bases 1 to 7059)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res."; date: "2018-05-18"; volume: "46(9):e55. doi"; pages: " 
            10.1093/nar/gky112"
COMMENT     Sequence Label: pUC19-Q5L8F8
FEATURES             Location/Qualifiers
     source          1..7059
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     109..128
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     322..344
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     336..353
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     337..353
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4716..4736
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4746..4762)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4746..4762)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4759..4781)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4770..4786
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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(join(4794..4800,4801..4818,4819..4824))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4839..4860
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5198..5786)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5278..5297)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5957..6748,6749..6817))
                     /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"
     primer_bind     6580..6599
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6818..6922)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6990..7008
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(join(7046..7059,1..9))
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
       61 agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc
      121 agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa
      181 aataccgcat caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
      241 tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa
      301 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgtgctgt
      361 ctcatcattt tggcaaagaa ttcgcggccg ccaccatgaa acgaattctg ggcctggatt
      421 tgggaacaaa ctctattggc tgggcgctgg tcaatgaggc cgagaacaaa gacgagcgga
      481 gtagtattgt gaaattgggc gtgagagtta acccattgac tgtcgatgaa ctgacgaatt
      541 ttgagaaagg caaaagtatc acgaccaatg ctgatcgcac gcttaagcgg ggaatgcgga
      601 ggaacctcca gcgatacaaa ctgaggagag agacattgac agaggttttg aaagagcaca
      661 aactgatcac tgaggataca atcctgagcg agaacggcaa cagaacaact tttgaaacct
      721 accgcctgcg ggcaaaggcc gtaaccgagg agatctctct cgaggagttt gccagagtct
      781 tgcttatgat aaacaagaag cgcggatata agtcttcccg gaaggcgaag ggcgtagagg
      841 agggaactct tatcgacggg atggacatcg cccgggagct gtacaacaat aatctgactc
      901 cgggggaact ctgtctgcag ttgctggacg caggcaagaa gtttcttcca gatttttatc
      961 ggtccgattt gcaaaacgag ctggatcgga tctgggaaaa acagaaggag tactatccag
     1021 agattctgac agatgttctg aaagaggagt tgagaggcaa aaagcgcgat gccgtatggg
     1081 ccatttgcgc caagtacttc gtttggaagg aaaattacac tgagtggaat aaggagaagg
     1141 gaaaaacgga gcagcaagaa cgggaacaca agctggaggg catctactcc aagaggaaac
     1201 gggatgaagc gaagagagag aacttgcagt ggcgggtcaa tggacttaag gaaaagctga
     1261 gtctggagca gctcgtgatc gtcttccagg aaatgaatac acagattaat aacagctcag
     1321 gctacctggg cgctatcagt gacagatcaa aagagttgta cttcaataag cagactgtcg
     1381 ggcagtatca gatggagatg ttggataaga atcccaacgc aagtcttagg aacatggtgt
     1441 tctacaggca agactacctg gacgaattta acatgttgtg ggaaaagcaa gcggtctacc
     1501 acaaggagct gaccgaggag ctgaagaaag agatcagaga tattatcatc ttctaccagc
     1561 gcaggcttaa gtcccagaaa ggcctgatag ggttctgcga gtttgagagc cggcaaatcg
     1621 aggtggacat cgatggcaag aagaaaatca aaaccgtggg gaacagagtg atatccagaa
     1681 gctcccctct gttccaggaa tttaaaatct ggcagatact taacaatata gaagtgaccg
     1741 ttgtagggaa gaaacggaag agacgaaagc tgaaagaaaa ctactctgcc ctcttcgaag
     1801 agcttaacga cgccgaacag ctggagctca acggcagtag acggctgtgc caagaggaaa
     1861 aagagctgct ggcacaggag ctgttcattc gggacaagat gacgaagagt gaagtgttga
     1921 agctgctgtt cgacaatcca caagaattgg accttaactt caaaactatc gatggaaata
     1981 agactggcta cgctttgttc caagcctatt ctaagatgat tgagatgagc ggacatgaac
     2041 ccgtggactt taagaagcca gttgagaagg tggtggagta cattaaggcc gtctttgatc
     2101 tgttgaattg gaataccgac atcctgggct ttaattcaaa cgaggaactc gacaaccagc
     2161 cctattataa gctctggcat ttgctttata gctttgaggg agataacaca ccaacaggaa
     2221 acggaagact gatccagaaa atgactgagc tttacgggtt tgaaaaggaa tacgcaacta
     2281 tcctggcgaa cgtgtctttc caggatgatt acgggtccct ctctgctaaa gctatccaca
     2341 aaattctgcc tcatcttaaa gaaggaaata gatacgatgt cgcctgcgtg tatgcagggt
     2401 acagacactc tgaatctagc ctgacccgcg aagaaatcgc taataaggtt cttaaagacc
     2461 ggctgatgct gcttcctaaa aactctcttc ataacccagt agtcgaaaag attcttaacc
     2521 agatggtgaa tgttattaat gttatcatcg acatctacgg aaaacccgac gagatcaggg
     2581 tagagctggc tcgcgagctg aagaagaatg ctaaagagcg cgaggaactg acaaagagca
     2641 tcgcccaaac cactaaagca cacgaggagt ataagaccct cttgcagact gagtttgggc
     2701 tcacaaacgt gtctagaacg gacattctta gatacaaact gtacaaggag cttgaatcat
     2761 gcggttataa aaccctctat agcaatacct acatttcacg cgagaagctg ttttccaagg
     2821 agttcgatat tgaacacatt ataccacagg ccagactgtt tgacgactct ttctccaata
     2881 agacactgga agctaggagc gtgaatatcg agaaaggtaa taaaacagct tacgacttcg
     2941 tcaaggagaa gtttggggaa tctggcgccg acaactccct cgaacactat ttgaacaata
     3001 ttgaggacct gttcaaaagt gggaaaatct caaagacaaa gtacaataag ctcaaaatgg
     3061 ctgagcagga tattcccgat gggttcattg agagagatct tagaaacaca caatatatcg
     3121 ccaaaaaagc cctgagtatg ctgaacgaga tcagccacag agtggtcgcc acctctggat
     3181 cagttaccga taagttgcgc gaagactggc agctgatcga tgtcatgaaa gagcttaact
     3241 gggaaaagta taaggccctg ggactggtag agtacttcga ggaccgcgac ggaaggcaga
     3301 ttggaagaat aaaagactgg accaagcgga atgaccatag gcaccacgct atggatgccc
     3361 ttacagtcgc attcacaaaa gacgtgttca tccaatattt caacaacaaa aacgccagtc
     3421 tggacccaaa cgctaatgag cacgctatta agaacaagta tttccagaac ggccgagcca
     3481 ttgccccaat gcccctgcgg gagtttcggg ccgaagctaa gaaacacctc gagaacaccc
     3541 tgatcagtat caaagcaaaa aacaaggtga taaccggcaa cattaacaag acccgcaaaa
     3601 aaggcggggt taataaaaac atgcagcaaa ccccccgggg ccaattgcac ctcgaaacaa
     3661 tctacgggtc tggcaagcag tacctcacca aggaagagaa ggttaatgcg tcattcgaca
     3721 tgaggaaaat aggaactgtg tcaaaatccg cttataggga cgcactgttg aaacggctgt
     3781 acgagaacga taatgaccct aagaaggcct ttgcaggcaa aaattctctc gataagcagc
     3841 caatctggct ggacaaagag caaatgcgga aagtccctga aaaggtgaag attgtgactt
     3901 tggaagctat ctacaccatt aggaaagaaa ttagtcctga cctcaaggta gataaggtaa
     3961 tcgacgtggg tgtccggaag atcctgattg acaggcttaa cgagtacggg aacgacgcta
     4021 agaaggcctt ctccaatttg gataagaacc cgatctggct taataaggag aaggggatta
     4081 gcataaagcg ggtgacaatt agcgggattt ccaacgctca gtccctgcac gtcaagaagg
     4141 ataaggacgg caaaccgatc ttggatgaga atggacggaa tatacctgtc gacttcgtta
     4201 acaccggcaa caaccaccat gtcgctgtct actaccgccc cgttatcgac aaacggggcc
     4261 agcttgtcgt ggacgaggcg ggcaatccga aatatgagct ggaagaggtt gtggtttcct
     4321 tctttgaagc cgtgacccga gccaatctgg gactcccgat tatcgacaag gactacaaga
     4381 ccactgaagg ctggcaattt ttgttcagca tgaagcagaa cgaatacttc gtcttcccaa
     4441 acgagaagac cggcttcaac ccaaaggaaa tcgacctcct cgatgtcgaa aactatggct
     4501 tgatctcccc taacctgttc agagttcaaa agttttcact gaagaattat gtattccggc
     4561 accatctgga gacaacgatt aaggacacta gcagcatact gcgagggatc acctggattg
     4621 actttaggtc cagtaaaggt ttggacacta tcgtcaaggt gcgcgtgaac catatcggac
     4681 agatagtgtc cgtgggagag tacagcaggg ctgaccccaa gaagaagagg aaggtgaggt
     4741 ccatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     4801 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     4861 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     4921 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     4981 ctcactgact cgctgcgctc ggtcgttcgg ctgcgtcttc cgcttcctcg ctcactgact
     5041 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     5101 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     5161 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     5221 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     5281 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     5341 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     5401 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     5461 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     5521 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     5581 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     5641 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     5701 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     5761 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     5821 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     5881 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     5941 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     6001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     6061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     6121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     6181 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     6241 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     6301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     6361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     6421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     6481 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     6541 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     6601 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     6661 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     6721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     6781 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     6841 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6901 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     6961 ccattattat catgacatta acctataaaa ataggcgtat cacgaggccc tttcgtctcg
     7021 cgcgtttcgg tgatgacggt gaaaacctct gacacatgc
//
