LOCUS       sequence-201810-        6201 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 6201)
  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 6201)
  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-E6I4Q5
FEATURES             Location/Qualifiers
     source          1..6201
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(183..771)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(263..282)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(942..1733,1734..1802))
                     /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     1565..1584
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1803..1907)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1975..1993
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2031..2053)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2153..2172
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2366..2388
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2380..2397
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2381..2397
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             5902..5922
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5932..5948)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5932..5948)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5945..5967)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5956..5972
                     /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(5980..5986,5987..6004,6005..6010))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6025..6046
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
       61 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
      121 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
      181 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
      241 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
      301 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
      361 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
      421 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
      481 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
      541 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
      601 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
      661 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
      721 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
      781 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
      841 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
      901 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
      961 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1021 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     1081 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     1141 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     1201 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     1261 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     1321 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     1381 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     1441 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     1501 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     1561 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     1621 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     1681 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     1741 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     1801 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     1861 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     1921 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     1981 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     2041 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     2101 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca
     2161 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     2221 agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     2281 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
     2341 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgtg
     2401 ctgtctcatc attttggcaa agaattcgcg gccgccacca tgtacagtat aggtctggat
     2461 ctggggatct ctagcgtggg ctggtctgtg attgatgagc gaactgggaa cgtcatagat
     2521 ctgggggtga gactcttttc agccaagaat agcgagaaga acctggagcg cagaacgaac
     2581 cgcggcgggc gccgcctgat caggcggaaa accaaccgac tcaaggatgc taaaaagatc
     2641 ctggctgccg tgggcttcta cgaagacaaa tctctgaaga actcctgccc ataccaactc
     2701 cgcgtaaagg gcctgactga gcctttgagc aggggggaga tctataaagt cacattgcac
     2761 attctcaaga agcggggcat ttcctatctg gatgaggtcg atactgaggc cgccaaggag
     2821 tctcaggact ataaggaaca agttcgcaaa aacgcccaac tgcttaccaa gtacaccccc
     2881 ggacagatcc agctgcagag gctgaaagaa aacaataggg ttaaaacagg tatcaatgcc
     2941 cagggaaact atcagttgaa cgtgttcaag gtgtccgcgt atgccaatga gttggccacc
     3001 atcctgaaga cacagcaggc cttttatccc aacgagctga ccgacgattg gattgccctg
     3061 ttcgttcagc ccggcatagc tgaggaggct ggcctgattt accggaaaag accttattat
     3121 cacgggcctg gcaatgaggc aaataatagc ccctacggca ggtggtctga cttccagaaa
     3181 acaggcgaac cggctacaaa catttttgat aagctgatcg gcaaagactt tcagggggag
     3241 ctgagagcta gtggcctgtc cctcagcgcc cagcagtaca acctgttgaa cgacttgacc
     3301 aatctgaaga ttgatggcga ggtccccctc agctccgagc agaaggagta tattctgacc
     3361 gagcttatga ctaaagagtt tacccggttt ggggtcaacg acgtggtgaa gctcctgggc
     3421 gtgaaaaaag agcggcttag cgggtggcgg ctggataaga aaggtaagcc tgaaattcac
     3481 acgctgaaag gttaccgaaa ttggagaaag atttttgccg aagccggcat cgaccttgcg
     3541 acgctgccta ctgagactat cgactgcctt gcgaaagtgc tcacactcaa taccgagaga
     3601 gaaggcattg aaaatacact ggctttcgag ctgccagaac ttagcgagtc cgttaaactg
     3661 ctggtccttg atcgctacaa agaactgagt cagagcatct ccacccagtc atggcaccga
     3721 ttcagtttga agactctgca cctgctgata cccgagctta tgaatgctac gtcagagcag
     3781 aataccctgc tcgagcaatt ccagcttaaa agcgatgtgc gaaaacgcta tagcgagtac
     3841 aaaaagctgc ccacaaaaga tgtgcttgcc gagatttata accccacagt caataagacc
     3901 gtcagccagg ccttcaaagt catagacgcc ctcctggtca agtacggtaa ggagcaaatt
     3961 aggtatatca ccattgaaat gccgcgggat gataacgagg aagacgagaa aaaacgaatc
     4021 aaggagctgc atgccaagaa ttcccaacgg aagaacgata gccagtccta tttcatgcaa
     4081 aaatccggct ggagtcagga gaaatttcaa acgactattc aaaaaaaccg gagatttctc
     4141 gctaagctcc tgtattacta tgaacaggat ggcatatgtg catacaccgg tctgccaatc
     4201 agccctgagc tgctggtcag tgactctacc gaaattgacc atatcatccc aattagtatc
     4261 tccctcgatg acagtattaa caacaaagtg ctcgttctct ccaaggccaa ccaagttaaa
     4321 ggacagcaaa ccccttatga tgcctggatg gatggctctt tcaaaaaaat caacggcaag
     4381 ttctctaatt gggatgatta ccagaagtgg gtggagtcta ggcacttctc tcataagaag
     4441 gagaacaatc tgctggaaac ccggaatatc ttcgattcag agcaggtgga aaagtttctg
     4501 gcgaggaacc tcaatgacac caggtacgcc tctaggctgg tgttgaatac actccaatct
     4561 ttttttacaa atcaggaaac caaggtgcgg gtggtaaacg ggagcttcac acacaccctg
     4621 agaaaaaaat ggggtgccga cctggacaag actagagaga ctcaccacca ccatgctgtt
     4681 gatgcaacac tgtgtgccgt aactagcttc gtgaaggtca gtcgatacca ctacgccgtg
     4741 aaggaagaga caggcgagaa agtgatgaga gagatcgatt tcgaaaccgg tgaaatagtc
     4801 aacgaaatgt cctactggga atttaagaaa tccaagaaat acgagagaaa gacataccag
     4861 gtgaagtggc cgaacttccg ggagcagctg aaacctgtga atctgcaccc ccgcatcaag
     4921 ttctcacacc aagtcgatag gaaagctaac agaaaactgt ccgatgccac catctacagc
     4981 gtccgcgaga aaaccgaagt taagacattg aagagcggga aacagaaaat aaccaccgac
     5041 gagtacacca tcgggaagat aaaagatata tataccctgg acgggtggga ggccttcaag
     5101 aaaaaacaag ataaactgct gatgaaagat cttgacgaaa agacctatga acgccttctg
     5161 tccatcgctg agacgacccc tgactttcag gaggttgagg agaagaacgg aaaagttaag
     5221 agggttaaac gctctccttt cgctgtgtac tgcgaagaga acgatatccc tgccattcaa
     5281 aagtacgcaa aaaagaacaa cggtcccctc attaggtccc tgaagtacta cgatggcaaa
     5341 cttaataaac atatcaacat caccaaggat tcccagggac gccccgtaga aaaaaccaag
     5401 aatgggagga aagtgacact gcaatccttg aagccgtaca gatatgatat ctaccaggac
     5461 ctggagacaa aagcctacta tactgtgcag ctctattata gtgacctccg atttgtggag
     5521 gggaagtatg gcattacaga aaaagaatac atgaaaaaag ttgccgaaca gacaaagggc
     5581 caggtagtcc gcttctgctt ctctcttcag aaaaacgacg gcctcgagat agagtggaag
     5641 gattcacaga gatacgacgt caggttttac aacttccagt ccgccaactc tatcaacttc
     5701 aagggtttgg agcaagagat gatgcctgct gagaatcaat tcaaacagaa accttataat
     5761 aacggtgcca taaatctgaa catcgctaag tatggtaaag agggcaagaa actgagaaag
     5821 ttcaatacag atattctggg aaaaaaacac tacctcttct atgaaaagga acccaaaaat
     5881 attataaaga gcagggctga ccccaagaag aagaggaagg tgaggtccat ggtcatagct
     5941 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     6001 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     6061 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     6121 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     6181 gcgctcggtc gttcggctgc g
//
