LOCUS       sequence-201890-        6867 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 6867)
  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 6867)
  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-Q5X8T6
FEATURES             Location/Qualifiers
     source          1..6867
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     70..89
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     283..305
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     297..314
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     298..314
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      351..360
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4485..4505
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4515..4531)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4515..4531)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4528..4550)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4539..4555
                     /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(4563..4569,4570..4587,4588..4593))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4608..4629
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4967..5555)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5047..5066)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5726..6517,6518..6586))
                     /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     6349..6368
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6587..6691)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6759..6777
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6815..6837)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc
       61 ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatatgcgg tgtgaaatac
      121 cgcacagatg cgtaaggaga aaataccgca tcaggcgcca ttcgccattc aggctgcgca
      181 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg gcgaaagggg
      241 gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca cgacgttgta
      301 aaacgacggc cagtgtgctg tctcatcatt ttggcaaaga attcgcggcc gccaccatgg
      361 agtctagtca aattctttct ccgattggaa ttgatctggg gggtaaattc acaggcgtct
      421 gtctgagcca cctcgaggcg ttcgctgagc tgcccaatca cgcaaatacg aaatattccg
      481 tgattctgat tgaccataat aacttccaac tgtctcaggc acagcgccgc gcaacgagac
      541 atagggtaag aaataaaaaa aggaatcaat ttgtaaagcg agttgcactg cagctgttcc
      601 aacacattct ttccagagat ctgaatgcaa aggaggaaac cgccctgtgc cactatttga
      661 acaaccgagg gtatacttac gtggacaccg acctggacga atacatcaag gatgaaacta
      721 caattaacct tctcaaagag ctcctgcctt ctgagtccga acacaatttt attgactggt
      781 tcctgcagaa gatgcagagt tccgagttca ggaagatcct ggtgagtaaa gtcgaggaga
      841 agaaggatga taaagagctg aaaaatgctg ttaaaaacat taaaaatttt atcaccggat
      901 ttgaaaagaa ttctgtcgag gggcaccggc ataggaaagt ttactttgaa aacattaaat
      961 ccgatatcac caaggataac cagctggatt ccattaaaaa gaaaatacct agtgtctgcc
     1021 tgtccaatct tctcggccac ctctctaatc tgcagtggaa gaatctgcac aggtacctgg
     1081 ctaaaaaccc caaacagttc gacgagcaga ccttcggtaa cgaatttttg cgcatgttga
     1141 agaacttcag acatttgaag gggtcccagg aatcactcgc cgtgagaaat ctgatccagc
     1201 agctggagca gtcacaggat tacatatcaa ttcttgagaa gaccccccca gaaatcacaa
     1261 tcccccctta tgaagcccgc acaaatacgg gaatggagaa ggatcagagt ctgttgttga
     1321 atcctgaaaa gctgaacaac ctttacccca actggcgcaa cctgatccct gggattatcg
     1381 atgcacaccc atttctggaa aaagacttgg aacacacgaa gctccgagat cgcaagcgaa
     1441 tcatcagtcc cagcaaacag gacgaaaagc gggacagcta cattctgcag aggtacctcg
     1501 acctgaataa aaagattgat aagttcaaaa taaagaagca gttgagcttt ctggggcagg
     1561 gcaaacagct tcccgccaat ctcatcgaaa cgcagaagga gatggaaacc cattttaaca
     1621 gctcactggt gagcgtgttg atacagattg catcagctta taataaagaa agggaggacg
     1681 cagcccaggg gatatggttt gataacgcct tctccctctg tgaactgtcc aacattaatc
     1741 ctccccggaa gcagaagatt ctgcccctgc tggtgggagc catactgagt gaagacttta
     1801 tcaataacaa agacaagtgg gctaaattca aaatcttctg gaacactcat aaaatcggtc
     1861 gcacctccct taagagcaag tgcaaagaaa tcgaggaagc tcggaagaac agtggtaatg
     1921 cgtttaagat cgactatgag gaggccttga accaccctga acactcaaac aataaagccc
     1981 ttatcaagat tatccagact atcccagaca taatccaggc tatccaatcc cacctggggc
     2041 acaatgattc tcaagcactg atttaccata atcccttttc actgagtcaa ctgtatacta
     2101 tccttgagac taagcgagat gggttccaca agaactgcgt ggccgtgacg tgtgagaatt
     2161 actggcggag tcagaagacc gaaatcgacc cggaaatctc ttacgcctca aggcttccag
     2221 ctgatagcgt gcgccctttc gacggcgttc tggctcggat gatgcagcgg cttgcctacg
     2281 agatagcgat ggccaaatgg gaacaaatca aacacattcc agacaattca agtctgctta
     2341 tccccatcta tctggaacag aaccggttcg agtttgagga gtcgtttaag aaaatcaagg
     2401 gctcttctag cgacaaaact ctggaacagg ctatcgagaa gcagaacatt cagtgggagg
     2461 agaaattcca gagaatcatt aatgccagta tgaatatctg cccctataag ggggcaagca
     2521 tcggcgggca gggggaaatt gatcatatat atcctagaag cttgagtaag aaacactttg
     2581 gagtgatctt taacagcgag gttaacttga tctattgtag tagtcagggg aatagagaga
     2641 agaaagagga gcattacctg ctcgaacatc ttagccctct gtacctcaag caccagttcg
     2701 gaaccgataa cgtctctgat attaaaaact ttatctccca gaatgttgca aatatcaaga
     2761 agtacatctc cttccacctt ctgacccctg agcagcagaa ggcagccaga catgccctgt
     2821 tcctggatta tgacgatgag gccttcaaga ccattacgaa attcttgatg agtcaacaga
     2881 aagctcgagt gaatggtact cagaagtttc tggggaagca aatcatggaa ttcttgtcca
     2941 ccctggcgga ttccaagcag ctgcagctgg agttcagtat aaaacagatc acagctgagg
     3001 aggttcacga ccatagggag ctgctgtcca agcaggaacc caaactcgtg aagtcacggc
     3061 agcagagctt ccctagccac gccattgatg ccacgttgac aatgagcatc ggactcaagg
     3121 agttccctca attttcacaa gaactcgata atagctggtt cattaaccac ctgatgcctg
     3181 acgaggtcca cctgaaccca gtgcgctcca aggaaaaata caataaacct aatatcagta
     3241 gcaccccact cttcaaggac agtctctacg ctgagcgctt cattcccgtg tgggtcaaag
     3301 gggaaacctt tgctatcgga ttcagcgaaa aggatctgtt tgaaatcaag ccctccaaca
     3361 aggagaagct gttcacgctg ctgaagacat acagcacaaa gaatcctggg gagtcacttc
     3421 aggagctgca ggcaaagtcc aaagcgaaat ggctgtattt ccccataaat aaaaccctgg
     3481 ccctggagtt tctgcatcat tattttcaca aggagatcgt gacccctgat gacaccacgg
     3541 tgtgtcactt tatcaactct ctgcggtatt acaccaagaa ggagagtatt acggtgaaga
     3601 tattgaaaga gcccatgccc gtgctgtccg ttaagttcga gtcctcaaag aagaatgtgc
     3661 tcgggtcatt caagcataca atcgccctgc ccgcgacaaa ggactgggag cgattgttta
     3721 atcaccccaa ctttttggcc ctcaaggcaa accccgcccc taatcccaaa gaatttaacg
     3781 aattcattcg gaagtatttc ctttcagaca ataatccaaa cagcgatatt ccaaataatg
     3841 ggcacaacat caaaccccag aaacacaaag cagttagaaa ggtgttttcc ttgcccgtca
     3901 tacccggcaa cgctggtacc atgatgagga ttcgccggaa ggataacaag ggccaacctc
     3961 tttaccagct ccagacaatc gacgacacgc caagcatggg aatccagatc aacgaagatc
     4021 ggctggtgaa acaggaagtg cttatggacg catataagac gcggaatctc tctaccattg
     4081 acggcataaa caacagcgag ggacaagctt acgcaacctt tgataactgg ctgactttgc
     4141 ccgtgtccac ttttaagcct gagataatca agctggaaat gaaaccacac tccaagaccc
     4201 gcagatacat ccggattact cagtcactgg ccgatttcat caaaacaatt gacgaagcgc
     4261 tgatgatcaa gccctccgac agtattgatg atcctctcaa catgcctaac gaaatagttt
     4321 gcaagaacaa gctgtttggt aacgaactga agccacgcga cgggaagatg aagattgtgt
     4381 ccacaggcaa aatagtgact tacgagtttg aatcagacag cacacctcag tggattcaga
     4441 cgttgtacgt gacgcagctg aagaaacagc caagcagggc tgaccccaag aagaagagga
     4501 aggtgaggtc catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     4561 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     4621 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     4681 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     4741 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcgtcttcc gcttcctcgc
     4801 tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg
     4861 cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag
     4921 gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc
     4981 gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag
     5041 gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga
     5101 ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc
     5161 atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg
     5221 tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt
     5281 ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca
     5341 gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca
     5401 ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag
     5461 ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca
     5521 agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg
     5581 ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa
     5641 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5701 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5761 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5821 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac
     5881 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5941 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     6001 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6061 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6121 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6181 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6241 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6301 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6361 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6421 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6481 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6541 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6601 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6661 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
     6721 tctaagaaac cattattatc atgacattaa cctataaaaa taggcgtatc acgaggccct
     6781 ttcgtctcgc gcgtttcggt gatgacggtg aaaacctctg acacatgcag ctcccggaga
     6841 cggtcacagc ttgtctgtaa gcggatg
//
