LOCUS       sequence-201804-        6405 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 6405)
  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 6405)
  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-D1VXP4
FEATURES             Location/Qualifiers
     source          1..6405
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(6..28)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     128..147
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     341..363
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     355..372
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     356..372
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4081..4101
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4124..4146)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4135..4151
                     /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(4159..4165,4166..4183,4184..4189))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4204..4225
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4563..5151)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4643..4662)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5322..6113,6114..6182))
                     /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     5945..5964
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6183..6287)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6355..6373
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc
       61 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt
      121 aactatgcgg catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg
      181 cacagatgcg taaggagaaa ataccgcatc aggcgccatt cgccattcag gctgcgcaac
      241 tgttgggaag ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga
      301 tgtgctgcaa ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa
      361 acgacggcca gtgtgctgtc tcatcatttt ggcaaagaat tcgcggccgc caccatgaac
      421 aaaagaattc ttggcctgga caccggcacc aattcactcg ggtgggcagt cgtggactgg
      481 gatgagcacg ctcagtctta cgaactcatc aagtatggag atgtgatctt ccaggaaggg
      541 gtcaagatcg agaaggggat agaatcttct aaggctgccg aaaggagcgg gtacaaggca
      601 attcggaaac agtatttcag gcgacgactc agaaaaatac aagtgctcaa ggttctggta
      661 aagtaccatt tgtgtccata tctctccgac gatgacctcc ggcaatggca tctccagaag
      721 cagtacccca aatcagatga actcatgctg tggcagagaa cgtcagatga ggaagggaag
      781 aatccatatt atgatcgcca cagatgtctg catgagaagc tggatctgac tgttgaagcg
      841 gaccgctata cccttggccg cgccctgtac cacctgacgc agaggcgcgg gttcctttca
      901 aatcggctgg acacatccgc cgataataag gaggatggtg tggtgaagtc cggtatctcc
      961 cagttgtcta ccgaaatgga ggaggcagga tgtgagtact tgggggatta cttttacaaa
     1021 ctttatgacg ctcagggcaa taaagtgagg attcggcaaa gatacacaga taggaacaaa
     1081 cattatcagc atgagttcga tgcgatctgt gaaaagcaag aactgagcag tgagcttatt
     1141 gaggatctgc aacgggccat cttcttccaa ctcccattga aaagtcagcg ccacggggtg
     1201 ggacggtgca ccttcgaaag ggggaaacca agatgtgcag actctcaccc agattacgaa
     1261 gaatttcgga tgttgtgctt tgtgaataac atccaggtaa aagggccgca cgaccttgag
     1321 ctgcggccgt tgacttacga ggaacgggaa aaaattgaac ccttgttctt ccgcaaatca
     1381 aagccaaact ttgatttcga agacattgcc aaggcacttg ccggtaaaaa gaactatgcg
     1441 tggatccacg ataaggaaga aagagcctac aaatttaact acaggatgac tcaaggagtg
     1501 cctgggtgcc ccactattgc acagctgaaa tccatcttcg gagatgactg gaagactggg
     1561 attgcggaga catataccct gatccagaaa aaaaatggca gcaagagcct gcaagaaatg
     1621 gtggatgacg tgtggaacgt tctctactcc ttttctagtg ttgagaagct gaaggagttc
     1681 gcgcatcata aactgcagct cgacgaagaa tcagctgaga aattcgcaaa aatcaagctc
     1741 tctcattcat ttgctgcgct cagcctgaag gccatccgaa agtttctccc ctttttgcgc
     1801 aagggaatgt actacacaca tgcctcattc ttcgctaaca tcccaaccat cgtaggcaag
     1861 gaaatatgga acaaggagca gaaccggaag tacatcatgg aaaatgtggg cgagctcgtt
     1921 ttcaactatc agcctaaaca ccgggaggtc cagggtacca tcgaaatgct cataaaagat
     1981 tttctcgcta acaattttga gctgccagct ggcgcaactg acaaactgta tcacccctct
     2041 atgatagaaa cctaccccaa cgcacagcgc aatgagttcg ggatcctcca gctcggcagc
     2101 ccccgcacca atgctataag aaatcccatg gctatgagga gcctccacat cctgcgaaga
     2161 gtggtgaatc agctgctgaa agagagcatc atcgacgaaa atacggaggt gcacgtagag
     2221 tacgccaggg aactcaatga cgctaacaag cggagggcaa ttgctgacag acagaaagaa
     2281 caggataaac aacacaaaaa gtacggagac gaaattagga agctttacaa ggaagaaacc
     2341 ggcaaggaca ttgaaccaac ccagacagat gttctgaaat tccagttgtg ggaggaacag
     2401 aatcaccatt gtctgtacac aggcgagcag ataggcatca cagattttat agggtctaat
     2461 cctaaattcg acatcgagca taccatacca cagtccgtcg gaggtgatag tactcagatg
     2521 aatttgactc tgtgtgacaa ccggttcaat agagaagtga aaaaggctaa actgcccacc
     2581 gaacttgcca atcacgagga gattctcacc cggatcgaac cttggaaaaa taaatacgaa
     2641 caattggtga aagaacgaga caagcagcgg acctttgcag gcatggacaa ggcagttaag
     2701 gatattcgca ttcagaagcg gcacaagctc caaatggaga tcgattattg gcgcggtaag
     2761 tatgaaagat ttacgatgac cgaggtgcca gagggtttca gtagaagaca aggaactgga
     2821 ataggtctta tttcaaggta cgccggactc tacctcaaga gtcttttcca ccaagctgac
     2881 tctaggaaca agtctaacgt gtatgttgtc aagggcgtcg cgacggcaga gttccggaag
     2941 atgtggggcc tgcaatctga gtatgaaaaa aagtgtcggg acaatcactc acaccactgt
     3001 atggatgcta tcaccatcgc ttgcatcggt aagcgcgaat acgacctgat ggccgagtac
     3061 tatcggatgg aggagacatt taaacagggt cgaggttcta aaccaaagtt tagcaaaccg
     3121 tgggccacct tcacagagga cgtcctcaac atatacaaga accttctggt tgtgcacgat
     3181 acgcctaaca atatgccgaa acacacgaag aaatacgtcc agacatctat cggaaaggtc
     3241 cttgcccagg gagacacagc ccgcggctcc ttgcatctcg atacttatta cggcgctata
     3301 gagagagacg gcgagatacg ctatgtcgtt cgccgacctc tgagtagttt tactaagcct
     3361 gaagaattgg agaatattgt agacgaaacc gtaaagagga ctataaaaga ggctatcgcc
     3421 gacaaaaatt tcaagcaagc aattgcagaa cccatctaca tgaacgaaga gaagggtatt
     3481 ctgataaaga aagtgcgatg ctttgccaaa agtgtgaagc agcctatcaa catccgccag
     3541 cacagggacc tctccaagaa agaatataaa cagcaatacc acgttatgaa cgaaaataat
     3601 tatctgttgg ctatttacga aggtctggtg aagaataaag ttgtgcgaga gttcgaaatc
     3661 gtgtcttaca tcgaggccgc taagtactac aagcggtcac aggatcggaa catttttagt
     3721 tctatcgtgc caactcatag caccaaatac ggcctccccc tgaagacgaa gctgctgatg
     3781 gggcagcttg tgcttatgtt tgaggaaaac ccggacgaga tccaagtaga caatacaaaa
     3841 gacctggtca aacgattgta taaagtggtt gggatcgaga aagacggccg gattaaattt
     3901 aaatatcacc aagaggctag gaaggaaggg ctgcctatct tctctactcc atataagaat
     3961 aacgatgatt acgcccctat cttccgacag tcaatcaata acataaatat tctggtcgac
     4021 ggaatcgact tcactattga tattctgggc aaggtgactt tgaaagagag cagggctgac
     4081 cccaagaaga agaggaaggt gaggtccatg ttcatagctg tttcctgtgt gaaattgtta
     4141 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4201 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4261 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4321 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4381 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4441 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4501 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     4561 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     4621 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4681 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4741 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4801 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4861 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4921 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4981 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     5041 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5101 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5161 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5221 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5281 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5341 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5401 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5461 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     5521 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     5581 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     5641 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5701 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5761 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5821 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5881 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5941 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6001 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6061 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     6121 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6181 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     6241 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6301 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     6361 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggt
//
