LOCUS       sequence-201894-        6936 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 6936)
  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 6936)
  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-Q73QW6
FEATURES             Location/Qualifiers
     source          1..6936
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(54..76)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     176..195
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     389..411
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     403..420
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     404..420
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4660..4680
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4690..4706)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4690..4706)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4703..4725)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4714..4730
                     /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(4738..4744,4745..4762,4763..4768))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4783..4804
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5142..5730)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5222..5241)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5901..6692,6693..6761))
                     /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     6524..6543
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6762..6866)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     join(6934..6936,1..16)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga aaacctctga
       61 cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg gagcagacaa
      121 gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa ctatgcggca
      181 tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca cagatgcgta
      241 aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg
      301 cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg tgctgcaagg
      361 cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac gacggccagt
      421 gtgctgtctc atcattttgg caaagaattc gcggccgcca ccatgaagaa ggaaattaaa
      481 gactacttcc tcgggctgga cgtgggtaca ggctctgtgg ggtgggccgt cacagacacc
      541 gactacaaac tgctcaaagc caatagaaag gatctgtggg gaatgcggtg cttcgaaacc
      601 gccgaaaccg ctgaggtcag gaggctccac cgaggcgccc gccgcagaat cgaacgccgg
      661 aagaaacgaa tcaagctcct tcaggagctt ttctcacaag agatcgctaa aactgacgag
      721 ggatttttcc agcggatgaa agagtccccc ttctacgctg aagataagac tatcctgcag
      781 gagaacaccc tgtttaatga taaggatttt gccgataaaa cctatcataa ggcttatcct
      841 accattaatc acctgattaa ggcgtggatc gagaataaag tgaagcccga tccgcgcctg
      901 ctctatctgg catgccacaa tatcatcaaa aagcgcgggc attttctgtt tgagggagac
      961 tttgatagtg agaatcagtt cgacacctct atccaagccc tttttgagta tctgagagag
     1021 gatatggaag tggacattga cgcagatagc cagaaggtga aagagatctt gaaggactca
     1081 agtcttaaga actctgagaa acagtctaga ctgaacaaga tcctgggtct gaagcccagc
     1141 gacaagcaga agaaagccat taccaatctt atctccggga ataaaatcaa ttttgcagat
     1201 ctgtacgaca atcctgatct gaaagatgcc gaaaaaaatt ccatcagctt cagcaaggac
     1261 gatttcgatg cgctcagcga tgacctcgcc agcatcctcg gtgattcatt tgagttgttg
     1321 ctgaaggcaa aggccgtgta caactgttct gttctgtcca aagtcatcgg ggatgaacag
     1381 tatctgagtt tcgcgaaggt aaaaatatac gaaaaacaca agaccgacct gaccaagctt
     1441 aaaaatgtta ttaagaagca cttccctaaa gactacaaga aagtgttcgg atataacaag
     1501 aatgagaaga ataacaataa ctactcagga tacgttggag tatgtaaaac caaaagcaag
     1561 aaactgataa taaataatag tgtgaaccaa gaggactttt ataagttttt gaagaccatt
     1621 ctctcagcaa aaagcgagat caaagaggtc aatgacattc tgaccgaaat cgagacgggt
     1681 acctttctgc cgaagcagat ctcaaaatcc aacgctgaaa tcccatatca actgcggaaa
     1741 atggaactcg agaagatcct gtccaatgct gagaaacatt tcagtttctt gaaacagaag
     1801 gacgagaagg ggttgtctca ctcagaaaag atcattatgc tgctcacgtt taaaatccct
     1861 tattatatcg gacccattaa cgataaccat aaaaaattct ttccggatcg gtgttgggtc
     1921 gtcaaaaagg agaagtctcc gtctggaaaa actaccccct ggaatttctt cgaccacatt
     1981 gataaagaga agacagccga ggcctttata actagtcgca ccaatttttg tacctacttg
     2041 gtaggggaga gtgtacttcc aaaaagctcc ctgctgtact ctgaatacac tgtactgaat
     2101 gagatcaata acctccagat tataatcgac ggcaagaata tctgcgacat taagcttaag
     2161 cagaaaatct acgaggacct gtttaagaaa tacaagaaga tcacccagaa gcagatttcc
     2221 acctttatca agcatgaggg catttgtaac aagacagacg aggtgattat cctgggtatt
     2281 gataaagaat gcactagttc tctgaagtct tacatagaac ttaagaacat atttgggaaa
     2341 caagtcgatg agatatctac aaagaatatg ctggaggaaa tcatccggtg ggccacgatc
     2401 tacgatgaag gggagggaaa gacaattctc aagacaaaaa ttaaagccga gtacggcaaa
     2461 tattgctcag acgaacagat taagaagata ctcaatctga aattctctgg atggggtaga
     2521 cttagccgga agtttttgga gacggtcacc agcgagatgc ctggattttc cgagccagtt
     2581 aacataatca ccgcaatgag agaaacacag aacaacttga tggagcttct ttcctctgag
     2641 ttcacattta cggaaaacat taagaaaata aatagtgggt ttgaggacgc cgaaaagcag
     2701 tttagctacg acggcctcgt taaacccctg tttctcagtc ccagtgtcaa gaagatgttg
     2761 tggcagacac tgaagctggt gaaagagatc tcccacataa cccaggcccc gcccaaaaag
     2821 atctttatcg aaatggcgaa gggcgctgag ctcgaaccag cgcggactaa aacgagactg
     2881 aagattctcc aggatttgta caacaactgt aagaacgacg ccgatgcctt tagttctgag
     2941 atcaaagacc ttagcgggaa aatagagaat gaagacaatc ttaggctcag gagcgacaag
     3001 ttgtacctct actatacgca actcgggaaa tgcatgtact gcggaaagcc catagagatt
     3061 ggtcatgtgt tcgacacttc caattatgac atcgatcata tctatcccca gtccaagatc
     3121 aaagatgact caatctccaa cagagtgctg gtatgctcct cctgtaacaa gaacaaggaa
     3181 gacaagtacc cgttgaaatc cgaaattcag agcaagcagc gcgggttttg gaatttcctc
     3241 cagcggaaca atttcatctc cctggagaaa ttgaatcgcc tcacgcgggc aacccctatt
     3301 tcagatgacg agactgctaa gttcatcgca cgccagctcg tggaaacacg ccaagctacc
     3361 aaagtagccg ctaaggtgct ggagaaaatg ttccccgaga ctaagatagt ttacagtaaa
     3421 gccgagactg tttccatgtt taggaataaa tttgatatcg ttaagtgtag agagataaat
     3481 gatttccacc atgcacacga cgcgtacctg aacattgtgg ttggaaacgt ttacaacacc
     3541 aagttcacaa acaatccatg gaattttata aaagagaaaa gagataaccc caaaattgcc
     3601 gacacataca actactacaa ggttttcgac tacgatgtga agagaaacaa catcacagca
     3661 tgggaaaagg ggaagacgat tatcaccgtg aaggatatgc tgaaaagaaa caccccgatc
     3721 tatacacggc aggccgcctg caagaaaggg gagctgttta accagacgat tatgaaaaag
     3781 ggccttggcc agcaccccct gaaaaaggag ggacctttct caaatatatc aaagtacggg
     3841 ggctataaca aggtttccgc cgcttactac acactcatcg aatacgaaga gaaaggcaac
     3901 aagatcagat ccctcgaaac catccctctg tacctggtga aggacataca gaaagaccag
     3961 gatgtgttga agtcttatct taccgacttg ttgggcaaga aagagttcaa aattctggtt
     4021 ccgaagatca agatcaattc tctgctgaag atcaacgggt tcccgtgcca cattacaggc
     4081 aagaccaacg attcatttct tctccgacct gctgttcagt tttgctgtag caataacgag
     4141 gtgctttact ttaaaaaaat catccgattt tccgaaatcc gaagccaaag ggagaagatc
     4201 ggcaagacaa ttagccctta cgaagatttg tccttcagga gctacatcaa ggagaatctg
     4261 tggaagaaaa ccaagaacga cgagatcgga gaaaaggagt tttacgatct cctgcagaaa
     4321 aaaaacctgg agatatacga catgcttctg acaaaacaca aggataccat ttacaagaag
     4381 cgtccaaact ccgcaactat tgatattctg gttaaaggta aggaaaagtt taaaagcctg
     4441 atcatcgaga atcagtttga ggtgatcctg gagatcctga aactcttttc tgctacccgg
     4501 aacgtgagcg acctccagca cattggcggt tccaagtaca gcggggttgc caagatcggg
     4561 aataaaatct cttccctgga taattgcatt ctgatctacc agagcatcac aggcattttc
     4621 gaaaagagga ttgatctgtt gaaggtgagc agggctgacc ccaagaagaa gaggaaggtg
     4681 aggtccatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     4741 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     4801 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     4861 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     4921 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgt cttccgcttc ctcgctcact
     4981 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     5041 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     5101 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     5161 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     5221 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     5281 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     5341 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     5401 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     5461 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     5521 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     5581 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     5641 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     5701 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     5761 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     5821 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     5881 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     5941 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     6001 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     6061 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     6121 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     6181 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     6241 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     6301 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     6361 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     6421 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     6481 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     6541 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     6601 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     6661 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     6721 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     6781 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     6841 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     6901 gaaaccatta ttatcatgac attaacctat aaaaat
//
