LOCUS       sequence-209043-        6039 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 6039)
  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 6039)
  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-A8REH9
FEATURES             Location/Qualifiers
     source          1..6039
                     /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             3547..3558
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             5740..5760
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5770..5786)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5770..5786)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5783..5805)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5794..5810
                     /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(5818..5824,5825..5842,5843..5848))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5863..5884
                     /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 tgatggaggt gtttatgggc
     2461 agattggtcc tcggtcttga cattggtatt acaagtgtag gtttcggcat cattgacctg
     2521 gacgaatctg agatagttga ctacggagtg aggctgttca aggaaggaac tgctgccgaa
     2581 aatgaaaccc gacgcaccaa aagaggaggc aggcgactga aaagacgccg cgttaccaga
     2641 agagaggaca tgctccacct gctgaaacag gcaggaatta taagtacaag cttccatcct
     2701 ttgaataacc cttatgatgt tcgggtgaag ggtttgaatg aaagactgaa cggagaggaa
     2761 ctggcaactg ctctgctcca tctctgcaag cacaggggca gctcagtcga aaccattgaa
     2821 gacgacgaag ctaaggcaaa ggaagccggg gaaactaaga aagtgttgtc aatgaatgat
     2881 cagctgctga aaagcggaaa gtatgtgtgc gaaatccaga aagaaagact gcggacgaat
     2941 ggtcatatca gagggcacga aaacaatttc aagactagag cttatgttga cgaggcattc
     3001 cagattcttt cacaccagga tctgtcaaat gaactgaaga gcgctatcat taccatcatc
     3061 tcacgaaaga ggatgtacta tgatgggcca ggcggccctc tgtctcccac gccctacggc
     3121 aggtacacat actttggcca gaaggaacca atagatctca tcgaaaagat gagaggtaag
     3181 tgctcacttt tccctaacga gccccgagcc cctaaactgg cttactctgc tgagctgttt
     3241 aatctgttga acgacctcaa caatttgtct atagagggag agaaacttac gagcgaacag
     3301 aaagcaatga tcttgaaaat cgtccacgag aagggcaaga ttacacctaa acagctggcc
     3361 aaggaggtgg gcgtttcact ggagcagata aggggcttca ggatcgacac taagggaagt
     3421 cccctgctgt ccgaactcac gggatataag atgatccgag aggtccttga aaagagcaat
     3481 gacgaacacc tggaggatca tgtgttttat gacgagatcg cggagatctt gacaaagacc
     3541 aaagacattg agggcagaaa gaagcagatt agcgagcttt ctagcgacct caacgaagaa
     3601 agtgtgcatc agctggccgg gcttacaaaa tttaccgctt accattcact tagcttcaag
     3661 gcactgcgcc tgatcaacga agaaatgctt aaaacagaac tcaatcagat gcaatctatt
     3721 acgctctttg ggctgaagca gaataatgag ctgtccgtga aaggcatgaa gaacattcaa
     3781 gccgacgata cagcaatcct ttctccggtc gccaaacgag ctcagcgcga aacctttaag
     3841 gtggtgaacc gcctgagaga gatttatggg gagttcgatt ctattgtagt ggagatggcc
     3901 agggagaaga actctgaaga gcagcgcaag gcgatcaggg aaagacaaaa gttcttcgaa
     3961 atgcgcaaca aacaggttgc agatatcatc ggagacgatc gcaagattaa cgcaaagctt
     4021 cgcgaaaagc tggtgctcta ccaggaacag gacggcaaaa cggcgtactc cctcgagccc
     4081 attgatttga aactgctcat cgacgatcca aacgcctatg aagtagatca tataattcct
     4141 atcagcatct ccctggacga ttctataact aataaggtgt tggttaccca tcgggagaac
     4201 caggagaagg gaaacctcac gcccatctct gcctttgtta aaggacgctt cactaagggg
     4261 tcccttgcgc aatacaaagc ctactgcctg aaactgaagg agaagaatat aaagactaat
     4321 aagggctaca gaaagaaagt tgaacagtac ctgctcaacg aaaatgacat ctataagtat
     4381 gatattcaga aagagttcat caaccggaat ctcgtcgata caagctatgc atctagggtg
     4441 gttctgaaca cactgaccac ctacttcaaa cagaatgaaa ttcctaccaa agtgtttaca
     4501 gtgaaaggat cactgacaaa tgcttttcgc cgaaagatta atctgaagaa agatcgagat
     4561 gaggattatg gccatcatgc cattgacgcg ctgattattg catccatgcc aaagatgagg
     4621 ctgctgagta caattttctc aaggtataag atcgaggaca tctacgatga atctacaggg
     4681 gaggtgttta gctcaggcga tgacagcatg tattatgacg accgatattt cgcttttatc
     4741 gcttcactga aggctatcaa agtcaggaaa ttttcacata agatcgacac caagcctaat
     4801 cggagtgttg ctgacgagac tatctacagt accagagtta tcgacggcaa ggaaaaggtt
     4861 gtgaagaagt ataaggacat ttacgaccca aagtttactg ctctggcaga agatatcttg
     4921 aataacgcct atcaggagaa gtatctcatg gctttgcacg atccacaaac cttcgatcaa
     4981 atagtgaagg tcgtgaatta ttacttcgag gaaatgtcca aaagtgaaaa gtatttcacg
     5041 aaagataaga agggccgaat caaaatttca gggatgaacc ctctgagcct ttaccgggat
     5101 gagcatggta tgttgaagaa gtacagcaag aaaggcgacg gccctgcgat aacccagatg
     5161 aagtattttg acggggttct cgggaaccac atagatatta gcgcgcacta tcaggtgagg
     5221 gataagaaag ttgttcttca gcaaatctct ccatacagga cggactttta ctattccaaa
     5281 gaaaacggct acaagttcgt aactatcagg tataaggatg tcaggtggtc cgagaagaag
     5341 aagaagtacg ttatcgacca acaggattat gccatgaaga aagccgagaa gaagattgat
     5401 gacacctacg agtttcagtt cagcatgcat cgcgatgagc tgattggcat caccaaggcc
     5461 gaaggcgaag ccctcatcta ccccgatgaa acgtggcata atttcaattt cttctttcat
     5521 gccggcgaga ctcccgaaat tctcaagttt acggccacca acaacgacaa gtctaacaag
     5581 atagaggtca agcctatcca ctgctattgt aaaatgaggc tgatgccaac gatctctaag
     5641 aagatagtga gaattgacaa gtatgcaact gacgtcgtcg ggaatctcta taaagtcaag
     5701 aagaataccc ttaagttcga gtttgatagc agggctgacc ccaagaagaa gaggaaggtg
     5761 aggtccatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5821 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5881 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     5941 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     6001 ctcgctcact gactcgctgc gctcggtcgt tcggctgcg
//
