LOCUS       sequence-201812-        6147 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 6147)
  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 6147)
  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-E6WZS9
FEATURES             Location/Qualifiers
     source          1..6147
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     99..118
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     312..334
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     326..343
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     327..343
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3794..3814
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3824..3840)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3824..3840)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3837..3859)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3848..3864
                     /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(3872..3878,3879..3896,3897..3902))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3917..3938
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4276..4864)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4356..4375)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5035..5826,5827..5895))
                     /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     5658..5677
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5896..6000)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6068..6086
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6124..6146)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc
       61 agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact
      121 gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
      181 caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
      241 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
      301 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgtgctgt ctcatcattt
      361 tggcaaagaa ttcgcggccg ccaccatgaa gaaaatcctg ggtgtcgacc tgggcatcac
      421 tagctttgga tacgcgattc ttcaagagac tggaaaggac ctttataggt gtctggacaa
      481 ttctgtagtg atgaggaata atccctatga cgaaaaatca ggtgagtcaa gccagtctat
      541 ccgcagcaca caaaagagca tgagaaggct gatcgaaaag cgcaaaaaaa ggatccgctg
      601 tgtggctcag actatggaac gatacggtat tcttgactac agcgaaacca tgaaaatcaa
      661 tgatccaaag aataatccta taaagaaccg gtggcaactg cgcgccgtgg acgcctggaa
      721 aaggcccctc agccctcagg aactgttcgc aatcttcgcc cacatggcaa aacacagagg
      781 gtacaaatct attgcaacag aagacctcat ctacgagctc gagcttgaac tgggactcaa
      841 cgatcctgaa aaggagtctg agaagaaagc agacgagcgc cgacaggtgt ataacgctct
      901 ccgacacttg gaggaattga ggaaaaagta tggaggagaa acaatcgcac aaacaattca
      961 cagggccgtg gaggcgggag atcttagaag ttacaggaac catgacgact atgaaaagat
     1021 gatcagacgc gaagatatag aagaagagat cgagaaggtg ttgcttaggc aggcggagct
     1081 tggcgctttg ggcttgcccg aggagcaagt ctcagagctc atagatgaac tcaaggcctg
     1141 tatcaccgac caggagatgc ctacaatcga tgagtctctg tttggtaaat gcacgttcta
     1201 caaggatgaa ctggccgccc cagcatactc ttacctgtac gatttgtacc gcttgtataa
     1261 gaaactcgcg gatttgaaca tcgatggtta cgaggttact caggaggaca gggaaaaggt
     1321 gatcgaatgg gtcgagaaga agattgccca gggcaaaaat ctgaagaaaa taacccataa
     1381 ggatctgcgg aaaatcctgg gccttgcccc cgagcagaag atcttcggcg tggaagacga
     1441 gagaattgtc aagggcaaaa aggaaccccg gaccttcgtg cccttctttt ttttggcaga
     1501 cattgcaaag tttaaagagc tcttcgcgag tatccagaaa cacccagacg ctttgcagat
     1561 tttccgcgag ctcgcagaaa ttctccagag gtccaaaact ccacaggagg ccctggatcg
     1621 gctccgggca ctgatggccg gcaagggcat tgatacggac gacagggaac ttctggagct
     1681 gtttaagaac aaaaggtccg ggacaagaga gctctctcat agatatattc tcgaggcctt
     1741 gccgctgttc ctggaaggat acgacgagaa ggaggtgcag cgcattctgg gcttcgatga
     1801 tcgcgaagac tacagccgat atcctaagag cctgcgacat ctgcacttga gggagggaaa
     1861 cctcttcgaa aaagaagaga acccaatcaa taaccacgcc gtaaagagcc tggcctcttg
     1921 ggctctgggc ctgatcgcgg atctctcttg gcggtacggg cctttcgatg aaatcatatt
     1981 ggaaaccacc agagacgcgc tgcccgaaaa aattcggaaa gaaatcgata aggcgatgag
     2041 agagcgcgag aaggctctgg acaaaatcat cggaaagtat aagaaggagt tcccctccat
     2101 cgataaacgc ctggcccgca agatccagct gtgggagcgg cagaaggggc tcgaccttta
     2161 tagcggcaag gtcatcaatt tgagccaact cctggacggc agtgccgata ttgagcacat
     2221 tgtaccccag agcctcgggg ggctctccac tgattacaat actatcgtca ccctcaagtc
     2281 agttaacgcc gcaaagggca accgccttcc aggggattgg ctggccggca accctgacta
     2341 ccgcgagaga attgggatgc tctctgagaa gggacttatt gactggaaaa agagaaagaa
     2401 tcttctggct cagagcctgg acgagattta taccgagaac actcactcaa agggtattcg
     2461 cgccacctcc tacctggaag ctctggtcgc ccaagtgctg aaaaggtatt accccttccc
     2521 cgatccagag ctgcgaaaaa acgggattgg tgttcgaatg attccgggga aagtaacctc
     2581 caaaactagg agccttcttg gcatcaaatc taaaagtaga gagactaact tccaccacgc
     2641 tgaggacgcc ctgatattgt ccactctgac ccgcgggtgg caaaatcgat tgcaccgcat
     2701 gctgcgagat aactatggaa agagtgaggc cgagctgaag gagctgtgga agaagtacat
     2761 gccgcatatc gagggcttga ccctggccga ctacatcgat gaggcgtttc gaaggttcat
     2821 gagcaaggga gaagaaagcc ttttttatcg cgacatgttc gataccataa gatccatctc
     2881 atactgggta gataagaagc cccttagtgc gtcaagccat aaagaaaccg tgtatagcag
     2941 cagacatgag gttcccaccc tgcgaaagaa tattctggaa gcattcgact ctttgaacgt
     3001 gattaaggac cggcacaagt tgaccactga ggaattcatg aaacggtacg acaaagagat
     3061 ccgccagaaa ctctggctgc atcgcattgg taatactaac gacgagagct accgcgccgt
     3121 ggaggagagg gccacccaga tcgctcagat actcaccagg tatcagttga tggatgcaca
     3181 gaacgataag gaaatcgatg aaaagtttca gcaagcactg aaggagctca ttaccagtcc
     3241 tatcgaggtt accggaaaac tgctgaggaa gatgcgattt gtttacgaca aactcaacgc
     3301 gatgcagatt gacaggggtc tggtggaaac agataagaac atgttgggaa tccacatcag
     3361 caaaggtcct aatgagaaat tgatcttccg ccggatggat gtcaacaatg cacacgagct
     3421 gcaaaaggag cgatcaggaa ttttgtgcta cctcaacgag atgctgttca tcttcaataa
     3481 gaagggtctg atacactacg ggtgcctgag gtcttatctc gaaaaggggc agggtagtaa
     3541 gtacattgcc ctctttaacc cacgctttcc agctaatccc aaggctcagc cctctaagtt
     3601 cacgtcagat agcaaaatca aacaggtggg gatcgggagc gctacaggaa tcatcaaggc
     3661 tcacttggac ctcgacggcc acgtgagaag ttatgaagtg ttcgggacac tgcccgaggg
     3721 gtctattgag tggtttaagg aagagagtgg atacggccgg gttgaggatg acccgcatca
     3781 tagcagggct gaccccaaga agaagaggaa ggtgaggtcc atggtcatag ctgtttcctg
     3841 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     3901 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     3961 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4021 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4081 tcgttcggct gcgtcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4141 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4201 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4261 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4321 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4381 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4441 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4501 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     4561 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     4621 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     4681 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     4741 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     4801 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     4861 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     4921 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     4981 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5041 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5101 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     5161 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     5221 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     5281 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     5341 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     5401 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     5461 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     5521 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     5581 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     5641 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     5701 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     5761 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     5821 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     5881 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     5941 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6001 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
     6061 ctataaaaat aggcgtatca cgaggccctt tcgtctcgcg cgtttcggtg atgacggtga
     6121 aaacctctga cacatgcagc tcccgga
//
