LOCUS       sequence-201713-        6255 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 6255)
  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 6255)
  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-D3NT09
FEATURES             Location/Qualifiers
     source          1..6255
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     108..127
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     321..343
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     335..352
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     336..352
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      389..398
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3941..3957)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3941..3957)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3954..3976)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3965..3981
                     /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(3989..3995,3996..4013,4014..4019))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4034..4055
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4393..4981)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4473..4492)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5152..5943,5944..6012))
                     /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     5775..5794
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6013..6117)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6185..6203
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(join(6241..6255,1..8))
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
       61 gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca
      121 gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa
      181 ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
      241 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
      301 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgtgctgtc
      361 tcatcatttt ggcaaagaat tcgcggccgc caccatggca agaccagcgt tccgcgcccc
      421 gaggagggaa catgtcaatg gttggactcc cgaccctcat agaatcagta agcctttctt
      481 catcctggtc agctggcacc tgttgtccag agttgtcatc gacagcagca gcggctgctt
      541 cccggggacc tctagggacc acactgacaa gttcgcagaa tgggaatgcg cagttcagcc
      601 ctataggctc agcttcgacc tcggtacaaa ttctattggc tggggtcttc ttaatctgga
      661 cagacagggc aaacctcgag agattagagc cttggggtcc agaatcttca gcgatggacg
      721 agatccacaa gacaaagcct cattggctgt ggctcggcgg ctggcccggc agatgaggcg
      781 gaggcgggat cgatatctga caagacgaac acgcctgatg ggtgccttgg tccggttcgg
      841 cctcatgcca gcagacccgg ccgcaagaaa aagactcgaa gtggccgtgg acccatattt
      901 ggcaagagaa agagccacac gcgaacggct tgaacccttt gagatcggca gagctttgtt
      961 ccacctgaat cagaggcggg gctacaagcc tgttagaacc gctacaaagc ctgacgagga
     1021 ggctggcaag gtgaaagagg ccgttgagag gctggaagcc gccattgccg ccgccggggc
     1081 gcccaccctc ggggcctggt tcgcttggag aaaaacccga ggcgagacac ttagggccag
     1141 gctggcaggc aaggggaaag aggcagcgta tcccttctac cccgcccgcc gcatgcttga
     1201 agcagagttc gataccctct gggcagagca ggcaagacac catcccgatc tcctgactgc
     1261 tgaggccagg gagatactga ggcaccggat atttcaccaa cgcccgttga agcccccacc
     1321 tgtaggtaga tgcaccttgt acccagacga cgggagagcc cccagagcac tgccttctgc
     1381 acaaaggctg agactgttcc aggagctggc tagtcttcga gtaattcatc tggacctttc
     1441 agagaggcca ctgactcctg ccgaacggga cagaattgta gccttcgtcc aaggccggcc
     1501 accaaaggcc ggtcggaaac caggaaaggt ccagaagagc gtgcctttcg agaagctccg
     1561 aggactgctg gaactgcccc cgggcacagg attcagcctg gagagcgata agcgaccgga
     1621 gctgctcggg gatgaaacag gcgcacgcat cgctcccgct ttcggccctg gctggaccgc
     1681 cctgcccttg gaggagcaag acgcacttgt cgagcttctc ctcaccgaag ccgagcccga
     1741 gagggcaatc gccgccctca ctgcccgctg ggcacttgat gaagctacag ctgccaagct
     1801 tgctggagcc acactccccg atttccacgg acggtacgga cggcgagctg tggctgaact
     1861 gctgccagtc ctggaaaggg aaaccagagg ggaccccgac ggcagagttc gcccaatccg
     1921 actggacgaa gccgtgaagc tgctgcgcgg ggggaaggat cattccgact tcagccgcga
     1981 gggtgcactt ctcgacgctc tcccctatta cggcgctgtg ctggagaggc acgtcgcatt
     2041 tggaactggc aacccggccg atccggagga gaagagagtt ggtcgcgtcg ccaaccccac
     2101 tgttcacatc gcccttaatc agctgaggca tttggtcaac gccattctgg ccaggcatgg
     2161 aaggcctgag gaaatcgtta tcgaacttgc aagggacctg aaaaggtccg cggaggaccg
     2221 gaggcgcgag gacaaacgcc aggctgataa tcagaaaaga aacgaagaac ggaaaaggct
     2281 gattctgtcc ctgggggaac ggccaacccc ccgaaatctg ctcaaactcc gcctgtggga
     2341 agagcagggc cctgtggaga ataggcggtg cccatattcc ggagagacaa tctcaatgcg
     2401 aatgctcctg agcgaacaag tggacatcga tcacatcctt ccctttagcg tgtctctgga
     2461 cgattccgcc gcgaacaagg tcgtttgcct ccgcgaagcc aaccgcatta aacggaaccg
     2521 gagcccttgg gaggcttttg gacacgactc cgaaaggtgg gctggaatcc tggcccgcgc
     2581 tgaagcgctc ccaaaaaata agaggtggag gtttgcaccc gatgctctgg agaagcttga
     2641 gggggaagga ggcttgcgcg ctcggcacct caatgatact cgccatctgt ccaggctggc
     2701 agtggagtac ctgcgctgtg tgtgtcccaa agtgcgcgtc tctcccggga gacttaccgc
     2761 cttgttgagg cgcaggtggg gaatcgatgc tattctcgct gaggcagacg gcccaccccc
     2821 tgaggtgccc gctgaaaccc tcgatccatc tcccgctgaa aagaaccggg cggatcatag
     2881 acatcatgct ctggacgctg tggtgatcgg gtgcattgat agatcaatgg tccagagggt
     2941 tcagctcgct gctgcttccg ctgagagaga ggccgccgca cgcgaggata acatccgcag
     3001 agtgctggaa gggttcaaag aagaaccctg ggacgggttc agggcggaac tcgagcgaag
     3061 ggccaggacg atcgtcgttt cccatcgccc ggagcacggc attggtggtg cgctgcacaa
     3121 agagactgca tatgggcctg ttgatccccc tgaagagggc tttaacctcg tggtgagaaa
     3181 accaattgac ggcctgagta aagatgagat caattccgtc cgcgacccaa ggcttcgcag
     3241 agctttgatc gaccgcctgg ccatccgcag aagggatgcg aacgaccctg ccacagccct
     3301 ggcaaaggcc gcggaagacc tggctgccca acccgcctcc cggggaatta ggagggtgag
     3361 agtgctcaaa aaagagtcta accccatacg ggtggaacac ggggggaatc cttctggtcc
     3421 acggagcggg ggaccctttc acaaactgct gctggctggc gaggtacacc acgtagatgt
     3481 cgcattgcgc gccgatggca ggaggtgggt aggccattgg gtaaccctgt ttgaagctca
     3541 cggaggtcgc ggcgcggatg gtgctgctgc acctccccgg ctcggcgatg gagaacggtt
     3601 tctcatgcgc ttgcataagg gcgactgcct taaattggaa cacaagggaa gagttagagt
     3661 gatgcaggtg gtgaaattgg aaccttcaag caatagcgtg gttgtggtcg agccccacca
     3721 ggtgaaaacc gaccgctcta agcacgtgaa aatcagttgc gatcagctgc gagcccgagg
     3781 agcccggaga gtaactgtcg accccctggg cagagtcaga gtccacgctc ctggagccag
     3841 ggtgggtatc ggcggcgatg caggacgcac cgcaatggag ccagccgaag atatctcaag
     3901 cagggctgac cacaagaaga agaggaaggt gaggtccatg gtcatagctg tttcctgtgt
     3961 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     4021 cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     4081 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     4141 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     4201 ttcggctgcg tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4261 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4321 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4381 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4441 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4501 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4561 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     4621 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     4681 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     4741 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     4801 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     4861 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     4921 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     4981 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5041 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5101 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5161 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5221 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5281 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5341 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     5401 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     5461 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     5521 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     5581 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     5641 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     5701 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     5761 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     5821 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     5881 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     5941 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6001 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6061 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6121 atttccccga aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta
     6181 taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa
     6241 cctctgacac atgca
//
