LOCUS       sequence-201740-        6093 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 6093)
  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 6093)
  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-C4ZA16
FEATURES             Location/Qualifiers
     source          1..6093
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     67..86
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     280..302
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     294..311
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     295..311
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3708..3728
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3738..3754)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3738..3754)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3751..3773)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3762..3778
                     /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(3786..3792,3793..3810,3811..3816))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3831..3852
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4190..4778)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4270..4289)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4949..5740,5741..5809))
                     /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     5572..5591
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5810..5914)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5982..6000
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6038..6060)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 ggagcagaca agcccgtcag ggcgcgtcag cgggtgttgg cgggtgtcgg ggctggctta
       61 actatgcggc atcagagcag attgtactga gagtgcacca tatgcggtgt gaaataccgc
      121 acagatgcgt aaggagaaaa taccgcatca ggcgccattc gccattcagg ctgcgcaact
      181 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg aaagggggat
      241 gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga cgttgtaaaa
      301 cgacggccag tgtgctgtct catcattttg gcaaagaatt cgcggccgcc accatgaact
      361 acactgagaa ggaaaagctc ttcatgaagt acattttggc tttggacata ggaattgcta
      421 gtgtcggctg ggccatcctt gataaggaat ctgagacagt gattgaagcg ggttctaata
      481 tcttccccga agcttctgct gcagacaatc agctgaggcg agacatgagg ggggccaagc
      541 ggaacaaccg caggctcaaa acccggatca atgacttcat caaactctgg gagaataata
      601 atctttctat tcctcagttt aaatctactg aaatagtggg gctgaaggtg agggcgatca
      661 cagaagaaat caccctcgat gagctctatc tgattctgta ttcttacctg aagcaccggg
      721 ggatttcata tctcgaggat gcgttggacg acactgtctc tggatcttcc gcatacgcca
      781 atgggctcaa actgaatgct aaggagttgg aaactcacta cccctgtgaa atccagcagg
      841 agcgcttgaa cacaatagga aagtatagag gccaaagcca gattatcaat gagaacggcg
      901 aggtgctcga cctgagcaat gttttcacga tcggggctta tcggaaagaa attcagagag
      961 tatttgagat ccagaaaaaa taccatccag agctcacaga tgagttttgt gatgggtaca
     1021 tgctgatctt taacagaaag aggaaatact atgaaggacc tggaaacgag aagtcacgga
     1081 cagactacgg ccgcttcact acgaagctgg acgccaatgg caattatatt actgaagaca
     1141 atatctttga gaaactgatc ggcaaatgtt ccgtttaccc cgacgagctc agggccgccg
     1201 cggccagcta taccgctcag gagtacaacg tccttaatga tctcaacaac ttgacgatta
     1261 acggtcgaaa gcttgaggaa aacgagaagc acgagattgt ggaaagaata aagtccagca
     1321 acacaattaa tatgcgcaag attatatccg attgcatggg cgagaacatt gatgatttcg
     1381 ctggagctcg gattgacaag agcgggaagg aaatcttcca taaatttgaa gtgtacaaca
     1441 agatgaggaa ggcccttttg gagattggca tcgatatcag taactattca cgagaagagc
     1501 tcgatgagat tggctatata atgactatca acaccgacaa agaagccatg atggaagcat
     1561 ttcagaagtc ttggattgat ctgtctgacg atgtgaagca gtgtttgatt aacatgagga
     1621 aaaccaacgg cgccctcttt aataagtggc aatcattcag cctgaagatc atgaacgagc
     1681 tcattcccga gatgtacgcc cagccaaagg aacagatgac tctgctgaca gaaatggggg
     1741 tcaccaaagg aacccaagag gagttcgcag gactgaagta catccctgtc gacgtggtgt
     1801 cagaggacat cttcaatccc gtggtgagga gaagcgtccg gatctctttc aagatcctta
     1861 atgccgttct taaaaaatac aaggctcttg acaccatagt gattgaaatg ccaagagatc
     1921 gcaactcaga ggagcaaaaa aagcgaatca acgattccca gaaactgaac gaaaaggaga
     1981 tggagtatat tgaaaagaaa ctggcagtga cctacggcat caagctgtcc cccagcgatt
     2041 tcagctccca gaagcaactg agcctcaagc tgaagttgtg gaacgagcag gacggaatct
     2101 gcctctattc cggaaaaaca attgatccga acgatattat taataatcct cagctgttcg
     2161 agatagacca tatcattccc agatctatct cattcgacga tgcccgctcc aataaggtcc
     2221 tggtgtacag gtccgagaat caaaagaagg gaaaccaaac tccatactat tatcttacgc
     2281 atagccactc agaatggagc ttcgaacagt acaaggcaac agtcatgaat ctgagtaaaa
     2341 aaaaggagta cgccatttca cggaagaaga tccagaatct tttgtacagc gaggatatca
     2401 ccaagatgga tgtacttaag ggcttcatta accgaaacat caacgataca tcctatgcca
     2461 gtcgccttgt actcaataca atccagaact tcttcatggc taatgaagct gatacgaagg
     2521 tcaaggtgat taaagggtca tatacccacc agatgagatg taacctcaag ctggacaaaa
     2581 accgcgacga gtcctacagt catcacgccg ttgatgccat gctcatcgga tactccgagc
     2641 tgggatatga agcttaccac aagctccagg gtgagtttat cgattttgag acaggtgaaa
     2701 tactccggaa agatatgtgg gacgagaata tgagcgacga ggtgtatgcc gactatttgt
     2761 acggaaagaa gtgggcaaac atacgcaacg aggtggtaaa ggctgagaag aacgtgaagt
     2821 attggcatta cgtgatgcgc aaatccaaca ggggcctctg taatcagaca attcggggaa
     2881 cacgggagta cgatggcaag caatacaaga tcaataagct ggatatcagg accaaggaag
     2941 gtatcaaggt gttcgcgaaa ttggcattta gcaagaaaga ctctgaccga gaaagactgc
     3001 ttgtgtacct gaatgaccgg cggacattcg acgatctttg caagatttat gaagactatt
     3061 ccgacgcagc caacccattt gttcaatacg aaaaggaaac tggagacatc attcggaagt
     3121 actctaagaa gcacaacggc ccccggatcg ataagctgaa gtacaaggat ggcgaggtgg
     3181 gggcctgtat tgacatcagt cataagtacg ggtttgagaa gggcagtaag aaggtgatcc
     3241 ttgagtctct ggtgccctac cgcatggacg tctattacaa ggaagagaat cactcctact
     3301 atcttgttgg agtgaagcaa tccgacatca aattcgaaaa aggaaggaac gttatagacg
     3361 aagaggccta tgctcgaatt ctggtgaatg aaaagatgat ccagccgggc cagagccgag
     3421 ccgacttgga gaacttggga ttcaaattca aactctcatt ctataagaat gacatcatcg
     3481 agtatgagaa agacggcaaa atctatacag aaagacttgt ctcccggacg atgcctaagc
     3541 aacgcaacta tattgaaacc aagcccattg acaaggctaa gttcgaaaag cagaaccttg
     3601 tggggctggg caaaactaag ttcatcaaaa agtatcgata cgacatcctc gggaataagt
     3661 attcttgtag tgaggagaag tttacgagct tctgtagcag ggctgacccc aagaagaaga
     3721 ggaaggtgag gtccatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     3781 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     3841 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     3901 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     3961 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcgtct tccgcttcct
     4021 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     4081 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4141 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4201 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4261 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4321 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4381 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4441 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4501 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4561 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4621 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4681 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4741 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     4801 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4861 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     4921 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     4981 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     5041 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     5101 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5161 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5221 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     5281 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5341 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5401 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5461 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5521 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     5581 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5641 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5701 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5761 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     5821 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     5881 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     5941 acgtctaaga aaccattatt atcatgacat taacctataa aaataggcgt atcacgaggc
     6001 cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg
     6061 agacggtcac agcttgtctg taagcggatg ccg
//
