LOCUS       sequence-201717-        5838 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 5838)
  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 5838)
  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-F4GDP9
FEATURES             Location/Qualifiers
     source          1..5838
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     109..128
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     322..344
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     336..353
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     337..353
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3495..3515
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3525..3541)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3525..3541)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3538..3560)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3549..3565
                     /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(3573..3579,3580..3597,3598..3603))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3618..3639
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3977..4565)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4057..4076)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4736..5527,5528..5596))
                     /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     5359..5378
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5597..5701)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5769..5787
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(join(5825..5838,1..9))
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga caagcccgtc
       61 agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg gcatcagagc
      121 agattgtact gagagtgcac catatgcggt gtgaaatacc gcacagatgc gtaaggagaa
      181 aataccgcat caggcgccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg
      241 tgcgggcctc ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa
      301 gttgggtaac gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgtgctgt
      361 ctcatcattt tggcaaagaa ttcgcggccg ccaccatgag gagcttgcgg tatcgactgg
      421 cactggacct ggggtcaact agcctgggat gggccctctt tcgcctggat gcctgcaaca
      481 ggccaacggc cgttatcaag gccggcgtgc ggatattttc tgacgggaga aaccctaagg
      541 atggctccag tctggccgtg acccgacggg cggctagagc catgcgccga aggcgcgata
      601 ggctgctgaa aaggaagact cgaatgcaag ccaagttggt ggaacatgga ttcttccccg
      661 ccgatgccgg aaaacgaaag gctctggagc agcttaaccc ttacgccttg agagctaagg
      721 ggttgcagga ggccctgctg ccgggagaat tcgcgcgggc actgtttcat atcaaccaga
      781 ggagaggatt taagagcaat aggaagactg acaagaagga caacgattca ggagtgttga
      841 agaaggcaat cggccaattg cgacagcaaa tggctgagca gggttcacgg accgttggcg
      901 aatacctgtg gaccagactg cagcagggcc agggggtcag ggcccgatac agagagaaac
      961 catatacgac cgaagaagga aagaaacgaa ttgataaaag ctacgacctg tacatcgaca
     1021 gagctatgat cgagcaggag tttgacgcac tgtgggctgc ccaggccgcc ttcaatccga
     1081 cactcttcca tgaggccgcg agagcggatc tcaaggatac tctgctccat cagcgaccac
     1141 tcagacccgt aaagccaggc agatgtacac tgctccccga agaagagcgg gcacctctcg
     1201 ctcttcctag cactcagcgg ttcaggatcc accaggaggt taaccacctg aggctcctcg
     1261 atgagaactt gcgagaggtg gccttgactc ttgcacaacg ggacgccgtt gtgacggccc
     1321 tcgaaactaa agctaagctg tccttcgagc agattagaaa gctgctcaag ttgagcggat
     1381 ctgtccaatt taatctggag gatgctaaga ggacagaact gaagggcaac gccacaagtg
     1441 ccgccctggc gagaaaagaa ctgttcggcg ccgcttggtc aggctttgat gaagctctcc
     1501 aagacgagat cgtctggcag ctggtgaccg aggaagggga aggcgcactt atcgcatggc
     1561 tgcagacaca taccggcgtc gatgaggctc gggcccaggc tattgtggac gtatccctgc
     1621 cagagggtta cgggaatctg agtcggaagg ccctggccag gatcgtgcct gccttgcgag
     1681 ccgccgtaat tacctacgat aaggccgtac aggcagctgg gttcgaccat cactcacagc
     1741 tgggcttcga atacgatgcg tcagaggtcg aggatctcgt gcatccagaa accggagaaa
     1801 tccgcagcgt gtttaagcag cttccttact atggaaaagc actccagcgc catgtggctt
     1861 ttgggtctgg gaaacctgaa gaccccgacg agaaacggta tgggaaaatt gcgaacccga
     1921 cagtgcatat tggcctgaac caggtccgca tggtggtgaa cgcgcttatt aggcgctacg
     1981 ggcggcccac tgaagtggtt attgaactgg ctcgagatct gaaacagtcc cgcgagcaga
     2041 aagtggaggc acagagaaga caggccgata accaaaggcg gaacgccagg attcggagaa
     2101 gcattgccga ggtccttggc atcggagagg aacgcgtgcg cgggagcgat attcagaaat
     2161 ggatttgttg ggaggaactg agctttgacg cagccgaccg cagatgccca tattctggag
     2221 tgcagatctc tgcagcaatg ctgctgtccg acgaggtcga agtcgagcac atcttgcctt
     2281 tttccaagac attggatgac tccttgaata acagaacagt ggctatgagg caggcaaaca
     2341 gaataaaaag gaaccgaact ccttgggatg ccagggccga atttgaggcc cagggctggt
     2401 catacgagga cattcttcaa cgcgctgaaa gaatgcccct caggaaacgg tatcgctttg
     2461 cacctgacgg gtatgagcgc tggctcggcg atgacaagga ttttctcgcg cgcgctctga
     2521 atgatacgcg gtacctcagt agagttgccg ccgaatacct gcgacttgtg tgtccaggaa
     2581 caagggtaat cccggggcag ttgaccgctc tgcttcgcgg caagttcgga ctgaatgacg
     2641 ttttggggct ggacggtgag aagaatcgaa acgatcaccg ccaccacgct gtagacgctt
     2701 gtgttattgg cgtgaccgac cagggactga tgcagcgctt cgctactgcc tctgcccaag
     2761 ctagggggga tggtctgacc cgcctggtgg acggtatgcc catgccatgg ccaacatata
     2821 gagatcacgt tgagcgagcc gtccgacaca tatgggtgag ccatcggcct gatcacggtt
     2881 tcgaaggagc tatgatggag gagacaagtt acggaatacg gaaagatggt agcatcaaac
     2941 agcgaaggaa ggccgatgga tcagcgggcc gcgagattag caatctgatt aggattcacg
     3001 aagcaacaca gcctctgcga cacggagtga gtgctgatgg ccaacctctc gcctacaagg
     3061 gatatgtagg gggtagtaat tattgcatcg aaataactgt taacgacaag ggtaagtggg
     3121 agggcgaagt gatcagtacc ttcagggctt atggcgtcgt ccgagcaggt ggcatgggta
     3181 ggcttcggaa tccgcacgag ggccagaacg gcaggaagct cattatgcga cttgttatag
     3241 gggactcagt ccgcctcgag gtcgatggtg ctgaacgcac tatgcggatc gtgaagattt
     3301 ccggaagcaa tggccagatt tttatggctc ccatccacga agctaatgta gatgcacgga
     3361 acacagataa acaggatgct ttcacctaca catctaaata cgccggaagt cttcagaagg
     3421 caaaaacaag gcgcgtgacg atttccccca tcggtgaagt tcgggatccg ggatttaagg
     3481 gcagcagggc tgaccccaag aagaagagga aggtgaggtc catggtcata gctgtttcct
     3541 gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt
     3601 aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc
     3661 gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg
     3721 agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg
     3781 gtcgttcggc tgcgtcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     3841 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     3901 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     3961 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4021 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4081 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4141 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4201 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4261 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4321 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4381 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     4441 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4501 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4561 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4621 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4681 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4741 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     4801 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     4861 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     4921 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     4981 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5041 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5101 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5161 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5221 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5281 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5341 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5401 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5461 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5521 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5581 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5641 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5701 gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc atgacattaa
     5761 cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt gatgacggtg
     5821 aaaacctctg acacatgc
//
