LOCUS       sequence-201719-        6090 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 6090)
  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 6090)
  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-K2FTY8
FEATURES             Location/Qualifiers
     source          1..6090
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     81..100
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     294..316
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     308..325
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     309..325
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3719..3739
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3749..3765)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3749..3765)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3762..3784)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3773..3789
                     /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(3797..3803,3804..3821,3822..3827))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3842..3863
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4201..4789)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4281..4300)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4960..5751,5752..5820))
                     /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     5583..5602
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5821..5925)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5993..6011
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6049..6071)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg
       61 tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatgcg
      121 gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc attcgccatt
      181 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
      241 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
      301 acgacgttgt aaaacgacgg ccagtgtgct gtctcatcat tttggcaaag aattcgcggc
      361 cgccaccatg cggtaccgcg taggcctgga tctgggaacc gcgtcagtgg gagctgccgt
      421 gttcagtatg gatgagcagg gtaaccccat ggagcttatc tggcactatg aacgcctttt
      481 ctctgaaccc ttggtccctg atatgggcca gcttaaaccc aagaaagccg cccgccgatt
      541 ggcgaggcag cagagaaggc agattgatcg aagggctagt cgactgcggc gaatagctat
      601 cgtttctcgc aggttgggga tagccccagg gagaaatgac tctggagtgc acggaaacga
      661 cgtccccact ctccgcgcca tggctgtgaa tgaacgaatc gagctggggc aattgcgcgc
      721 tgtgcttctc cgaatgggca aaaagcgggg gtacggagga acatttaaag ccgtacgcaa
      781 agttggagag gctggcgagg tggcctcagg agcctcacgc ctggaggagg aaatggtggc
      841 actggcgagt gtgcagaata aggacagcgt aacagttggc gaatatttgg ctgcacgggt
      901 agagcatgga ctgcctagta aattgaaagt ggccgcgaac aacgaatact acgccccgga
      961 atacgctctt ttcaggcagt acttggggct ccctgcgatt aagggccgcc cagactgtct
     1021 gcccaatatg tacgccctgc gccaccagat tgagcacgag tttgaacgaa tctgggccac
     1081 ccagtctcaa tttcatgacg tgatgaaaga ccatggcgtc aaggaagaaa ttaggaacgc
     1141 gatatttttc cagcgacccc tcaagtctcc agccgataag gtgggacgat gttcactgca
     1201 gacgaacctg ccacgcgccc caagggccca aatcgcagcc cagaacttcc gcatcgagaa
     1261 gcagatggcc gatctgcggt ggggcatggg ccgcagagcc gagatgctta acgaccacca
     1321 aaaggctgtg atcagagagc tcttgaacca gcagaaagag ctgagtttcc gaaaaatcta
     1381 caaggaattg gaaagggctg gttgccctgg ccccgaaggg aagggcctga acatggacag
     1441 ggctgcactc gggggaagag atgacctgtc cggcaacaca actctcgccg catggaggaa
     1501 acttggcctt gaggatcgct ggcaggaact ggatgaagtg acacagatcc aggtcattaa
     1561 tttcctcgcc gaccttggct cccccgagca gcttgatacc gatgactgga gttgcaggtt
     1621 tatggggaag aatgggcggc ccaggaattt ttcagatgag tttgtggcat ttatgaatga
     1681 gttgaggatg acagacggct tcgacaggct ttctaagatg ggattcgaag ggggcaggtc
     1741 ttcctactca atcaaagccc ttaaagctct cactgagtgg atgatagctc ctcactggcg
     1801 ggagacacca gagacgcacc gggttgacga agaggcagct atcagggaat gctacccaga
     1861 gagcttggca accccagcac aggggggccg acagtctaag ctcgagcctc cacccctgac
     1921 tggaaatgaa gtagtggacg tcgctctgag acaagttagg cataccatca atatgatgat
     1981 cgacgacctg ggttccgtgc ctgcccagat cgtggtggag atggccaggg agatgaaggg
     2041 gggcgtcacc cggaggaacg acatcgagaa gcagaataag cgatttgctt ccgaacgaaa
     2101 aaaagccgcc cagtccatcg aggagaacgg caagactcca acaccagccc gcatcctccg
     2161 ataccagctg tggattgaac aaggtcacca gtgcccatat tgcgagtcaa atataagcct
     2221 cgagcaggca ctgtccggcg cttatacaaa tttcgaacat atcctgccaa ggacgctcac
     2281 ccaaattgga cgcaaaagat ccgagcttgt tctcgctcac agagagtgta atgacgaaaa
     2341 gggcaaccga actccctatc aggcttttgg acacgacgat aggaggtggc gcatagtcga
     2401 gcaacgggct aacgcattgc ccaagaaatc atctaggaag acccggctgt tgctcctgaa
     2461 agactttgag ggggaggccc tgaccgacga gtccattgac gaatttgccg acaggcagct
     2521 gcatgaaagt tcatggcttg ctaaggtaac aacacagtgg ctcagttctc tggggtcaga
     2581 cgtctatgtc tctaggggta gtttgacggc tgaattgagg cggcgctggg gccttgacac
     2641 cgtcatcccc caggtacgat tcgagagcgg catgccagtg gttgacgagg agggtgcaga
     2701 gatcacgcct gaggaatttg agaaattcag gctccagtgg gaaggtcacc gggtcacacg
     2761 agagatgaga acagaccggc ggccagacaa gaggatagac cacaggcatc acctggtcga
     2821 tgccatcgtg actgccctta cttctcgctc tttgtatcag caatacgcaa aggcatggaa
     2881 ggtcgcggat gagaagcaga gacacggccg ggtggatgtg aaggtcgaac tgcctatgcc
     2941 tatccttaca atccgcgata tcgccttgga ggccgtgcgg agtgtgagaa tcagccacaa
     3001 gccggaccgc taccctgacg gccggttttt cgaagctacc gcttatggca tcgctcaacg
     3061 gctggacgag aggtctggtg agaaagttga ctggctcgtg tcacgcaagt ccctcaccga
     3121 tctggctccc gagaagaaat ctatagatgt agataaagta agagcgaata tctccaggat
     3181 cgtcggcgag gcgataagac tgcacatctc caacatcttc gagaaacgag tgagtaaagg
     3241 catgacacca cagcaggcac tgagagaacc aatcgaattt cagggaaata tcctgcggaa
     3301 agtgcgctgc ttttattcaa aagcggacga ttgtgtcagg atcgagcact caagtagacg
     3361 gggacatcac tacaaaatgc tgctgaacga cggtttcgcc tacatggagg taccatgcaa
     3421 ggaggggatt ctctacggag tgcctaatct cgttcgcccc agcgaggccg taggcatcaa
     3481 aagggccccc gagtcaggtg attttatacg attctacaaa ggagataccg ttaaaaatat
     3541 caagactgga agagtctaca cgatcaagca gatcctggga gacggaggag gtaagttgat
     3601 cctcacccca gtcacagaga caaagcctgc cgacctgctt agcgctaagt ggggacgact
     3661 caaagtgggg gggcgcaata tccatctgct gcggctctgt gctgagagca gggctgaccc
     3721 caagaagaag aggaaggtga ggtccatggt catagctgtt tcctgtgtga aattgttatc
     3781 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     3841 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     3901 acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta
     3961 ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcgtc
     4021 ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc
     4081 agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa
     4141 catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt
     4201 tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg
     4261 gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg
     4321 ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag
     4381 cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc
     4441 caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa
     4501 ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg
     4561 taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc
     4621 taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac
     4681 cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg
     4741 tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt
     4801 gatcttttct acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt
     4861 catgagatta tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa
     4921 atcaatctaa agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga
     4981 ggcacctatc tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt
     5041 gtagataact acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg
     5101 agacccacgc tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga
     5161 gcgcagaagt ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga
     5221 agctagagta agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg
     5281 catcgtggtg tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc
     5341 aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc
     5401 gatcgttgtc agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca
     5461 taattctctt actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac
     5521 caagtcattc tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg
     5581 ggataatacc gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc
     5641 ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg
     5701 tgcacccaac tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac
     5761 aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat
     5821 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
     5881 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
     5941 agtgccacct gacgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     6001 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     6061 gcagctcccg gagacggtca cagcttgtct
//
