LOCUS       sequence-201871-        6917 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 6917)
  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 6917)
  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-G9WGU4
FEATURES             Location/Qualifiers
     source          1..6917
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     96..115
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     309..331
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     323..340
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     324..340
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      377..386
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4562..4582
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4591..4607)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4591..4607)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4604..4626)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4615..4631
                     /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(4639..4645,4646..4663,4664..4669))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4684..4705
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5043..5631)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5123..5142)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5802..6593,6594..6662))
                     /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     6425..6444
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6663..6767)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6835..6853
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6891..6913)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
       61 gggtgttggc gggtgtcggg gctggcttaa ctatgcggca tcagagcaga ttgtactgag
      121 agtgcaccat atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
      181 gcgccattcg ccattcaggc tgcgcaactg ttgggaaggg cgatcggtgc gggcctcttc
      241 gctattacgc cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc
      301 agggttttcc cagtcacgac gttgtaaaac gacggccagt gtgctgtctc atcattttgg
      361 caaagaattc gcggccgcca ccatggcccg agactacagt gtcggcctcg acataggaac
      421 ttctagcgtg ggttgggcag ccatcgacaa taaatatcat ttgatcaggg ccaagagcaa
      481 aaacctgata ggcgtgcgcc tgtttgattc cgccgtgacg gcagagaaaa gacgggggta
      541 tcgaaccact cgaaggcggc tgtcccggcg ccattggcgg ctgaggttgc tgaacgacat
      601 tttcgccggc ccactgacgg atttcggcga cgagaacttt cttgcacgcc tgaagtactc
      661 ctgggtgcac cctcaggatc aaagcaatca ggctcacttc gcagccggac tgctgtttga
      721 ctccaaggag caagacaagg atttttacag aaagtacccc actatctatc acctgagact
      781 ggcactgatg aacgacgatc aaaagcatga ccttagggag gtgtacctcg ccattcatca
      841 tcttgtcaaa tacagaggcc atttcctgat cgagggcgac gtcaaagccg actctgcctt
      901 tgacgtgcac actttcgccg atgccattca gaggtacgca gaatccaaca acagcgatga
      961 aaacctgttg ggaaaaatcg atgagaagaa gctgagtgct gctctcactg ataagcacgg
     1021 ttctaaaagt cagcgagccg aaaccgctga gacggccttc gacattttgg acctgcaatc
     1081 aaaaaaacag atccaggcca tactgaagag tgttgtcggc aaccaagcta acctcatggc
     1141 aatcttcggg cttgatagca gtgccatctc aaaagatgag cagaagaatt acaagttttc
     1201 atttgacgac gcagatattg acgagaagat tgccgattcc gaggctctgt tgtccgacac
     1261 cgagttcgaa ttcctgtgtg acctgaaggc agcattcgat ggtctgacac tgaagatgct
     1321 gttgggagat gacaaaacgg tatctgccgc tatggtaaga cggtttaatg agcatcagaa
     1381 ggactgggag tatatcaaat cccatattcg caacgctaag aatgccggaa atggcctcta
     1441 cgagaagagc aaaaaatttg acgggatcaa cgcagcctat ctcgccttgc agtctgacaa
     1501 cgaagatgac agaaaaaaag caaaaaaaat tttccaggac gagatctcct cagcagatat
     1561 tcctgatgac gttaaggctg actttctgaa aaaaatcgac gatgaccagt tcctgcccat
     1621 ccagagaacc aagaacaatg ggactatccc acatcagctg caccgaaacg agctggagca
     1681 gatcatcgaa aagcagggca tctattatcc ctttcttaaa gacacctacc aggagaactc
     1741 acacgaactc aacaagatca ccgcattgat caactttagg gtgccttact atgtggggcc
     1801 gctcgttgag gaggagcaga aaatcgctga cgatgggaag aacatccccg atccgacaaa
     1861 tcactggatg gtacgcaaat caaacgatac aatcacgccg tggaatttga gccaagtggt
     1921 tgacctggac aagagtggca gaaggtttat tgagcgcttg accggaaccg atacatactt
     1981 gatcggagag ccaacactgc ccaagaactc actgttgtat cagaaattcg atgttctcca
     2041 ggaactgaac aatattcggg tgtccggtcg cagactggac atcagagcca agcaggatgc
     2101 ctttgaacac ctgttcaaag tgcagaaaac cgtctcagcc acgaatctta aggactttct
     2161 ggttcaagcc ggttatattt ccgaggacac acaaatcgaa ggcctggccg atgtgaacgg
     2221 caaaaatttc aacaacgccc tcacgacata caactacctt gtcagcgtat tgggccgcga
     2281 gttcgtggag aatccaagca acgaggaact cctggaagag atcacagagc ttcaaaccgt
     2341 attcgaggat aaaaaagttt tgcgccggca gctcgatcag ctggacgggc tctccgacca
     2401 taaccgggag aagctgagta ggaagcacta tactgggtgg ggaagaattt caaagaagtt
     2461 gctgactacc aaaatcgtac agaatgccga caagattgat aatcagacct ttgatgtgcc
     2521 tcggatgaac cagtcaatca tcgacactct gtacaataca aaaatgaatt tgatggagat
     2581 aattaacaat gccgaggacg atttcggggt cagagcatgg attgataagc agaataccac
     2641 tgatggagac gagcaggacg tttacagtct tatcgacgag ctggcgggcc ctaaggagat
     2701 caagaggggg atagtccagt ctttccgcat tcttgacgac atcactaaag cggtcggcta
     2761 cgccccgaaa cgggtatacc tggaattcgc gcgcaagacc caggagagtc acctgactaa
     2821 tagcaggaag aaccagctga gcaccctgct gaagaacgcg ggactgtccg agctggtcac
     2881 acaagtgtct cagtatgatg cagctgccct gcagaatgat agactgtatc tttatttcct
     2941 gcaacagggg aaggatatgt actcaggcga aaagctgaat ctcgacaatc tctctaacta
     3001 cgatatcgat catatcatcc cacaggccta taccaaggat aactcactcg ataatcgcgt
     3061 gctggtcagc aatatcacca atagacggaa gagcgattct tctaactacc tgccagccct
     3121 gatagataaa atgcgacctt tctggtcagt gctgagtaag cagggtctgt tgagtaaaca
     3181 caaattcgct aatctgaccc ggacgaggga ttttgacgac atggagaaag aaagattcat
     3241 cgctcgaagc ctggtggaga ctaggcaaat tattaaaaac gtagcgagcc tcatcgacag
     3301 tcatttcggt ggagagacaa aagccgtcgc catacgctca agccttaccg cggacatgag
     3361 acgctacgtg gacatcccta aaaaccggga tatcaatgac tatcatcacg cctttgacgc
     3421 ccttcttttc agcaccgtag gccagtatac tgagaactcc ggactgatga agaaggggca
     3481 gctttccgac agcgctggta accagtataa ccggtatatc aaggaatgga ttcacgccgc
     3541 ccggctgaac gcccagtctc agcgcgtgaa cccctttgga ttcgtggtgg gctccatgag
     3601 aaacgctgcg cccggcaaac ttaatcctga gactggcgag ataacaccag aggagaatgc
     3661 agactggtca attgcagacc ttgactatct gcataaggtg atgaacttta ggaagataac
     3721 agtaacccga aggctgaaag accaaaaggg acagctttat gacgagagca gatacccatc
     3781 tgtcctccac gacgcaaaga gcaaagctag catcaatttc gataaacaca aaccagtcga
     3841 cctttatgga ggtttctctt cagccaagcc cgcttacgcc gcattgatca agttcaaaaa
     3901 taagttccgg ttggtgaatg tcctgaggca atggacttat tcagacaaga attccgagga
     3961 ttacattctt gagcagatca gaggaaagta cccgaaggct gagatggtgc tttcacatat
     4021 accttacgga cagttggtca agaaagacgg ggctttggtg actatatcaa gcgccactga
     4081 actccataat ttcgaacaac tttggctgcc gttggctgac tacaaattga ttaacacttt
     4141 gctcaagacc aaggaggata acctggtcga catccttcat aaccgcctgg atctgcccga
     4201 gatgaccatc gagtccgcct tttataaagc atttgacagt atcctgtcct ttgctttcaa
     4261 caggtatgcc ctgcatcaga acgcgcttgt gaagctgcag gctcatcggg acgacttcaa
     4321 tgctctgaat tacgaggaca aacagcagac cctggaacga atactggatg ccctgcacgc
     4381 atcccccgcc tctagcgacc ttaaaaaaat caatctctct tccggcttcg gtcggctgtt
     4441 tagcccaagt cacttcaccc tggccgacac ggatgagttc attttccagt ctgtgaccgg
     4501 acttttcagc actcagaaaa ctgtggcaca gctttaccag gaaaccaaaa gcagggctga
     4561 ccccaagaag aagaggaagg tgaggtccat gtcatagctg tttcctgtgt gaaattgtta
     4621 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4681 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4741 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4801 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4861 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     4921 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     4981 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     5041 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     5101 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     5161 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     5221 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     5281 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     5341 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     5401 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     5461 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     5521 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     5581 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     5641 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     5701 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     5761 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     5821 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     5881 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     5941 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     6001 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     6061 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     6121 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     6181 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     6241 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     6301 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     6361 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     6421 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     6481 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     6541 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     6601 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     6661 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     6721 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     6781 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     6841 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     6901 atgcagctcc cggagac
//
