LOCUS       sequence-201883-        6249 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 6249)
  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 6249)
  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-Q1QGC9
FEATURES             Location/Qualifiers
     source          1..6249
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(4..26)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     126..145
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     339..361
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     353..370
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     354..370
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(1604..1615)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             3923..3943
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3953..3969)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3953..3969)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3966..3988)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3977..3993
                     /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(4001..4007,4008..4025,4026..4031))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4046..4067
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4405..4993)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4485..4504)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5164..5955,5956..6024))
                     /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     5787..5806
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6025..6129)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6197..6215
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 aaacctctga cacatgcagc tcccggagac ggtcacagct tgtctgtaag cggatgccgg
       61 gagcagacaa gcccgtcagg gcgcgtcagc gggtgttggc gggtgtcggg gctggcttaa
      121 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atgcggtgtg aaataccgca
      181 cagatgcgta aggagaaaat accgcatcag gcgccattcg ccattcaggc tgcgcaactg
      241 ttgggaaggg cgatcggtgc gggcctcttc gctattacgc cagctggcga aagggggatg
      301 tgctgcaagg cgattaagtt gggtaacgcc agggttttcc cagtcacgac gttgtaaaac
      361 gacggccagt gtgctgtctc atcattttgg caaagaattc gcggccgcca ccatgcacgt
      421 tgagattgat ttcccgcact tcagcagagg tgacagccat ctcgcaatga acaagaatga
      481 aattctgcgg ggcagctctg tgctctatag actgggcctc gacctgggga gtaattctct
      541 gggctggttt gttacacacc tcgaaaagag gggggacagg catgaacccg tggctctcgg
      601 ccctggtggt gtcagaattt tccccgatgg gagagatcca cagtcaggta caagtaatgc
      661 ggtggacagg aggatggccc gaggtgccag aaaaaggcga gatcgatttg tcgaaagaag
      721 aaaagaattg atcgcggccc tgattaagta taatctgctg ccggacgatg ctcgagaacg
      781 ccgggctctg gaagtacttg atccttacgc actgcggaag accgctctga ctgacacact
      841 tcccgcacac catgttggga gagctctttt ccatctcaat cagaggcgcg gcttccaaag
      901 caacagaaaa accgacagca agcagtctga ggacggtgcc attaaacagg ctgccagtcg
      961 ccttgctacc gacaagggga acgagacgtt gggggtgttc tttgctgata tgcacctgcg
     1021 aaaatcctat gaagatcggc agactgccat tcgcgcggag ctggttaggc tgggcaaaga
     1081 ccacctcaca ggcaacgctc gcaagaagat ctgggcaaag gtcaggaaaa gattgtttgg
     1141 tgatgaggtc ttgccacggg cggacgctcc gcacggcgtt cgagccagag caaccattac
     1201 tggcaccaaa gccagctacg attattaccc cacacgcgat atgctccgag acgagttcaa
     1261 cgcgatctgg gccggtcaat ctgcccacca tgctactatc actgacgaag ctaggacaga
     1321 aattgaacat atcatttttt atcagaggcc ccttaagccg gcaatcgtgg gaaagtgtac
     1381 acttgacccg gctactaggc ccttcaagga ggaccccgaa ggatatcggg ccccttggag
     1441 ccaccccctc gcccagcgct ttcgcatcct gagtgaagcc agaaatctgg agatcaggga
     1501 taccggtaaa ggcagcagac gcctgaccaa ggaacaatca gaccttgtcg tcgcagctct
     1561 gctggccaac cgggaagtca agttcgataa acttagaaca ctgctgaagc tcccagccga
     1621 agccagattt aaccttgaga gcgaccgccg ggccgcactt gatggagatc aaactgccgc
     1681 aagactgagc gacaagaagg ggtttaataa ggcctggcgc ggattcccac ccgagcgcca
     1741 aatcgctatt gttgctagac tcgaagagac tgaggatgag aacgagctga ttgcctggct
     1801 ggaaaaggag tgcgccctcg acggcgcagc tgctgctcgc gtggccaaca cgacactccc
     1861 cgatgggcac tgcaggctgg gcctcagggc gattaaaaag attgtgccaa taatgcagga
     1921 cggactcgac gaggatgggg tggcaggggc aggctaccat atcgccgcca agagagcagg
     1981 atatgatcac gcaaaacttc ccactggaga acagctggga cgactgcctt actacggcca
     2041 gtggctccag gacgccgtgg tgggatccgg cgatgcaaga gatcagaagg agaaacagta
     2101 cgggcaattt cctaatccta ctgtgcacat tggcctcggc cagctgcgga gagtcgtgaa
     2161 cgatttgatt gacaagtatg gccccccaac cgagatctct attgagttta cacgggcact
     2221 gaaattgtct gaacagcaaa aggccgagcg gcagcgggaa cagcggcgaa atcaggataa
     2281 gaacaaagcc cgcgccgaag aactggcgaa gttcgggcgc cccgcaaacc caagaaacct
     2341 cctgaaaatg cgactgtggg aagaattggc ccatgatcca ctggatagaa agtgcgttta
     2401 tacgggggag cagataagca tagagcggct gctttctgac gaggtcgaca tagatcacat
     2461 tctccccgtg gccatgacac tggacgactc acctgcaaac aagatcattt gcatgcgata
     2521 tgctaaccgc cacaagagaa agcagacacc ctcagaggcc tttgggtcat caccgacact
     2581 tcaggggcac agatataatt gggacgatat agctgcccgg gcaaccggtc tgccacggaa
     2641 caaacgctgg cggtttgacg caaacgccag agaggaattc gacaagaggg gcggatttct
     2701 ggccaggcaa ctgaacgaga ctgggtggct ggcccgcctg gcaaagcaat accttggcgc
     2761 cgtgacggac ccgaatcaga tctgggttgt ccctggtcgg ctcacctcaa tgctgcgcgg
     2821 gaagtggggc cttaatggtc tgcttcctag tgacaactat gcgggtgtgc aggacaaggc
     2881 cgaagagttc ctcgcatcca ccgacgacat ggagttcagc ggcgtgaaga acagggccga
     2941 tcaccggcac cacgcaattg acgggctggt gactgccttg accgacagga gtctcctctg
     3001 gaaaatggct aatgcatacg acgaagagca tgaaaaattt gtgatcgagc caccttggcc
     3061 taccatgagg gacgacctga aggccgcgct ggaaaaaatg gtcgtctctc ataagccaga
     3121 tcacggaata gaaggcaagc tgcacgagga ttccgcttac gggttcgtga aaccacttga
     3181 cgccaccgga ctcaaggagg aggaggctgg caacctggtg tatagaaagg ctatcgagtc
     3241 tcttaacgag aacgaggtgg atcggatcag ggatatccag cttcggacta ttgtcaggga
     3301 tcacgtcaat gtagaaaaga ccaagggtgt tgctctcgct gatgcactcc gccagcttca
     3361 ggccccgagc gatgattacc ctcaatttaa gcacggtctg cggcacgtga ggatattgaa
     3421 aaaggagaag ggcgattatt tggtacccat agctaaccgg gcttccggag tggcctacaa
     3481 ggcttactca gcgggagaga acttctgcgt agaagtgttt gaaaccgctg gaggtaagtg
     3541 ggatggggaa gcagtcaggc gcttcgacgc caataagaag aacgccggcc ccaagattgc
     3601 tcacgctcct cagtggagag atgccaacga gggggccaaa ctggtaatgc ggattcacaa
     3661 aggggatctg ataaggttgg accacgaagg cagagcgaga ataatggttg tgcacagact
     3721 tgatgccgct gcggggcgct tcaaactggc tgatcataat gaaaccggta acctcgacaa
     3781 gagacacgca acaaacaacg acatcgaccc ctttaggtgg ctgatggcca gctacaatac
     3841 cctgaagaag ctggccgcgg tgcccgtgcg agtagatgaa ctgggcagag tgtggcgggt
     3901 catgcccaac agcagggctg accccaagaa gaagaggaag gtgaggtcca tggtcatagc
     3961 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     4021 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     4081 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     4141 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     4201 tgcgctcggt cgttcggctg cgtcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4261 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4321 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4381 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4441 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4501 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4561 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4621 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4681 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4741 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4801 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     4861 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4921 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4981 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5041 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5101 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5161 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5221 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5281 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5341 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5401 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5461 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5521 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5581 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5641 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5701 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5761 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5821 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5881 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5941 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6001 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6061 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6121 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6181 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc gtttcggtga
     6241 tgacggtga
//
