LOCUS       sequence-201803-        6840 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 6840)
  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 6840)
  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-C7TEQ6
FEATURES             Location/Qualifiers
     source          1..6840
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     89..108
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     302..324
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     316..333
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     317..333
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4477..4497
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4507..4523)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4507..4523)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4520..4542)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4531..4547
                     /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(4555..4561,4562..4579,4580..4585))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4600..4621
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4959..5547)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5039..5058)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5718..6509,6510..6578))
                     /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     6341..6360
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6579..6683)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6751..6769
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6807..6829)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gcttgtctgt aagcggatgc cgggagcaga caagcccgtc agggcgcgtc agcgggtgtt
       61 ggcgggtgtc ggggctggct taactatgcg gcatcagagc agattgtact gagagtgcac
      121 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgccat
      181 tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc ttcgctatta
      241 cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac gccagggttt
      301 tcccagtcac gacgttgtaa aacgacggcc agtgtgctgt ctcatcattt tggcaaagaa
      361 ttcgcggccg ccaccatgac aaagctgaac cagccctatg gaatcggcct cgatattgga
      421 agtaatagca tcggcttcgc cgtggtagat gcgaactcac acctgcttcg cctgaaaggg
      481 gagactgcca tcggagctag gctgtttcgc gaagggcaaa gtgcagccga tcggagaggt
      541 agccgcacaa caaggagaag actcagtcga actagatgga ggctgtcctt cctgagggac
      601 tttttcgccc cacacataac caagattgat cctgattttt tcctgagaca gaagtacagc
      661 gaaatcagcc ccaaagataa ggaccggttt aagtatgaaa aaagattgtt taatgacaga
      721 acagacgcgg aattctacga ggattatccc agcatgtacc atcttaggct ccaccttatg
      781 acacacacac ataaggctga tccccgcgag attttcctcg ccattcacca tattctgaag
      841 tctcgagggc atttccttac ccccggggcg gccaaagact tcaacaccga taaagtggac
      901 cttgaggaca ttttccctgc cctgacagaa gcatacgcgc aggtgtatcc cgacctggag
      961 ttgacctttg atctggctaa ggccgacgat ttcaaggcaa agctgcttga tgaacaggcg
     1021 acacccagcg acactcagaa ggcgctcgtc aatttgcttc tgtcaagcga cggggaaaaa
     1081 gaaatcgtca agaagagaaa gcaggtcttg accgaattcg ctaaggctat caccggactt
     1141 aagacaaaat tcaaccttgc actggggacc gaagtcgatg aggccgatgc ttctaactgg
     1201 cagttcagca tgggccagct tgacgacaaa tggtctaata ttgagacttc tatgactgac
     1261 caagggaccg aaatttttga acagatccag gagctctatc gcgcaaggct gctgaatggg
     1321 attgtgcctg ccggaatgtc tctgtcccag gccaaagtgg ctgactacgg gcagcataag
     1381 gaggatctcg agctcttcaa gacctatctg aagaaactga acgatcacga actcgctaaa
     1441 accatccgcg gcctctacga caggtacatc aacggggatg acgctaagcc cttcctcaga
     1501 gaagactttg tgaaagccct gacaaaagag gttacagcgc atccaaatga ggtaagcgag
     1561 cagctgctga atcgaatggg acaggctaac tttatgctga agcagcgaac caaagccaac
     1621 ggggcgatcc caatccagct gcaacagagg gagctggacc agattatcgc gaatcagagt
     1681 aagtactacg attggcttgc agccccaaac ccagtcgagg cacacagatg gaagatgcct
     1741 tatcagctgg atgaacttct gaatttccac attccatact atgtcgggcc attgatcaca
     1801 ccgaagcagc aggctgagtc cggggagaac gtctttgctt ggatggtgcg caaggatcct
     1861 tcaggcaata taacccccta caatttcgac gagaaggttg atagagaagc ctccgctaat
     1921 acatttatac agcggatgaa aaccacagat acctatctga ttggcgaaga tgtactcccc
     1981 aaacagtcct tgctgtatca aaagtatgag gtgcttaacg agctgaataa tgtaaggatt
     2041 aataatgagt gccttggcac cgaccaaaag cagcgactca tccgggaagt gttcgaacgg
     2101 cacagttcag tgaccatcaa acaggtggcg gacaacctcg tagcacatgg cgacttcgca
     2161 cgcaggcctg aaatccgcgg ccttgcagat gaaaagcgct ttctctcatc tctgtcaaca
     2221 taccaccagc tgaaagagat tctgcacgaa gcgattgatg acccaactaa acttctggat
     2281 atagagaata ttatcacctg gtctacagtg ttcgaggatc atactatctt cgagacaaaa
     2341 ctggctgaga tcgagtggtt ggaccccaaa aagatcaatg aactgagtgg aatcaggtat
     2401 cgaggatggg ggcaattcag ccggaaactg cttgacggtc tgaagctggg aaacgggcat
     2461 acagtgattc aggagctcat gctctccaat cataatctca tgcagattct ggcagacgaa
     2521 acgctcaaag aaacaatgac agaactcaac caggacaagc tgaaaactga tgatatagag
     2581 gatgtgataa acgacgcata cacaagtcca agtaataaaa aggcccttcg ccaggtactc
     2641 agagtggtgg aggatattaa gcacgccgcc aacggccaag accctagctg gctgtttatc
     2701 gagactgctg acgggactgg taccgctggt aagcggaccc aaagtcgaca gaaacagata
     2761 cagaccgtct acgcaaacgc cgcccaggaa ttgattgatt cagctgtcag aggagagctc
     2821 gaggacaaga tcgccgataa ggcatctttt acggataggc ttgtcctcta tttcatgcag
     2881 ggcggacgcg acatctacac gggcgcaccc ctcaacattg accaactgtc tcattacgat
     2941 atcgaccata ttctgcctca aagcctgatt aaagatgatt cccttgacaa ccgagtcctc
     3001 gtcaatgcca ctattaatcg ggaaaaaaat aacgtgtttg catcaacatt gttcgccggt
     3061 aaaatgaagg ctacatggcg gaaatggcat gaagccggac tgattagtgg aaggaagctt
     3121 cggaatctga tgctccgacc agatgaaatc gataaatttg caaagggatt cgtagctcgc
     3181 caactggtgg agactcgaca gataattaaa ttgaccgaac aaattgccgc tgcgcagtac
     3241 ccaaatacta agatcatcgc agtcaaggca ggtctgagtc atcagctgcg cgaggagctc
     3301 gacttcccaa aaaaccggga tgtaaatcac tatcatcatg cttttgacgc ctttctcgcc
     3361 gcccgaatag ggacctatct cctgaaaaga taccccaaac tcgctccttt cttcacttac
     3421 ggcgagtttg cgaaggttga cgtcaagaag ttccgggaat tcaacttcat tggcgcactc
     3481 acacacgcta agaagaatat aattgccaag gacaccgggg aaattgtgtg ggacaaagag
     3541 agggacatac gggagctgga ccgaatctat aatttcaaga ggatgcttat tacacacgag
     3601 gtatattttg aaaccgctga cttgttcaag cagacgatct acgccgccaa agactctaaa
     3661 gagcggggag gttccaaaca gctcattccc aagaagcaag gctaccctac tcaggtgtac
     3721 ggtgggtata cccaggagtc tggaagctac aacgcactgg tgcgagtggc agaagcagat
     3781 acaaccgcct atcaggttat caagatcagc gcacagaatg ccagcaaaat cgcatctgca
     3841 aacctgaaat cccgggaaaa gggtaagcag ctgttgaacg aaatagtcgt gaagcagctg
     3901 gccaagagac gcaagaactg gaagccttct gcgaattctt tcaagatcgt gatccctcgg
     3961 ttcggaatgg gaacactctt ccaaaacgcc aagtatggcc tgtttatggt gaacagtgac
     4021 acgtactata ggaattatca ggagctgtgg ctgagccgag agaaccagaa actcctgaaa
     4081 aaacttttct ctatcaagta cgaaaagacc cagatgaatc atgacgctct tcaggtgtat
     4141 aaagcaatta tcgaccaagt ggagaaattc tttaagctgt atgatattaa tcagtttagg
     4201 gctaaactgt ccgatgcgat cgagcggttc gagaagctcc ctatcaacac cgacggaaat
     4261 aaaatcggga agaccgaaac cttgcgccag atactcattg gccttcaagc caacggaacc
     4321 aggagcaatg tcaaaaactt gggcattaag acggatctgg ggctccttca ggtggggagt
     4381 ggcatcaagc tcgacaaaga cacacagatt gtgtaccagt ctcctagtgg gctgttcaag
     4441 cgcagaattc ccctcgccga tctgagcagg gctgacccca agaagaagag gaaggtgagg
     4501 tccatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
     4561 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
     4621 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
     4681 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
     4741 gctcactgac tcgctgcgct cggtcgttcg gctgcgtctt ccgcttcctc gctcactgac
     4801 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4861 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4921 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4981 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5041 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5101 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     5161 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     5221 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     5281 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5341 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     5401 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5461 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5521 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5581 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5641 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5701 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5761 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5821 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5881 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5941 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     6001 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     6061 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     6121 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     6181 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     6241 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     6301 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     6361 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     6421 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     6481 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     6541 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     6601 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     6661 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtctaagaa
     6721 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc
     6781 gcgcgtttcg gtgatgacgg tgaaaacctc tgacacatgc agctcccgga gacggtcaca
//
