LOCUS       sequence-201692-        6165 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 6165)
  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 6165)
  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-A0LWB3
FEATURES             Location/Qualifiers
     source          1..6165
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     110..129
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     323..345
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     337..354
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     338..354
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      391..400
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3823..3843
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3853..3869)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3853..3869)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3866..3888)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3877..3893
                     /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(3901..3907,3908..3925,3926..3931))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3946..3967
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4305..4893)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4385..4404)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5064..5855,5856..5924))
                     /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     5687..5706
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5925..6029)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6097..6115
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(join(6153..6165,1..10))
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag acaagcccgt
       61 cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc ggcatcagag
      121 cagattgtac tgagagtgca ccatatgcgg tgtgaaatac cgcacagatg cgtaaggaga
      181 aaataccgca tcaggcgcca ttcgccattc aggctgcgca actgttggga agggcgatcg
      241 gtgcgggcct cttcgctatt acgccagctg gcgaaagggg gatgtgctgc aaggcgatta
      301 agttgggtaa cgccagggtt ttcccagtca cgacgttgta aaacgacggc cagtgtgctg
      361 tctcatcatt ttggcaaaga attcgcggcc gccaccatgg ggggttcaga ggtcggtacc
      421 gtgccggtaa cctggcggct cggcgtggac gtgggggaaa gatcaatcgg gcttgctgcc
      481 gtgtcatatg aagaggacaa gcctaaggaa atcttggctg cagtgtcctg gatccatgat
      541 gggggggttg gcgatgaacg gtccggggca agtcgacttg ccctccgagg catggcaaga
      601 agagcccgaa ggctgcgccg gtttaggaga gcccgcctcc gcgacctgga catgttgctg
      661 agtgagttgg gatggacccc cctccctgac aaaaacgtct caccagttga tgcctggctg
      721 gcacgcaaaa gactggccga ggaatatgtg gtggatgaaa ctgagaggcg aagactgctg
      781 ggctacgccg tgtctcatat ggcccggcac cgagggtggc gcaatccatg gactacgatt
      841 aaggacctga aaaatctgcc acagccctca gactcatggg agcggactag agagtcactg
      901 gaagccaggt atagcgtgtc tctggagccc ggcactgtcg ggcaatgggc aggctatctt
      961 ctgcagagag caccgggcat aagacttaat cccacacaac agtccgccgg tcgaagggcc
     1021 gagttgagta acgcaacagc ctttgagaca agactgcgac aagaagacgt cctttgggaa
     1081 ctgagatgca tcgccgatgt gcaagggttg cccgaggacg tcgttagcaa cgttatagac
     1141 gcagtgtttt gccagaaacg gccctccgta ccagcggaaa gaatcggcag agatccgctg
     1201 gaccccagcc agcttcgcgc tagcagagcc tgtctggagt ttcaagagta tcggattgtg
     1261 gcagccgtcg cgaatctgag aattagagat ggcagcggta gtcggccact gtctttggag
     1321 gagcggaatg ccgttataga ggctctgctt gcgcagaccg aaaggtcttt gacgtggagc
     1381 gacattgcgc tggaaattct gaaactgcct aacgaatccg acctgacctc tgtcccggag
     1441 gaagacgggc ctagctccct ggcctactct caatttgctc cgttcgatga gacttcagcc
     1501 cgaatcgcgg aattcattgc aaaaaaccgg cgcaagatcc ccacctttgc tcagtggtgg
     1561 caagagcaag atcggacaag ccgcagtgac ctggtggctg ctctggcaga taactcaatc
     1621 gctggcgagg aagaacagga gctgcttgtc cacctgccgg acgctgagct cgaggccctt
     1681 gaagggctcg ccttgcctag cggccgggtg gcctactccc gcctgacgct gtccggactt
     1741 accagggtga tgagagacga tggagtggac gttcacaacg ctcgcaagac ttgctttgga
     1801 gttgacgaca actggcgccc cccgctgcct gccctgcacg aagctaccgg acatcccgtg
     1861 gtcgatcgga atttggcgat cctgcggaag tttctctcat ccgctaccat gaggtgggga
     1921 ccacctcaat ctatcgtggt ggagctggcc cggggcgcca gcgagtctcg agagcgacag
     1981 gccgaggagg aagccgccag aagagcccat cggaaagcca acgatcgaat aagagccgag
     2041 ctgcgggcat ccggactcag cgatcctagc cctgctgacc ttgtccgcgc aagactgctg
     2101 gaactgtatg actgccactg catgtattgt ggggcaccta taagctggga gaactctgaa
     2161 ctggaccaca tcgtcccacg gactgatggc ggatcaaatc gccacgagaa tctggccatt
     2221 acttgcggcg catgtaacaa ggagaaaggt cgacggccgt ttgcgagttg ggccgagacg
     2281 tcaaataggg tgcagctgag ggatgtcatc gatcgcgtgc agaaactcaa atactcagga
     2341 aatatgtatt ggaccagaga cgaatttagc agatacaaaa agtctgtggt agctagactc
     2401 aagcgaagga cgtcagaccc cgaggtcatc cagagtattg agagtaccgg ctatgccgca
     2461 gtggcgctcc gagacagact gctgagctac ggcgagaaaa acggggtggc tcaggtcgca
     2521 gtctttcgcg gaggcgttac agctgaagcg agacgctggc ttgatatatc catagagagg
     2581 ctgttttcta gggtggccat ttttgcgcag tccactagca caaagaggct cgacagacga
     2641 catcatgccg tggatgctgt ggttctgaca acgctcacac ctggtgttgc gaagactttg
     2701 gctgatgcgc gcagcagacg agttagcgcc gagttctggc ggaggcctag cgacgtcaac
     2761 cgacacagca ctgaggagcc gcagagcccg gcctacaggc aatggaagga atcttgttct
     2821 ggtctgggcg acttgctcat ttccactgct gcacgagatt ccattgcagt agctgcaccc
     2881 ttgaggctcc ggccgactgg agcactgcac gaagaaaccc ttagggcctt cagtgaacac
     2941 acggtcgggg cagcctggaa aggcgcagag ctgcgcagga ttgtggaacc cgaagtatac
     3001 gccgcctttc tggctttgac cgaccctggc ggccgcttcc tgaaagtgtc accttcagaa
     3061 gatgtgctgc ccgcggacga aaacagacat atcgtgctgt cagatagagt cctggggcca
     3121 cgggacagag tgaagctgtt cccagacgac cggggaagta taagggtgcg agggggtgcg
     3181 gcgtatattg ctagcttcca ccacgctcgc gtgtttaggt ggggaagctc tcacagcccc
     3241 tcatttgccc tgctgcgcgt cagtcttgca gatcttgcgg tagccggact gttgagggat
     3301 ggggtggacg tcttcactgc cgaactccct ccttggacac ctgcctggag gtatgcgtca
     3361 atcgctctgg tcaaagcggt cgagagtggc gatgctaaac aggttggctg gttggtccct
     3421 ggcgacgagc tggacttcgg gcctgaaggg gtcaccacag cggcgggcga cctcagtatg
     3481 ttcttgaaat attttccaga gcgccactgg gttgttacag gatttgaaga cgataagaga
     3541 atcaacctta agcccgcctt cctctctgct gaacaggcag aggtgctgcg caccgagcgc
     3601 agtgaccggc cggacaccct gaccgaggct ggagagatac tcgcccaatt tttcccccgc
     3661 tgttggagag caaccgtcgc caaggtgctc tgtcacccgg gactgacggt tattagaaga
     3721 acagcccttg gccagccaag gtggcgaagg ggtcacctgc catatagctg gcgtccctgg
     3781 tcagccgacc catggagcgg tggaacccca agcagggctg accccaagaa gaagaggaag
     3841 gtgaggtcca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
     3901 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
     3961 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
     4021 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
     4081 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cgtcttccgc ttcctcgctc
     4141 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4201 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     4261 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     4321 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     4381 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     4441 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     4501 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     4561 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     4621 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     4681 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     4741 agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     4801 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     4861 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     4921 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     4981 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5041 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5101 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5161 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     5221 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     5281 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     5341 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     5401 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     5461 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     5521 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     5581 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     5641 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     5701 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     5761 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     5821 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     5881 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     5941 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6001 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
     6061 taagaaacca ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt
     6121 cgtctcgcgc gtttcggtga tgacggtgaa aacctctgac acatg
//
