LOCUS       sequence-201808-        5784 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 5784)
  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 5784)
  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-E0XXB7
FEATURES             Location/Qualifiers
     source          1..5784
                     /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             3421..3441
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(3451..3467)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3451..3467)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3464..3486)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3475..3491
                     /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(3499..3505,3506..3523,3524..3529))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3544..3565
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3903..4491)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3983..4002)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4662..5453,5454..5522))
                     /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     5285..5304
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5523..5627)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5695..5713
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5751..5773)
                     /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 ccaccatgag ctcaaaggct attgactccc tggagcagct cgacctgttc
      421 aaaccccaag aatatactct tgggctggat ctgggcatca agtcaattgg ctgggccata
      481 ctttcaggcg agaggattgc caacgcagga gtttacctgt tcgagactgc cgaagagctg
      541 aattctacag ggaataagct gatctctaaa gctgcagaac gcggaaggaa aagacggatc
      601 cgccgcatgc tggacagaaa ggcgcgaaga ggccgccata tcagatatct gctggaacgc
      661 gaaggcctgc cgacagacga actggaggaa gttgtcgtgc atcaatccaa cagaactctc
      721 tgggacgtca gagccgaggc ggtggagcgc aagctgacca agcaggagct cgccgccgtt
      781 ctcttccacc tcgtgagaca ccggggctac ttccccaaca cgaaaaagct gccacccgat
      841 gacgagagcg attccgctga tgaggagcag ggcaagataa accgggcgac aagcagactt
      901 agggaagagc tcaaggcctc agattgtaaa acaatcggtc aatttttggc ccagaatcgc
      961 gataggcaga ggaacagaga gggagactac tctaacctga tggctcgcaa actggtgttc
     1021 gaagaagcgc tccagatact ggcatttcaa cgcaaacagg ggcacgagct ctcaaaggac
     1081 ttcgagaaaa cctacctcga tgtacttatg ggtcagagaa gcggccgatc tcccaaactc
     1141 ggcaactgca gtctgatacc ttctgaactg agagcccctt catccgcccc cagcacagaa
     1201 tggttcaagt ttctgcaaaa cctgggcaat ttgcagattt ccaatgccta cagagaggaa
     1261 tggtccattg acgctcctag aagagcacag atcattgacg catgttcaca gaggtctacc
     1321 agcagttact ggcagattcg gagggacttc cagatcccgg acgaataccg gtttaacttg
     1381 gtgaactatg agaggcgcga ccccgatgtg gatctgcagg agtaccttca gcagcaggag
     1441 agaaaaaccc tcgccaattt ccgcaactgg aaacaactcg agaagattat cggcactggc
     1501 caccctatcc agactcttga cgaagcagca agacttatta ccctgataaa ggatgatgaa
     1561 aagctgagcg accaactggc tgacctgctg cccgaggcta gcgacaaagc aattactcag
     1621 ctgtgtgaac tggacttcac aaccgcagca aaaatctccc tcgaagcaat gtacagaatc
     1681 ctgcctcata tgaaccaggg aatgggattc tttgatgcat gtcagcagga gtctcttccg
     1741 gagattgggg ttccaccagc tggcgacagg gtgcccccct tcgacgaaat gtacaacccc
     1801 gtggtaaatc gagtgcttag ccaaagcaga aagctcataa acgccgtgat tgacgagtat
     1861 ggaatgcccg ccaagattag agtggagctg gctagagatc tggggaaggg ccgcgaactt
     1921 cgggaacgaa taaaactgga ccagttggac aagtctaagc agaatgatca gagagccgag
     1981 gactttcggg ccgaattcca gcaggccccc agaggggacc agtcactgcg gtaccggctg
     2041 tggaaggaac agaattgcac ctgtccttac tctgggcgga tgatccctgt gaattcagtg
     2101 ctgagcgaag atacccagat cgatcatata ctgcctatct cccagagttt cgataatagc
     2161 ttgtccaaca aggtattgtg ctttacagag gaaaacgccc agaagtcaaa caggacacca
     2221 tttgagtacc tggacgccgc ggacttccag cgacttgaag ccatctccgg taactggcca
     2281 gaagcgaaac ggaataagtt gctccacaag tcctttggta aggtggctga ggagtggaaa
     2341 tctagagctc tcaacgacac tcgatatctc acatcagccc tggcagatca tctgcggcac
     2401 cacctgccgg actcaaaaat ccagaccgtg aacggacgga ttacagggta cctgcggaaa
     2461 caatggggcc tggagaaaga ccgagacaag cacactcacc acgccgtgga cgccatcgtc
     2521 gtcgcctgca ccacacccgc catcgttcag caggtgaccc tgtaccacca ggacatcagg
     2581 agatataaaa aactgggcga aaagaggcct acaccatggc ctgaaacatt tcgacaggac
     2641 gttcttgacg ttgaagaaga gattttcata actcggcagc ccaagaaagt gtctggaggg
     2701 atccagacca aagacactct caggaaacac aggtccaaac cagacaggca gcgcgtcgca
     2761 ctcacgaaag tgaagctggc ggacctggag cggctcgtcg agaaggatgc ttctaataga
     2821 aacttgtacg agcacctgaa acagtgcttg gaggaatcag gggaccagcc caccaaagcg
     2881 ttcaaggcac ccttctatat gccctctggc ccggaggcga aacagaggcc aatcctgtca
     2941 aaagtgacgc tcctgcgcga gaaacctgag cctcctaaac aactgaccga gttgagcggg
     3001 gggcgaaggt acgattccat ggcacaaggc cgactcgata tttatcggta taagcccgga
     3061 ggcaaaagga aggacgagta ccgggtcgtg cttcagagaa tgatcgatct tatgaggggg
     3121 gaagagaatg tacacgtgtt tcagaaaggc gtgccctacg accaaggccc agagatagaa
     3181 cagaattata cattcttgtt cagcctgtat ttcgacgacc ttgtggagtt ccagagaagc
     3241 gctgacagcg aggtcattag agggtattac cggaccttta acatcgcgaa cggacagctg
     3301 aagatctcta cctacctgga gggtcgccaa gactttgact tttttggggc caaccggctg
     3361 gcacattttg ccaaagtcca ggtgaacctc ctcgggaagg tgattaaaag cagggctgac
     3421 cccaagaaga agaggaaggt gaggtccatg gtcatagctg tttcctgtgt gaaattgtta
     3481 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     3541 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     3601 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     3661 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     3721 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
     3781 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
     3841 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
     3901 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
     3961 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
     4021 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
     4081 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
     4141 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
     4201 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
     4261 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
     4321 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
     4381 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
     4441 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
     4501 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
     4561 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
     4621 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
     4681 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     4741 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     4801 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     4861 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     4921 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     4981 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     5041 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     5101 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     5161 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     5221 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     5281 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     5341 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     5401 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     5461 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     5521 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     5581 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     5641 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     5701 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     5761 atgcagctcc cggagacggt caca
//
