LOCUS       sequence-201816-        7035 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 7035)
  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 7035)
  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-F3QMY7
FEATURES             Location/Qualifiers
     source          1..7035
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     122..141
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     335..357
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     349..366
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     350..366
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      403..412
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4705..4725
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4735..4751)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4735..4751)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4748..4770)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4759..4775
                     /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(4783..4789,4790..4807,4808..4813))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4828..4849
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5187..5775)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5267..5286)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5946..6737,6738..6806))
                     /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     6569..6588
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6807..6911)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6979..6997
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(join(7035,1..22))
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga tgccgggagc
       61 agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg gcttaactat
      121 gcggcatcag agcagattgt actgagagtg caccatatgc ggtgtgaaat accgcacaga
      181 tgcgtaagga gaaaataccg catcaggcgc cattcgccat tcaggctgcg caactgttgg
      241 gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct
      301 gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg
      361 gccagtgtgc tgtctcatca ttttggcaaa gaattcgcgg ccgccaccat gggtaagaca
      421 cacatcatcg gcgtcggtct tgacctgggc ggaacctata ccggtacctt tataacctcc
      481 catcccagtg acgaagccga gcaccgcgat catagctctg catttaccgt ggtgaactcc
      541 gagaagctct cttttagctc taagtcacgg acagccgttc gccatcgggt acggagctac
      601 aaggggtttg acctgagacg gagattgctc ctgctcgtgg ctgagtacca acttctgcag
      661 aaaaagcaaa ccctggcgcc agaagaacgc gaaaacctcc gcatagccct tagtgggtac
      721 ctcaagagac gggggtacgc aaggacggag gccgaaaccg acacttccgt cctcgaatct
      781 ttggatcctt ccgtattctc atctgcccca agttttacta acttcttcaa tgactcagag
      841 cccctcaata tccagtggga agccatcgcc aactcaccag aaactacaaa ggcactcaac
      901 aaggagctca gcggtcagaa agaggctgat tttaaaaaat atatcaagac atctttccca
      961 gagtactccg ccaaggaaat cttggctaat tacgtcgagg gtaggcgagc aatcctcgac
     1021 gccagcaagt acattgcgaa ccttcagagt ctggggcaca aacacagaag caagtacctg
     1081 agcgatattc ttcaagacat gaaaagggat agccggatta cccgcctttc tgaagccttt
     1141 ggctctaccg acaatctctg gagaataatt ggcaatatta gtaacttgca ggagcgggcg
     1201 gtacggtggt atttcaatga cgcaaaattt gaacaggggc aggaacagct ggatgccgtc
     1261 aagctgaaga acgtactcgt ccgggccctg aagtatctga gaagcgacga taaggaatgg
     1321 agcgcgagcc aaaagcagat catccagtca ctggaacagt ccggggacgt tctggacgtg
     1381 ctggctgggc tggatccaga caggacaatc cctccttacg aagaccaaaa caatcggcgt
     1441 cccccagaag accagacact gtaccttaac cccaaagctt tgtcttccga gtatggagaa
     1501 aagtggaagt cttgggcaaa caagtttgct ggcgcatatc ccctgcttac ggaggatctc
     1561 accgagattc tcaagaatac ggacaggaag agccggatca agatccgcag cgatgtgctc
     1621 cccgactctg attaccggtt ggcgtacatc ctccaacgag cttttgatcg gtccatcgcc
     1681 ctggatgaat gctctatcag gcgaactgca gaggacttcg agaacggggt cgtaatcaag
     1741 aatgagaaat tggaggacgt gttgtccggc caccagctgg aggagttcct ggaatttgcc
     1801 aataggtatt accaagaaac cgctaaggcg aagaacgggc tctggtttcc agaaaacgct
     1861 cttctcgaac gcgccgatct gcatccccct atgaagaata agattctgaa tgttatagtg
     1921 gggcaggctc tcggagtttc accggcggag ggtacagatt ttatcgaaga aatctggaat
     1981 agcaaggtta aaggccgatc taccgtccgc agcatctgta acgcaatcga aaatgagcga
     2041 aaaacctacg gtccctactt cagcgaggac tacaagttcg tcaagaccgc cctgaaggaa
     2101 ggaaagacag aaaaggaact ctccaagaaa ttcgccgcag tgatcaaagt gctgaagatg
     2161 gtgtcagagg tggtgccctt catcgggaag gagctccggc tctccgacga agcccagtcc
     2221 aaattcgata acctgtattc tctggcccag ctgtacaatc tgatcgaaac cgaaaggaac
     2281 gggttctcca aagtgtctct cgctgcccac ctggagaacg catggcggat gaccatgaca
     2341 gatgggtccg cacaatgttg taggctgccc gctgattgtg tgcgaccatt cgatggtttt
     2401 atcaggaagg cgatcgacag aaattcctgg gaagtggcta agcggatagc cgaggaagtg
     2461 aagaagagcg ttgatttcac caatgggaca gtcaaaatcc ccgttgcaat cgaggctaac
     2521 agcttcaact ttaccgcttc actgactgat ctgaagtaca tacagcttaa ggagcaaaag
     2581 ctcaagaaga aactggagga catccaacgg aacgaagaaa atcaggagaa aaggtggctc
     2641 tcaaaggagg agcgcatacg ggccgacagc cacgggattt gcgcatacac cggacggccc
     2701 cttgatgacg tgggtgagat tgaccacatt atcccaaggt ccctcacact gaaaaaatca
     2761 gagtcaatct ataattcaga ggtcaatttg atattcgtgt cagcccaggg gaatcaggaa
     2821 aagaagaata atatctatct cctcagtaac cttgccaaaa attatctggc cgccgttttc
     2881 ggtactagcg acctttctca gatcacgaat gagatagaga gcaccgtgct ccagctgaaa
     2941 gccgcagggc gccttggata tttcgatttg ttgagcgaga aagaacgcgc atgcgcacgg
     3001 cacgcacttt tcctgaatag cgactccgaa gcgagaaggg ctgtgatcga cgtcctgggt
     3061 tcccgccgca aggccagcgt taatggaacc caggcatggt ttgtgcgcag catcttcagc
     3121 aaggttagac aggccctggc tgcctggact caagaaaccg gtaacgaact gatcttcgac
     3181 gcaatatccg tgcctgcagc agactccagc gagatgcgca agagattcgc tgagtatcga
     3241 ccagagtttc ggaagcctaa agtacaaccc gttgcttctc atagcattga tgccatgtgt
     3301 atctatctgg cggcttgtag cgaccctttt aaaaccaagc gcatggggtc ccagcttgcc
     3361 atatacgaac caattaactt cgacaacctg ttcactggaa gttgccaggt tattcagaat
     3421 acacccagaa atttcagcga taagaccaac atcgctaata gccccatttt taaagaaacc
     3481 atctacgcgg agcggttcct ggatattatt gtgagccggg gtgaaatctt catcggatac
     3541 ccgtctaaca tgccatttga ggagaagcct aacagaattt ccatcggagg gaaagatcct
     3601 ttcagtattc tctctgtgct gggtgcgtat ctggataagg caccttctag cgagaaagag
     3661 aagctcacaa tatacagagt ggtgaagaac aaagccttcg agctctttag caaggtggcc
     3721 ggaagtaaat tcacggccga ggaggacaag gctgctaaga tcctggaggc actccacttc
     3781 gtgacagtga agcaggacgt ggctgcaaca gtttccgatc tgatcaaatc caagaaggaa
     3841 ttgtctaaag attctataga gaatttggcc aagcaaaaag ggtgccttaa gaaggtcgaa
     3901 tacagtagca aggagttcaa atttaagggg agtctgatta ttccagccgc agtggagtgg
     3961 ggaaaggtac tttggaacgt gtttaaggag aacaccgcgg aagagctgaa ggatgagaat
     4021 gccctgcgca aagcccttga agctgcctgg ccgtcttctt tcgggactag gaatttgcac
     4081 agcaaggcta aacgggtatt ctctctccct gtggtggcta ctcagtccgg agctgtgcgc
     4141 atccggcgca aaacagcctt cggagacttt gtgtaccagt cacaggatac aaataacctc
     4201 tattcatcct ttccggtcaa aaatggcaaa ctggactggt ccagtccaat tatccatccc
     4261 gccctgcaga accggaatct gaccgcttat ggataccgat ttgtagatca cgacagaagc
     4321 attagcatgt ccgagttccg cgaagtctat aataaagacg atttgatgag gatcgaactg
     4381 gctcagggca cctcttccag gcgctacctg cgggtggaaa tgcccggcga gaaattcctg
     4441 gcctggtttg gggagaattc catttccctc gggtcctcct tcaagtttag cgtgtcagaa
     4501 gtcttcgata acaaaatata taccgaaaac gctgaattca caaagttttt gcccaagccg
     4561 agggaagaca ataaacataa cggcacaatc ttctttgaac tggtggggcc tcgagtcatc
     4621 ttcaattaca tcgtgggagg ggctgcctca tcacttaaag agatttttag cgaagccggt
     4681 aaagagcggt ccagcagggc tgaccccaag aagaagagga aggtgaggtc catggtcata
     4741 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     4801 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     4861 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     4921 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     4981 gctgcgctcg gtcgttcggc tgcgtcttcc gcttcctcgc tcactgactc gctgcgctcg
     5041 gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca
     5101 gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac
     5161 cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac
     5221 aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg
     5281 tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac
     5341 ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat
     5401 ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag
     5461 cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac
     5521 ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt
     5581 gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt
     5641 atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc
     5701 aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga
     5761 aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac
     5821 gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc
     5881 cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct
     5941 gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca
     6001 tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct
     6061 ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca
     6121 ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc
     6181 atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg
     6241 cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct
     6301 tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa
     6361 aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta
     6421 tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc
     6481 ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg
     6541 agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa
     6601 gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg
     6661 agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc
     6721 accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg
     6781 gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat
     6841 cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata
     6901 ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tctaagaaac cattattatc
     6961 atgacattaa cctataaaaa taggcgtatc acgaggccct ttcgtctcgc gcgtttcggt
     7021 gatgacggtg aaaac
//
