LOCUS       sequence-209052-        6335 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 6335)
  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 6335)
  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-B2KB46
FEATURES             Location/Qualifiers
     source          1..6335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(183..771)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(263..282)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(942..1733,1734..1802))
                     /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     1565..1584
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1803..1907)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1975..1993
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2031..2053)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2153..2172
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2366..2388
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2380..2397
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2381..2397
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             6036..6056
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(6066..6082)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6066..6082)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6079..6101)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6090..6106
                     /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(6114..6120,6121..6138,6139..6144))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6159..6180
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta
       61 tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag
      121 aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg
      181 tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg
      241 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg
      301 cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga
      361 agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc
      421 tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt
      481 aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact
      541 ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg
      601 cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt
      661 accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt
      721 ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct
      781 ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg
      841 gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt
      901 aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt
      961 gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc
     1021 gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg
     1081 cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc
     1141 gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg
     1201 gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca
     1261 ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga
     1321 tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct
     1381 ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg
     1441 cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca
     1501 accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata
     1561 cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct
     1621 tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact
     1681 cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa
     1741 acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc
     1801 atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga
     1861 tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga
     1921 aaagtgccac ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg
     1981 cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac
     2041 atgcagctcc cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc
     2101 cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca
     2161 gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag atgcgtaagg
     2221 agaaaatacc gcatcaggcg ccattcgcca ttcaggctgc gcaactgttg ggaagggcga
     2281 tcggtgcggg cctcttcgct attacgccag ctggcgaaag ggggatgtgc tgcaaggcga
     2341 ttaagttggg taacgccagg gttttcccag tcacgacgtt gtaaaacgac ggccagtgtg
     2401 ctgtctcatc attttggcaa agaattcgcg gccgccacca tgcaaaagaa tatcaacaca
     2461 aaacagaacc acatttatat caaacaggcc cagaaaatca aagaaaagct cggagataag
     2521 ccctacagga ttggcttgga cctgggtgtg ggcagtattg ggtttgccat agtttccatg
     2581 gaagaaaacg acggtaacgt gctgcttcct aaggagatta tcatggtggg aagccgcatc
     2641 tttaaggcct ctgccggggc cgccgaccgg aaactcagca ggggccagag aaacaaccat
     2701 cgccacaccc gggagaggat gaggtatctt tggaaggtct tggctgaaca gaagcttgcc
     2761 ctgccagtcc cggccgacct cgatcgcaaa gagaacagca gtgaaggtga gacaagcgct
     2821 aaacgattct tgggagacgt cctgcaaaaa gatatttatg aactgcgcgt caagtctctg
     2881 gacgagaggt tgagcctcca ggagctggga tacgtgctgt atcacatcgc cggacaccgc
     2941 ggaagttcag ccatccgcac ctttgagaac gatagtgagg aggcccagaa ggaaaatact
     3001 gagaacaaga aaatcgccgg caatatcaag cgactgatgg cgaagaaaaa ctatcggaca
     3061 tacggcgagt atctgtataa ggagtttttc gaaaataaag aaaagcacaa acgggaaaaa
     3121 atctcaaacg cagcgaataa ccacaagttt agtcctaccc gagatctggt gattaaggaa
     3181 gctgaagcga ttctgaagaa acaggcgggg aaagacggct ttcacaagga gctcaccgag
     3241 gaatatatcg agaagctcac taaagctatc ggatatgaga gtgaaaaact catacctgaa
     3301 tccgggttct gcccatacct gaaggacgag aaacgactgc ctgcaagcca taagcttaac
     3361 gaagagagac gcctttggga gacacttaac aacgcaaggt actctgatcc catcgtggat
     3421 atcgtcactg gtgaaatcac aggatattac gagaagcagt ttaccaagga gcagaagcaa
     3481 aagcttttcg actacttgct gaccggatca gaactgaccc ctgcccagac taagaaactg
     3541 ctggggctca agaatactaa ctttgaagac attatactcc aaggaagaga taagaaagcc
     3601 cagaaaatca agggctacaa gcttatcaag ctcgagagta tgcctttttg ggcccgcctc
     3661 agtgaggcac agcaggactc attcctttat gactggaatt cctgcccaga tgagaaactg
     3721 ctgacagaga agttgtcaaa tgaataccac ctgacggagg aggagattga caacgcgttc
     3781 aatgagatag ttctgagctc cagctacgcc cctctgggca aaagtgcgat gctgatcatc
     3841 ctcgagaaaa tcaaaaatga tctgtcatac acggaagcag ttgaggaggc actgaaggaa
     3901 ggaaaactga cgaaggaaaa acaggccatt aaggatcggc tcccatatta tggagccgtg
     3961 cttcaggaat caacccagaa aattatagca aagggtttct ccccgcagtt taaagataag
     4021 gggtacaaga ctcctcacac gaacaagtac gagctggaat acggtagaat tgcgaaccca
     4081 gtcgtgcatc agactcttaa cgaactgcga aaacttgtaa atgagatcat cgacatcctg
     4141 ggcaagaagc cctgcgagat cgggctggaa accgccaggg aattgaagaa gagtgcagag
     4201 gatcggtcta aactgtctcg cgagcagaat gataacgagt ccaatagaaa tcgaatctat
     4261 gaaatctaca tccgacctca gcagcaggtt atcattacca ggcgggagaa ccctcggaac
     4321 tatattctta aattcgaact gctcgaggag caaaaatccc agtgcccttt ctgcggcggt
     4381 cagatcagcc caaatgacat tattaataac caggccgata tagagcacct ttttcccata
     4441 gcagaaagtg aagacaacgg caggaataat ctggtcatca gccacagcgc ctgtaacgcc
     4501 gacaaggcca agagatctcc gtgggcagct tttgcatccg ccgctaagga tagcaaatat
     4561 gattataaca gaattctctc caacgtcaag gagaacatcc ctcacaaagc gtggagattc
     4621 aaccagggag catttgagaa attcatagaa aataaaccta tggccgccag gtttaaaacc
     4681 gataacagct acatctcaaa ggtggcccac aagtacttgg cttgcctgtt cgaaaaacca
     4741 aatattattt gtgttaaggg atccctgact gcccaattga ggatggcttg gggcctgcag
     4801 ggccttatga tccctttcgc gaagcagctc atcacagaga aagagtccga aagtttcaat
     4861 aaagatgtca actccaacaa gaagattaga ctcgacaata gacaccacgc cttggacgct
     4921 attgtgattg cctacgcgtc ccggggctat ggcaatttgc tgaacaaaat ggcaggcaaa
     4981 gactataaaa ttaactactc agagcgcaat tggctgtcaa aaattctgct gcctccgaat
     5041 aatatcgtct gggagaatat tgacgctgat ctcgagagtt tcgagagctc cgtgaaaact
     5101 gcactgaaaa acgccttcat tagcgttaaa cacgatcata gtgacaacgg agagcttgtc
     5161 aagggtacaa tgtacaaaat cttctatagc gagcgcggct acaccctgac tacgtacaaa
     5221 aaacttagtg ccttgaagct caccgatcct cagaaaaaaa agacccccaa ggactttctt
     5281 gagacagcgc ttctgaaatt taagggccgg gagtccgaga tgaagaacga gaaaatcaag
     5341 agcgccattg aaaataacaa gaggctgttc gacgtcatac aagataatct ggagaaagca
     5401 aagaagcttt tggaggagga gaacgagaag agtaaggcgg agggtaaaaa ggagaagaat
     5461 atcaacgacg cgtccattta ccagaaggcg attagcctgt ctggcgacaa gtatgttcaa
     5521 ctctccaaga aggaacctgg gaagtttttt gcgattagta agcctactcc cactaccaca
     5581 ggatacgggt atgatacagg ggattcactt tgcgttgatc tctattacga taacaagggc
     5641 aaactgtgcg gagagatcat acggaaaata gatgctcagc agaaaaaccc tcttaaatac
     5701 aaagaacagg gcttcacctt gttcgaaagg atttacggcg gcgacatttt ggaagtggat
     5761 ttcgatattc atagcgacaa aaacagcttt agaaacaaca ctggcagcgc tccagagaac
     5821 cgcgtcttta tcaaagtcgg tacattcact gaaatcacta acaataatat ccagatctgg
     5881 tttgggaata tcataaaaag tacaggcggg caggacgatt cttttaccat taattccatg
     5941 cagcagtaca atcccaggaa actgatcctg agttcctgtg gtttcattaa atacagatct
     6001 ccaattctga agaacaagga gggcagcagg ctgaccccaa gaagaagagg aaggtgaggt
     6061 ccatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
     6121 cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta
     6181 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
     6241 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
     6301 ctcactgact cgctgcgctc ggtcgttcgg ctgcg
//
