LOCUS       sequence-201805-        5822 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 5822)
  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 5822)
  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-D3UFL8
FEATURES             Location/Qualifiers
     source          1..5822
                     /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             5523..5543
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5553..5569)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5553..5569)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5566..5588)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5577..5593
                     /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(5601..5607,5608..5625,5626..5631))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5646..5667
                     /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 atttggcaaa gaattcgcgg ccgccaccat gattcgaact ctgggcatcg
     2461 atatcggaat tgcctcaatt ggatgggccg tcattgaggg cgagtataca gataaggggc
     2521 tggaaaacaa ggaaatcgtt gcttctggcg tgcgggtgtt tacgaaggca gagaatccaa
     2581 agaataaaga gagcctcgcc ttgcctcgga ccctggcacg cagtgcaaga aggcggaacg
     2641 ctagaaagaa agggcggatc cagcaggtga aacactacct gtcaaaagcc cttggcctcg
     2701 acctcgagtg tttcgtgcag ggagagaagc tggccacttt gtttcagact tccaaggatt
     2761 ttctgagccc atgggaactt cgggaaagag cgttgtaccg ggtcctggat aaagaggagc
     2821 tggcccgagt aatactgcat attgccaaac gcaggggcta tgatgacatc acctatgggg
     2881 tcgaggataa tgactcaggc aagattaaga aggcaattgc cgaaaacagc aagaggataa
     2941 aagaagaaca atgtaagact atcggggaga tgatgtacaa gctttatttt cagaaaagtc
     3001 ttaatgtaag gaataagaaa gaaagttaca ataggtgcgt gggtagaagt gaacttagag
     3061 aagaactgaa gaccattttc caaattcagc aggagctgaa atccccttgg gtgaacgagg
     3121 aacttattta caaattgctt ggaaaccctg atgcccagtc caagcaggag cgggagggac
     3181 tgattttcta tcagcgccct cttaaaggct tcggcgacaa aattggaaaa tgcagtcaca
     3241 tcaagaaggg cgagaatagc ccctatcgcg cctgcaagca cgctccttcc gccgaggagt
     3301 tcgtggccct cacaaagtca atcaatttcc tcaagaatct tacaaaccgg catggcctct
     3361 gcttttcaca ggaggacatg tgcgtttacc tggggaaaat cctccaggag gcccaaaaga
     3421 acgagaaagg actgacatac tccaagctca agcttcttct ggatcttccc tctgattttg
     3481 agtttctggg cttggattat agtggcaaga accccgagaa ggccgttttc ctctccctgc
     3541 cctctacttt taagctgaat aaaatcaccc aagatcggaa aacacaagac aaaatcgcca
     3601 acatcctcgg cgcgaataaa gactgggagg ccatccttaa ggaactggaa tctctgcagt
     3661 tgtctaagga acagatccaa actattaagg acgccaaact caattttagc aagcacatta
     3721 atctgtccct cgaagcactc taccaccttc tccctctgat gcgcgaaggg aagagatatg
     3781 atgaaggcgt ggaaatcctg caggagcgcg ggatattctc caagcctcag cctaagaacc
     3841 gacagcttct ccctcctctg tcagagttgg ctaaggagga atcttatttt gatatcccta
     3901 atccagtgtt gcgcagggcc cttagtgagt tccgaaaggt ggtaaacgcc cttcttgaaa
     3961 agtacggagg gttccattat tttcacattg aattgacgcg cgatgtctgt aaggctaagt
     4021 cagcacgcat gcagctggaa aagataaaca agaagaacaa gtctgaaaat gacgccgcca
     4081 gtcagctgtt ggaggtcttg gggcttccca acacctacaa taatcgcttg aagtgtaagc
     4141 tgtggaaaca gcaggaagag tattgtctct actccggaga aaagattacc atagatcacc
     4201 tgaaggatca gcgcgccctc cagatcgatc acgcatttcc cctgagtcga tccctcgacg
     4261 actcacagtc aaacaaggtg ctgtgtctga cttcctcaaa tcaggagaag agcaataaga
     4321 ccccatacga atggctggga agcgacgaga agaagtggga catgtacgtt ggccgggtgt
     4381 actcctccaa cttctcccct tccaagaaga ggaagttgac gcagaagaat ttcaaagagc
     4441 gcaatgaaga agatttcctc gcccgcaact tggtcgacac cggttatatc ggccgagtta
     4501 ctaaggagta tattaaacat tccctcagtt tcctgccact gccggatggt aagaaagagc
     4561 atattaggat catttcaggc tccatgacat ctacaatgcg gtcattttgg ggtgttcagg
     4621 aaaagaatcg agatcaccac ttgcatcatg cacaggacgc aatcatcatt gcatgtatcg
     4681 agcccagcat gattcaaaag tataccacgt atctcaaaga taaggaaacg cacaggttga
     4741 agtcccacca gaaagcccag atactgcgag agggtgacca caaattgtca ctgaggtggc
     4801 cgatgagcaa tttcaaagat aaaatacagg agtccattca gaatatcatc ccatcacacc
     4861 acgtgagcca caaggtcacc ggtgaactcc accaggagac tgtgagaacc aaagagttct
     4921 actaccaagc ctttggaggg gaggaaggag tgaagaaagc actgaagttc ggaaagatca
     4981 gggagatcaa tcagggaatc gtagacaacg gggctatggt gcgggtggac atcttcaagt
     5041 ccaaggacaa ggggaaattc tacgcggttc cgatctacac atacgacttt gccattggca
     5101 agcttccaaa caaggcaatc gtgcaaggga agaagaacgg catcataaag gattggttgg
     5161 agatggatga gaactatgaa ttctgtttca gtttgttcaa gaatgattgc atcaagatac
     5221 agactaaaga gatgcaggag gccgtgcttg caatatataa gtcaacaaat agtgcaaaag
     5281 ctacaataga gctcgaacac ctgagtaagt atgccctgaa gaacgaagac gaggagaaaa
     5341 tgttcactga caccgacaag gagaagaata agaccatgac tcgggagagt tgcgggattc
     5401 aagggctcaa ggttttccag aaagtgaaac tctcagtgct gggtgaggtt ttggaacaca
     5461 aacctcgcaa cagacagaat atcgccctga agacaacacc caaacatgtc agcagggctg
     5521 accccaagaa gaagaggaag gtgaggtcca tggtcatagc tgtttcctgt gtgaaattgt
     5581 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     5641 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     5701 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     5761 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     5821 cg
//
