LOCUS       sequence-403617-        5729 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human expression vector for human codon optimized N-terminal 
            NLS-tagged SpuFz1-D487K.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5729)
  AUTHORS   Saito M, Xu P, Faure G, Maguire S, Kannan S, Altae-Tran H, Vo S, 
            Desimone A, Macrae RK, Zhang F
  TITLE     Fanzor is a eukaryotic programmable RNA-guided endonuclease.
  JOURNAL   Nature. 2023 Jun 28. doi: 10.1038/s41586-023-06356-2.
  PUBMED    37380027
REFERENCE   2  (bases 1 to 5729)
  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: "Nature. 
            2023 Jun 28. doi: 10.1038/s41586-023-06356-2."
COMMENT     Sequence Label: pMSSM58: pCMV-HA-NLS-SpuFz1-D487K
FEATURES             Location/Qualifiers
     source          1..5729
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             563..589
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             590..616
                     /codon_start=1
                     /product="nuclear localization signal (Makkerh et al., 
                     1996)"
                     /label=NLS
                     /translation="PAAKKKKLD"
     primer_bind     complement(2675..2694)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2740..2795
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2794..2813)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3156..3172)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3156..3172)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3169..3191)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3180..3196
                     /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(3204..3210,3211..3228,3229..3234))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3249..3270
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3329..3524
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3375..3510
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    3530..3664
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3567..3586)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3621..3640
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3732..3749)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     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
     enhancer        join(5658..5729,1..304)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atcgccacca tgtaccccta cgacgtgcca gactacgccc ctgcggcaaa
      601 aaagaaaaag ttggacggcg gctctggcgg cggaggatct ggcggaggtg gaagcggagg
      661 accgcctaag aagaagcaga agctggaacg gctgaagaaa ctggacaagc ccacactgca
      721 cacctgtaac aagaccagct tcgccaaggc cttcctgcct aacgagacat accggcagcg
      781 gctgctggac tatatcgcca tcattcacca gctggccgac cacgcctctc acgccctgaa
      841 gttttacatc ctgagcacct ccaccagcag cttccctgtg gtgcacgagg ataccatcga
      901 ggccatcctg tacctgctga acaagggcga agcctggcat cccagaaaag aggccaagaa
      961 ggcctggcgg gattgcctgc tgccttacgt gcagagatac tgccagatcg tgggcttcat
     1021 tcaccccaac ctgagaggcg agcagcagag catcaactac ctgaccgtgt ccatgatgac
     1081 caacctgaaa gtgaacgtgc aagagcactt catgcagatg ctgctgcggt acatcaatct
     1141 gcgcttcgac gtgaagggcc agaagcagag actgcctcct aagagcgacg cccgcaaggc
     1201 ctttttcacc cggctgagat acctgaagtc cgtgttcctg ttcgacgtgg tgcccgagct
     1261 ggaattcctg gacgatctga cccctctgga aagcgaggtg ctggaagaga tctggtccct
     1321 ggacctgcct tttctgccca acgatcctct ggcctacgcc atcgtggccg atcctatgag
     1381 cttcttccca gcctactgca agctgagcgg cctgtacgag cagtacggct tccagcggtt
     1441 tagcgccatt cctctgcgga gaagcctgat ccagtctcac gtgcggatcg acaccatcat
     1501 cctgtatcag cacatcctgt gcatcaccag acgggacgcc gagacagtgg aaaaggacga
     1561 cctgtggatg agagtgtgca acctgtgcac caaagccttc cggtccagat gcggcatgca
     1621 cttcgagggc agcatcacaa cagatggcgc cagcgtgtcc gtgtacctga aacaccctga
     1681 ggccgataag tacggcaaga gaggcgccag aaagtccgcc aatactgtgg ccgccgaagt
     1741 gaaggccctg tacgtggaaa acaacctgcc tgcctgtaga gccgccgaga acgtggtggt
     1801 catcgacccc aacaagcggg atatcctgta ctgccaggac agcaacggca ccaccttcag
     1861 atacaccgcc aaccagagag ccgtggaaac cggcagcaga agattcgcca agcggcggga
     1921 agccatgaag gaagaagccg gcgttgacct gatcgagagc agaatcccta gccacaagac
     1981 catgaacctg atggacttca ccagatacct gctcgtgcgg agagccgact gggacagacg
     2041 gaaagagttc tactctcacc ccgctcacac ccggtggaag tggcacagct tcatcaaccg
     2101 gcagaagtcc gagagcaagc tgatcagcaa catgcggaac aaatacggcg agaacttcac
     2161 cgtggtcatg ggcgattgga gcgacgctgg aagaaccgcc agattccaga ccagcagcaa
     2221 gaccaaaggc tggcggaccc tgttcaagcg gaaccggatc gattgcttcc tgctggacga
     2281 gtacaagacc tccagcgtgt gccctagatg cagcagcagc gagttcgtgg aaaagaagtt
     2341 caagacccgg cctcacagca gaccctggcg aagaagagag ggcaagatcg agaaggtgca
     2401 cggcctgctg ggctgcacca atcctaattg tctgcagcag gcctggacca gcggcatgag
     2461 atactggaac cgggacatgc tgagcacatg caacatgctg ctgatcgtgc ggtccatgct
     2521 ggatggccat ggcagacctg aggtgttctc tagatctgtg cctgccgtgg cctgactagc
     2581 ataacccctt ggggcgcact cctcaggtgc aggctgccta tcagaaggtg gtggctggtg
     2641 tggccaatgc cctggctcac aaataccact gagatctttt tccctctgcc aaaaattatg
     2701 gggacatcat gaagcccctt gagcatctga cttctggcta ataaaggaaa tttattttca
     2761 ttgcaatagt gtgttggaat tttttgtgtc tctcactcgg aaggacatat gggagggcaa
     2821 atcatttaaa acatcagaat gagtatttgg tttagagttt ggcaacatat gcccatatgc
     2881 tggctgccat gaacaaaggt tggctataaa gaggtcatca gtatatgaaa cagccccctg
     2941 ctgtccattc cttattccat agaaaagcct tgacttgagg ttagattttt tttatatttt
     3001 gttttgtgtt atttttttct ttaacatccc taaaattttc cttacatgtt ttactagcca
     3061 gatttttcct cctctcctga ctactcccag tcatagctgt ccctcttctc ttatggagat
     3121 ccctcgacct gcagcccaag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat
     3181 tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg
     3241 ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag
     3301 tcgggaaacc tgtcgtgcca gcggatccgc atctcaatta gtcagcaacc atagtcccgc
     3361 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3421 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctcggcctct gagctattcc
     3481 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagcta acttgtttat
     3541 tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt
     3601 tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg
     3661 gatccgctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc
     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 ctgggtcgac attgattatt gactagtaat agtaatcaat tacggggtca
     5701 ttagttcata gcccatatat ggagttccg
//
