LOCUS       NmeN7A.1_Cas9           8180 bp    ds-DNA  circular SYN 23-NOV-2025
DEFINITION  Protein2PAM NmeN7A.1_Cas9 plasmid.
ACCESSION   NmeN7A.1_Cas9
VERSION     NmeN7A.1_Cas9
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
            .
FEATURES             Location/Qualifiers
     enhancer        1..304
                     /label="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        306..508
                     /label="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate earlypromoter"
     CDS             3879..3899
                     /label="SV40 NLS"
                     /product="nuclear localization signal of SV40 (simian
                     virus40) large T antigen"
                     /translation="PKKKRKV"
     CDS             3912..3977
                     /label="3xFLAG"
                     /product="three tandem FLAG(R) epitope tags, followed by
                     anenterokinase cleavage site"
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3996..4052
                     /label="P2A"
                     /note="Eukaryotic ribosomes fail to insert a peptide bond
                     between the Gly and Pro residues, yielding separate
                     polypeptides."
                     /product="2A peptide from porcine teschovirus-1
                     polyprotein"
                     /translation="ATNFSLLKQAGDVEENPGP"
     rep_origin      complement(5417..6005)
                     /label="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
                     replication"
                     /direction=LEFT
     CDS             6751..7611
                     /label="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin,
                     andrelated antibiotics"
                     /gene="bla"
                     /product="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW*"
     CDS             579..3824
                     /label="NmeN7A.1_Cas9"
                     /gene="cas9"
                     /product="NmeN7A.1_Cas9 variant"
                     /note="full ORF CDS for engineered NmeN7A.1_Cas9 variant"
                     /translation="MAAFKPNSINYILGLDIGIASVGWAMVEIDEEENPIRLIDLGVRV
                     FERAEVPKTGDSLAMARRLARSVRRLTRRRAHRLLRTRRLLKREGVLQAANFDENGLIK
                     SLPNTPWQLRAAALDRKLTPLEWSAVLLHLIKHRGYLSQRKNEGETADKELGALLKGVA
                     GNAHALQTGDFRTPAELALNKFEKESGHIRNQRSDYSHTFSRKDLQAELILLFEKQKEF
                     GNPHVSGGLKEGIETLLMTQRPALSGDAVQKMLGHCTFEPAEPKAAKNTYTAERFIWLT
                     KLNNLRILEQGSERPLTDTERATLMDEPYRKSKLTYAQARKLLGLEDTAFFKGLRYGKD
                     NAEASTLMEMKAYHAISRALEKEGLKDKKSPLNLSPELQDEIGTAFSLFKTDEDITGRL
                     KDRIQPEILEALLKHISFDKFVQISLKALRRIVPLMEQGKRYDEACAEIYGDHYGKKNT
                     EEKIYLPPIPADEIRNPVVLRALSQARKVINGVVRRYGSPARIHIETAREVGKSFKDRK
                     EIEKRQEENRKDREKAAAKFREYFPNFVGEPKSKDILKLRLYEQQHGKCLYSGKEINLG
                     RLNEKGYVEIDHALPFSRTWDDSFNNKVLVLGSENQNKGNQTPYEYFNGKDNSREWQEF
                     KARVETSRFPRSKKQRILLQKFDEDGFKERNLNDTRYVNRFLCQFVADRMRLTGKGKKR
                     VFASNGQITNLLRGFWGLRKVRAENDRHHALDAVVVACSTVAMQQKITRFVRYKEMNAF
                     DGKTIDKETGEVLHQKTHFPQPWEFFAQEVMIRVFGKPDGKPEFEEADTLEKLRTLLAE
                     KLSSRPEAVHEYVTPLFVSRAPNRKMSGQGHMETVKSAKRLDEGVSVLRVPLTQLKLKD
                     LEKMVNREREPKLYEALKARLEAHKDDPAKAFAEPFYKYDKAGNRTQQVKAVRVEQVQK
                     TGVWVRNHNGIADNATMVRVDVFEKAGKYYLVPIYSWQVAKGILPDRAIIAYKDEEDWQ
                     LIDDSFNFKFSLHPNDLIKVTTKKKSRFAYYASCDRANGAINIRIHDLDHKIGKNGILE
                     GIGVKTALSFQKYQIDELGKEIRPCRLKKRPPVR"
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 catgctgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatca gatctttgtc
      541 gatcctacca tccactcgac acacccgcca gcggccgcat ggcagcattc aagccaaact
      601 caatcaatta catcctggga ctggacatcg gcatcgcatc cgtcgggtgg gctatggtcg
      661 aaatcgacga ggaggagaac cccatccgcc tgatcgatct gggcgtgcgc gtgtttgaga
      721 gggcagaggt gcctaagacc ggcgacagcc tggccatggc acggagactg gcacgctccg
      781 tgaggcgcct gacccggaga agggcccaca gactgctgag gacacgccgg ctgctgaaga
      841 gggagggcgt gctgcaggcc gccaacttcg atgagaatgg cctgatcaag tccctgccca
      901 ataccccttg gcagctgagg gcagccgccc tggaccgcaa gctgacacct ctggagtggt
      961 ccgccgtgct gctgcacctg atcaagcacc ggggctacct gtctcagaga aagaacgagg
     1021 gcgagacagc cgataaggag ctgggcgccc tgctgaaggg agtggcagga aatgcacacg
     1081 ccctgcagac cggcgacttt cgcacaccag ccgagctggc cctgaacaag ttcgagaagg
     1141 agagcggcca catccgcaat cagcggtctg actatagcca caccttctcc cggaaggatc
     1201 tgcaggccga gctgatcctg ctgtttgaga agcagaagga gttcggcaac ccacacgtgt
     1261 ctggcggcct gaaggagggc atcgagacac tgctgatgac acagcggccc gccctgagcg
     1321 gcgacgcagt gcagaagatg ctgggacact gcacctttga gccagccgag cccaaggccg
     1381 ccaagaatac ctacacagcc gagcggttca tctggctgac aaagctgaac aatctgagga
     1441 tcctggagca gggaagcgag cgcccactga ccgacacaga gagggccacc ctgatggatg
     1501 agccctaccg caagtccaag ctgacatatg cacaggcaag gaagctgctg ggcctggagg
     1561 acaccgcctt ctttaagggc ctgagatacg gcaaggataa cgccgaggcc tctacactga
     1621 tggagatgaa ggcctatcac gccatcagca gggccctgga gaaggagggc ctgaaggaca
     1681 agaagtcccc actgaatctg tctcccgagc tgcaggatga gatcggcacc gcctttagcc
     1741 tgttcaagac cgacgaggat atcacaggca gactgaagga caggatccag ccagagatcc
     1801 tggaggccct gctgaagcac atcagctttg ataagttcgt gcagatcagc ctgaaggccc
     1861 tgcggaggat cgtgccactg atggagcagg gcaagaggta cgacgaggcc tgcgccgaaa
     1921 tctacggcga tcactatggc aagaagaaca cagaggagaa aatctacctg ccccctatcc
     1981 ccgccgatga gatcaggaac cctgtggtgc tgcgcgccct gtctcaggca agaaaagtga
     2041 tcaacggagt ggtgcgccgg tacggcagcc ccgccagaat ccacatcgag acagccaggg
     2101 aagtgggcaa gtcctttaag gacagaaagg agatcgagaa gaggcaggag gagaacagaa
     2161 aggataggga gaaggccgcc gccaagttca gagagtactt tcctaatttc gtgggcgagc
     2221 caaagtccaa ggatatcctg aagctgaggc tgtacgagca gcagcacggc aagtgtctgt
     2281 attctggcaa ggagatcaac ctgggccgcc tgaatgagaa gggctatgtg gagatcgacc
     2341 acgccctgcc tttttctcgg acctgggacg atagcttcaa caataaggtg ctggtgctgg
     2401 gctctgagaa ccagaataag ggcaaccaga caccctacga gtatttcaac ggcaaggaca
     2461 atagccgcga gtggcaggag tttaaggcaa gggtggagac aagcaggttc cctcggtcca
     2521 agaagcagag aatcctgctg cagaagtttg acgaggatgg cttcaaggag aggaacctga
     2581 atgacacccg ctacgtgaat cggtttctgt gccagttcgt ggccgataga atgaggctga
     2641 ccggcaaggg caagaagaga gtgtttgcct ccaacggcca gatcacaaat ctgctgaggg
     2701 gcttctgggg cctgagaaag gtgagggcag agaacgacag gcaccacgca ctggatgcag
     2761 tggtggtggc atgttctacc gtggccatgc agcagaagat cacacgcttt gtgcggtata
     2821 aggagatgaa tgccttcgac ggcaagacca tcgataagga gacaggcgag gtgctgcacc
     2881 agaagacaca ctttcctcag ccatgggagt tctttgccca ggaagtgatg atccgggtgt
     2941 ttggcaagcc tgacggcaag ccagagttcg aggaggccga taccctggag aagctgagaa
     3001 cactgctggc agagaagctg agctccaggc ccgaggcagt gcacgagtac gtgaccccac
     3061 tgttcgtgtc tagagccccc aacaggaaga tgagcggcca gggccacatg gagacagtga
     3121 agtccgccaa gagactggac gagggcgtgt ctgtgctgag ggtgcctctg acacagctga
     3181 agctgaagga tctggagaag atggtgaatc gcgagcggga gccaaagctg tatgaggccc
     3241 tgaaggcaag gctggaggca cacaaggacg atcctgccaa ggcctttgcc gagccattct
     3301 acaagtatga taaggccggc aacagaaccc aacaggtgaa ggccgtgagg gtggaacagg
     3361 tgcagaagac aggcgtgtgg gtccgcaacc acaatggcat cgccgacaat gctaccatgg
     3421 tgcgggtgga cgtgtttgag aaggccggta agtactatct ggtgcccatc tacagctggc
     3481 aagtggccaa gggcatcctg cctgatagag ccatcatagc gtacaaggac gaggaggatt
     3541 ggcagctgat cgacgattcc ttcaacttta agttctctct gcaccccaat gacctgatca
     3601 aagtgactac caagaagaaa tctcgatttg cctactacgc ctcctgcgac cgcgccaacg
     3661 gcgcaatcaa tatccggatc cacgacctgg atcacaagat cggcaagaac ggcatcctgg
     3721 agggcatcgg cgtgaagaca gccctgtcct tccagaagta tcagatcgac gagctgggca
     3781 aggagatcag accttgtagg ctgaagaagc gcccacccgt gcggggatcc ggcggaggcg
     3841 gaagcgggaa aagaaccgcc gacggcagcg aattcgagcc caagaagaag aggaaagtct
     3901 cgagcggagg cgactacaaa gaccatgacg gtgattataa agatcatgac atcgattaca
     3961 aggatgacga tgacaaggga ggcggtggaa gcggcgcaac aaacttctct ctgctgaaac
     4021 aagccggaga tgtcgaagag aatcctggac cgatggtgag caagggcgag gagctgttca
     4081 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
     4141 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
     4201 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     4261 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     4321 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     4381 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     4441 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     4501 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     4561 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     4621 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     4681 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     4741 tcactctcgg catggacgag ctgtacaagt gaccggtcat catcaccatc accatggatc
     4801 cgcaggcctc tgctagcttg actgactgag atacagcgta ccttcagctc acagacatga
     4861 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     4921 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     4981 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg tgggaggttt
     5041 tttaaagcaa gtaaaacctc tacaaatgtg gtattggccc atctctatcg gtatcgcgat
     5101 ctctcgattt cgatcaagac attcctttaa tggtcttttc tggacaccac taggggtcag
     5161 aagtagttca tcaaactttc ttccctccct aatctcattg gttaccttgg gctatcgaaa
     5221 cttaattaac cagtcaagtc agctacttgg cgagatcgac ttgtctgggt ttcgactacg
     5281 ctcagaattg cgtcagtcaa gttcgatctg gtccttgcta ttgcacccgt tctccgatta
     5341 cgagtttcat ttaaatcatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     5401 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     5461 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     5521 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     5581 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     5641 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     5701 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     5761 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     5821 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
     5881 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     5941 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     6001 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     6061 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     6121 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     6181 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     6241 catttaaatt tccgaactct ccaaggccct cgtcggaaaa tcttcaaacc tttcgtccga
     6301 tccatcttgc aggctacctc tcgaacgaac tatcgcaagt ctcttggccg gccttgcgcc
     6361 ttggctattg cttggcagcg cctatcgcca ggtattactc caatcccgaa tatccgagat
     6421 cgggatcacc cgagagaagt tcaacctaca tcctcaatcc cgatctatcc gagatccgag
     6481 gaatatcgaa atcggggcgc gcctggtgta ccgagaacga tcctctcagt gcgagtctcg
     6541 acgatccata tcgttgcttg gcagtcagcc agtcggaatc cagcttggga cccaggaagt
     6601 ccaatcgtca gatattgtac tcaagcctgg tcacggcagc gtaccgatct gtttaaacct
     6661 agatattgat agtctgatcg gtcaacgtat aatcgagtcc tagcttttgc aaacatctat
     6721 caagagacag gatcagcagg aggctttcgc atgagtattc aacatttccg tgtcgccctt
     6781 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     6841 gtaaaagatg ctgaagatca gttgggtgcg cgagtgggtt acatcgaact ggatctcaac
     6901 agcggtaaga tccttgagag ttttcgcccc gaagaacgct ttccaatgat gagcactttt
     6961 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     7021 cgccgcatac actattctca gaatgacttg gttgagtatt caccagtcac agaaaagcat
     7081 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     7141 actgcggcca acttacttct gacaacgatt ggaggaccga aggagctaac cgcttttttg
     7201 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     7261 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac cttgcgtaaa
     7321 ctattaactg gcgaactact tactctagct tcccggcaac agttgataga ctggatggag
     7381 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     7441 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     7501 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     7561 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta accgattcta
     7621 ggtgcattgg cgcagaaaaa aatgcctgat gcgacgctgc gcgtcttata ctcccacata
     7681 tgccagattc agcaacggat acggcttccc caacttgccc acttccatac gtgtcctcct
     7741 taccagaaat ttatccttaa gatcccgaat cgtttaaact cgactctggc tctatcgaat
     7801 ctccgtcgtt tcgagcttac gcgaacagcc gtggcgctca tttgctcgtc gggcatcgaa
     7861 tctcgtcagc tatcgtcagc ttaccttttt ggcagcgatc gcggctcccg acatcttgga
     7921 ccattagctc cacaggtatc ttcttccctc tagtggtcat aacagcagct tcagctacct
     7981 ctcaattcaa aaaacccctc aagacccgtt tagaggcccc aaggggttat gctatcaatc
     8041 gttgcgttac acacacaaaa aaccaacaca catccatctt cgatggatag cgattttatt
     8101 atctaactgc tgatcgagtg tagccagatc tagtaatcaa ttacggggtc attagttcat
     8161 agcccatata tggagttccg
//
