LOCUS       sequence-332821-        8898 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian Expression, sauCas9 based prime editor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8898)
  AUTHORS   Liu P, Liang SQ, Zheng C, Mintzer E, Zhao YG, Ponnienselvan K, Mir 
            A, Sontheimer EJ, Gao G, Flotte TR, Wolfe SA, Xue W
  TITLE     Improved prime editors enable pathogenic allele correction and 
            cancer modelling in adult mice.
  JOURNAL   Nat Commun. 2021 Apr 9;12(1):2121. doi: 10.1038/s41467-021-22295-w.
  PUBMED    33837189
REFERENCE   2  (bases 1 to 8898)
  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: "Nat 
            Commun."; date: "2021-04-9"; volume: "12(1):2121. doi"; pages: " 
            10.1038/s41467-021-22295-w"
COMMENT     Sequence Label: SauCas9 PE2*
FEATURES             Location/Qualifiers
     source          1..8898
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(5317..5337)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(5377..5403)
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     regulatory      5406..5415
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5456..5659)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5485..5505)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5660..6039
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     6211..6230
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        6309..6413
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6414..6482,6483..7274)
                     /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     complement(6632..6651)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7445..8033
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7934..7953
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8187..8204
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    8321..8342
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(8357..8362,8363..8380,8381..8387)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8395..8411
                     /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)."
     primer_bind     8400..8422
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     8419..8435
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     8419..8435
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    8506..8730
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     8719..8736
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(8759..8776)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(8785..8805)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 gccagagggt gatgaatttt ctatgaggag ggtagaggtg tctggagtct ctgtgatggc
       61 tgcctttcgg gccgcttggt cagccatccg gttgcctcta gcctcggcgc tgtgtccctt
      121 ttgatgtcct ggacaatgga ttatgctaag tcttttgggc agaaagaggg cttttagtag
      181 ggccaagatc tcgtctttat ttttgatctc tttgccttct gatgtgagcc acccacgcct
      241 tctgtatatt tctccatgga tatgggcagt agcaaaagca taacggctat cagtataaac
      301 atttagcttc ttaccttctg ccatctttag ggcctgggtg agtgctatca gttcagcccg
      361 ctgagcggat gtcccggctg gcagggcttt agcccagatt acctcggtct cggtggtcac
      421 cgcagctccc gccttacgct gtccctcttg taagagactg cttccatccg tgtaccaggt
      481 gtggtcggcg tctgggagcg gctggtccgt taggtcgggt cgggttccgt gggcttcggc
      541 caggatatca aggcagttgt gttgcagccc ttcctcaggc agtgggagca gcgtagccgg
      601 gttcagggct accaccggtc cgaactggac ccggtccgtg tccaaaagca aggcctgata
      661 gtgagtcatc cgggcgttgg aaagccagcg gtcggggggt tgtttgacta gtgcctctac
      721 tgcatggggg gccagaatga ctagtggctg tcccatggtt agcttgcctg catcctttgt
      781 cagtacggca atggctgcta ccatccgtag gcaagggggc cacccagctg ctactgggtc
      841 tagctttttg gacaggtagg ccaccggccg acgccaaggt cccagttttt gcgttaggac
      901 acctttggcg tagccctgct tctcgtcgac aaagagttca aagggcttag tcaaatctgg
      961 caaccccagg gctggggcag ttagaagagc ttgcttgatt tcttgatagg ccttttgttg
     1021 gtctgggccc caattaaaca gagtccccgg tttggtgaga gggtacaggg gggctgccat
     1081 ttctgcaaac ccagggatga agaggcgaca gaagcctgcc ttccctagga actcccttag
     1141 ttgtcgaggg gtcttcggag taggctgccc catcacagtc tcttttctgg cctcagtcag
     1201 ccatctctga ccctctttta gaagataccc cagatacttg acctgtttct ggcaaatttg
     1261 ggctttcttg gccgaggccc gatacccgag gttccctagg gtttgtaaca gggcccgagt
     1321 accttgttgg cagtctagct cagaagtggc ggccagcagt aagtcatcca cgtactgtag
     1381 caggatcaag tctgggtgct ggatccggaa gtctgctagg tctctgtgca gtgcctcatt
     1441 aaacagggtg ggactgtttt tgaaaccctg tgggagtctg gtccaggtca attgtcctga
     1501 gattcccatc tctggatctc tccactcaaa ggcgaagaga ggctgactgg tggggtggag
     1561 tctcaggcag aaaaaggcat cctttaaatc aagcacagtg taccactggt gggacggtgg
     1621 gagcccgctc aagaggttgt aagggttggg cacggtgggg tggatgtctt ccacccgctt
     1681 gttgacttct ctcagatcct ggacaggcct ataatcatta gtccctggtt tcttaacggg
     1741 tagcaggggc gtgttccagg gggactggca gggtaccagt attccctggt ccaacagtct
     1801 ctgtatgtgg ggcttgatcc ccagtctggc ttcttgtgac atggggtatt gttttatgga
     1861 cacgggggta gaggttgctt tcagaggtat gatcagagga gcttggcgaa ctgccagtcc
     1921 catgcccccg gtttccgccc aggcctgagg aaaatcagac agccatgtgg accctagaga
     1981 aacatctggc tcttttgagg tctcatgtag ccgatactca tcttctatat ttagggtgct
     2041 gctgccgccg ctgctgccgc cactgctctc tggtgttgct gactcgcttg ttcctggtgt
     2101 ctcgctgcca gaggatcctc cgctagatcc tccagagccc tttttgatga tctgagggtg
     2161 cttcttagat ttcacttcat acaggttgcc cagaatgtct gtgctgtact tcttaatgct
     2221 ctgggtcttg gaggcgattg tcttaatgat cctggggggc ctcttgtcgt tcatgttttc
     2281 caggtactcg cggtaggtga tgtcgatcat gttcacttcg atccggttca gcaggtcgtt
     2341 gttcacgccg atcactctat acagctcgcc gttgatcttg atcagatcgt tgttgtagaa
     2401 ggaggcgata aactcggcct ggttgctgat cttcttcagc ttcttagctt cctcatagca
     2461 cttgctattc acttcgtagt agttttcttt tttgatcaca tccagattct tcacggtcac
     2521 gaacttgtac acgccattgt ccaggtacac gtcgaatctg tagggcttca gggacagctt
     2581 cacgaccttg tttctgctgt tggggtagtc gtcggtgatg tccagatggg cgttcagttt
     2641 gttgccgtaa tacttaatct tcttgatcac ggggccgttg tcctttttgg agtacttggt
     2701 caggtagttc ccggtttcct cgtagtactt gtacagggga ttcttctcgt cgccgtactg
     2761 ttccataatc agcttcagtt tctggtaggt ctgggggtcg tggtggtaca tcagcagctt
     2821 ttcggggctc ttgttgatca gctttttcag cttgtcattg tccttgtcgt acaggccgtt
     2881 cagattgttc acgatcaggg tgttgccctt gtcgtccttc cgggtggagt acagggtgtc
     2941 gttaatcagc tctctattag gcttcttgtc cacccggtgg ctgtacttgt agtccttgaa
     3001 gtccttaatg tgcttgatct ggtggggggt gatgaagatc tctttgtact cctgctcggt
     3061 ttcgatctcg ggcatgctct cggcctgctt ttcctcgaac atctggtttt ccatcacttt
     3121 tttggccttg tccagtttct tccactcttt gaagatgaaa tcggcgttgg caatgatcag
     3181 ggcgtcctcg gcgtggtgct tgtacccctt gttccgctct ttcttaaact tccacttccg
     3241 ccgcagaaag ctggtgaagc cgccattgat ggacttcact ttcacgtcca ggttgttcac
     3301 tctgaagtag ctccgcagca ggttcatcag gcctctggtg gcgtatctgg tatccaccag
     3361 gttccggttg atgaagtctt tctgcacgga gaacctgttg atgtcccgtt cttccagcag
     3421 atactctttc ttggtcttgc tgattctgcc cttgcccttg gccagattca ggatgtgctt
     3481 cttgaaggtt tcgtagctga tcttgctgtc gctgctgctc aggtactgga atggggtccg
     3541 gttgcccttc ttgctggctt cttcctgctt cacgagcacc ttgttgttga agctgttgtc
     3601 gaaggacacg cttctgggga tgatgtggtc cacctcatag ttgaaggggt tgttcagcag
     3661 atcttccaga gggatggctt ccaggctgta caggcacttg ccttcctgca tgtcgtgcag
     3721 cttgatcttc tcgatcaggt acttggcgtt ctctttgccg gtggtccgga tgatttcctc
     3781 gatccgctcg ttggtctgcc ggttccgctt ctgcatctcg ttgatcattt tctgggcgtc
     3841 cttggagttc ttctcgcggg ccagctcgat aatgatgtcg ttgggcaggc cgtacttctt
     3901 gatgatggcg ttgatcactt tgatgctctg gatgaagctt ctcttcacga cggggctcag
     3961 gatgaagtcg tccaccaggg tggtggggat ctctttctgc tgggacaggt ccaccttctt
     4021 gggcaccagc ttcagccggt tgaagatagc gatctggttg tcgttggtgt gccacagctc
     4081 gtccaggatc aggttgatgg ccttcaggct caggttgtgg gtgccggtat agcccttcag
     4141 attagagatc tgctcgatct cttcctgggt cagctcggag ttcagattgg tcagttcttc
     4201 ctggatgtcc tcgctgctct ggtagatggt caggatcttg gcaatctgat ccagcagctc
     4261 ggcgttctca ataatctctt tccgggcggt aatgtccttg atgtcgtggt acaccttcag
     4321 gttggtgaac tcgggcttgc cggtgctggt cactctgtag cccttaatat cctcttcgtt
     4381 cacgaggatt tctttggcga tctgcttcag ggtgggcttc ttcttctgct tgaacacgtt
     4441 ctcgatgatc tggaacttct cgtaatattc cagcttctcg ttctcgtccc tggtgatcac
     4501 gagattgttc aggtcgttca gggcgttgta caggtcggcg ttgtaggcgt acttcacgct
     4561 ccgcagttcc tcggggaagt aggtgcagtg gcccatcagc atctcgtacc attctttgat
     4621 gtccttccag ccgaaggggc tgccctcgcc aggtccctca tagtaggtcc gccgggtttc
     4681 cagcaggtcg atgtaggtgt cgatgaagct ctggtccagc tggtggtagg ccttctgcac
     4741 cttcagcagc tgtttggctt ctttcacgta gtcgctggtc ttgaatctgt tgatgctgcc
     4801 ccgcacttcg ccgtctttct tcagccgttc cagctgcagt tcggccacgt atttctcttc
     4861 cagggccttg ctgttccggc tgatctgctc tttggtggac agctcgttgc cggtgtcctc
     4921 ttccacctcg ttcacgttgt gcacgcctct tctcttggcc aggtgcagca gggcggcaga
     4981 gaactcttcc tcgctcagct tctggctcag gcccttcact ctggcctcgt aggggttgat
     5041 gccgctcagc tcgctgtggt cggtcagcag gttgtagtcg aacagcagct tcttcactct
     5101 ctggattcta tgccgcctcc gccgcttcag ccttctggcg cctctcttgc tccgcctgcc
     5161 ctcgttgttt tccacgttgg cctctttgaa cagccgcacg ccggcatcga tcacgtcccg
     5221 tgtctcgtag tcgatgatgc cgtagcccac gctggtgatg ccgatgtcca ggcccaggat
     5281 gtagttccgc ttggctgctg ggactccgtg gataccgact ttccgcttct tctttggtga
     5341 ctcgaactcg cttccgtcgg ctgtccgttt gccgccgtcc agcttcacgc gcttggcggc
     5401 gggccccatg gtggcgcgcc gcggatctct agcggatctg acggttcact aaaccagctc
     5461 tgcttatata gacctcccac cgtacacgcc taccgcccat ttgcgtcaat ggggcggagt
     5521 tgttacgaca ttttggaaag tcccgttgat tttggtgcca aaacaaactc ccattgacgt
     5581 caatggggtg gagacttgga aatccccgtg agtcaaaccg ctatccacgc ccattgatgt
     5641 actgccaaaa ccgcatcacc atggtaatag cgatgactaa tacgtagatg tactgccaag
     5701 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
     5761 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
     5821 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
     5881 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
     5941 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
     6001 taattgatta ctattaataa ctagtcaata atcaatgtca acgcgtatat ctggcccgta
     6061 catcgcgaag cagcgcaaaa cgcctaaccc taagcagatt cttcatgcaa ttgtcggtca
     6121 agccttgcct tgttgtagct taaattttgc tcgcgcacta ctcagcgacc tccaacacac
     6181 aagcagggag cagatactgg cttaactatg cggcatcaga gcagattgta ctgagagtag
     6241 accctagggg atcgggagat cgatctcccg atccgtcgac gtcaggtggc acttttcggg
     6301 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     6361 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     6421 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     6481 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     6541 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     6601 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     6661 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     6721 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     6781 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     6841 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     6901 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     6961 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     7021 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     7081 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     7141 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     7201 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     7261 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     7321 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     7381 tcccttaacg tgagttttcg ttccactgag tgtcagaccc cgtagaaaag atcaaaggat
     7441 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     7501 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     7561 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     7621 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     7681 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     7741 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     7801 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     7861 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     7921 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     7981 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     8041 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     8101 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
     8161 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
     8221 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
     8281 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
     8341 attaggcacc ctaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga
     8401 gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag ctctagctag
     8461 aggtcgacgg tatcgagccc cagctggttc tttccgcctc agaagccata gagcccaccg
     8521 catccccagc atgcctgcta ttgtcttccc aatcctcccc cttgctgtcc tgccccaccc
     8581 caccccccag aatagaatga cacctactca gacaatgcga tgcaattttc tcattttatt
     8641 aggaaaggac agtgggagtg gcaccttcca gggtcaagga aggcacgggg gaggggcaaa
     8701 caacagatgg ctggcaacta gaaggcacag tcgaggctga tcagcgggtt taaactcaat
     8761 ggtgatggtg atgatgaccg gttagacttt cctcttcttc ttgggctcga attccacctt
     8821 gcgcttggtc ttgggggggg tgctgtgctg gctgtcggcg gtgcgcttct cgctgccgtc
     8881 ggcggttctt tttgagcc
//
