LOCUS       sequence-329392-        8899 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 8899)
  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 8899)
  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..8899
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      626..635
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             638..664
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             704..724
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6134..6154
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6163..6180
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6203..6220)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6209..6433
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6504..6520)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6504..6520)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6517..6539)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6528..6544
                     /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(6552..6558,6559..6576,6577..6582))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6597..6618
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6735..6752)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6906..7494)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6986..7005)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7665..8456,8457..8525))
                     /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     8288..8307
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8526..8630)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(8709..8728)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatccgcta gagatccgcg gccgcgccac catggggccc gccgccaagc gcgtgaagct
      661 ggacggcggc aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa
      721 agtcggtatc cacggagtcc cagcagccaa gcggaactac atcctgggcc tggacatcgg
      781 catcaccagc gtgggctacg gcatcatcga ctacgagaca cgggacgtga tcgatgccgg
      841 cgtgcggctg ttcaaagagg ccaacgtgga aaacaacgag ggcaggcgga gcaagagagg
      901 cgccagaagg ctgaagcggc ggaggcggca tagaatccag agagtgaaga agctgctgtt
      961 cgactacaac ctgctgaccg accacagcga gctgagcggc atcaacccct acgaggccag
     1021 agtgaagggc ctgagccaga agctgagcga ggaagagttc tctgccgccc tgctgcacct
     1081 ggccaagaga agaggcgtgc acaacgtgaa cgaggtggaa gaggacaccg gcaacgagct
     1141 gtccaccaaa gagcagatca gccggaacag caaggccctg gaagagaaat acgtggccga
     1201 actgcagctg gaacggctga agaaagacgg cgaagtgcgg ggcagcatca acagattcaa
     1261 gaccagcgac tacgtgaaag aagccaaaca gctgctgaag gtgcagaagg cctaccacca
     1321 gctggaccag agcttcatcg acacctacat cgacctgctg gaaacccggc ggacctacta
     1381 tgagggacct ggcgagggca gccccttcgg ctggaaggac atcaaagaat ggtacgagat
     1441 gctgatgggc cactgcacct acttccccga ggaactgcgg agcgtgaagt acgcctacaa
     1501 cgccgacctg tacaacgccc tgaacgacct gaacaatctc gtgatcacca gggacgagaa
     1561 cgagaagctg gaatattacg agaagttcca gatcatcgag aacgtgttca agcagaagaa
     1621 gaagcccacc ctgaagcaga tcgccaaaga aatcctcgtg aacgaagagg atattaaggg
     1681 ctacagagtg accagcaccg gcaagcccga gttcaccaac ctgaaggtgt accacgacat
     1741 caaggacatt accgcccgga aagagattat tgagaacgcc gagctgctgg atcagattgc
     1801 caagatcctg accatctacc agagcagcga ggacatccag gaagaactga ccaatctgaa
     1861 ctccgagctg acccaggaag agatcgagca gatctctaat ctgaagggct ataccggcac
     1921 ccacaacctg agcctgaagg ccatcaacct gatcctggac gagctgtggc acaccaacga
     1981 caaccagatc gctatcttca accggctgaa gctggtgccc aagaaggtgg acctgtccca
     2041 gcagaaagag atccccacca ccctggtgga cgacttcatc ctgagccccg tcgtgaagag
     2101 aagcttcatc cagagcatca aagtgatcaa cgccatcatc aagaagtacg gcctgcccaa
     2161 cgacatcatt atcgagctgg cccgcgagaa gaactccaag gacgcccaga aaatgatcaa
     2221 cgagatgcag aagcggaacc ggcagaccaa cgagcggatc gaggaaatca tccggaccac
     2281 cggcaaagag aacgccaagt acctgatcga gaagatcaag ctgcacgaca tgcaggaagg
     2341 caagtgcctg tacagcctgg aagccatccc tctggaagat ctgctgaaca accccttcaa
     2401 ctatgaggtg gaccacatca tccccagaag cgtgtccttc gacaacagct tcaacaacaa
     2461 ggtgctcgtg aagcaggaag aagccagcaa gaagggcaac cggaccccat tccagtacct
     2521 gagcagcagc gacagcaaga tcagctacga aaccttcaag aagcacatcc tgaatctggc
     2581 caagggcaag ggcagaatca gcaagaccaa gaaagagtat ctgctggaag aacgggacat
     2641 caacaggttc tccgtgcaga aagacttcat caaccggaac ctggtggata ccagatacgc
     2701 caccagaggc ctgatgaacc tgctgcggag ctacttcaga gtgaacaacc tggacgtgaa
     2761 agtgaagtcc atcaatggcg gcttcaccag ctttctgcgg cggaagtgga agtttaagaa
     2821 agagcggaac aaggggtaca agcaccacgc cgaggacgcc ctgatcattg ccaacgccga
     2881 tttcatcttc aaagagtgga agaaactgga caaggccaaa aaagtgatgg aaaaccagat
     2941 gttcgaggaa aagcaggccg agagcatgcc cgagatcgaa accgagcagg agtacaaaga
     3001 gatcttcatc accccccacc agatcaagca cattaaggac ttcaaggact acaagtacag
     3061 ccaccgggtg gacaagaagc ctaatagaga gctgattaac gacaccctgt actccacccg
     3121 gaaggacgac aagggcaaca ccctgatcgt gaacaatctg aacggcctgt acgacaagga
     3181 caatgacaag ctgaaaaagc tgatcaacaa gagccccgaa aagctgctga tgtaccacca
     3241 cgacccccag acctaccaga aactgaagct gattatggaa cagtacggcg acgagaagaa
     3301 tcccctgtac aagtactacg aggaaaccgg gaactacctg accaagtact ccaaaaagga
     3361 caacggcccc gtgatcaaga agattaagta ttacggcaac aaactgaacg cccatctgga
     3421 catcaccgac gactacccca acagcagaaa caaggtcgtg aagctgtccc tgaagcccta
     3481 cagattcgac gtgtacctgg acaatggcgt gtacaagttc gtgaccgtga agaatctgga
     3541 tgtgatcaaa aaagaaaact actacgaagt gaatagcaag tgctatgagg aagctaagaa
     3601 gctgaagaag atcagcaacc aggccgagtt tatcgcctcc ttctacaaca acgatctgat
     3661 caagatcaac ggcgagctgt atagagtgat cggcgtgaac aacgacctgc tgaaccggat
     3721 cgaagtgaac atgatcgaca tcacctaccg cgagtacctg gaaaacatga acgacaagag
     3781 gccccccagg atcattaaga caatcgcctc caagacccag agcattaaga agtacagcac
     3841 agacattctg ggcaacctgt atgaagtgaa atctaagaag caccctcaga tcatcaaaaa
     3901 gggctctgga ggatctagcg gaggatcctc tggcagcgag acaccaggaa caagcgagtc
     3961 agcaacacca gagagcagtg gcggcagcag cggcggcagc agcaccctaa atatagaaga
     4021 tgagtatcgg ctacatgaga cctcaaaaga gccagatgtt tctctagggt ccacatggct
     4081 gtctgatttt cctcaggcct gggcggaaac cgggggcatg ggactggcag ttcgccaagc
     4141 tcctctgatc atacctctga aagcaacctc tacccccgtg tccataaaac aataccccat
     4201 gtcacaagaa gccagactgg ggatcaagcc ccacatacag agactgttgg accagggaat
     4261 actggtaccc tgccagtccc cctggaacac gcccctgcta cccgttaaga aaccagggac
     4321 taatgattat aggcctgtcc aggatctgag agaagtcaac aagcgggtgg aagacatcca
     4381 ccccaccgtg cccaaccctt acaacctctt gagcgggctc ccaccgtccc accagtggta
     4441 cactgtgctt gatttaaagg atgccttttt ctgcctgaga ctccacccca ccagtcagcc
     4501 tctcttcgcc tttgagtgga gagatccaga gatgggaatc tcaggacaat tgacctggac
     4561 cagactccca cagggtttca aaaacagtcc caccctgttt aatgaggcac tgcacagaga
     4621 cctagcagac ttccggatcc agcacccaga cttgatcctg ctacagtacg tggatgactt
     4681 actgctggcc gccacttctg agctagactg ccaacaaggt actcgggccc tgttacaaac
     4741 cctagggaac ctcgggtatc gggcctcggc caagaaagcc caaatttgcc agaaacaggt
     4801 caagtatctg gggtatcttc taaaagaggg tcagagatgg ctgactgagg ccagaaaaga
     4861 gactgtgatg gggcagccta ctccgaagac ccctcgacaa ctaagggagt tcctagggaa
     4921 ggcaggcttc tgtcgcctct tcatccctgg gtttgcagaa atggcagccc ccctgtaccc
     4981 tctcaccaaa ccggggactc tgtttaattg gggcccagac caacaaaagg cctatcaaga
     5041 aatcaagcaa gctcttctaa ctgccccagc cctggggttg ccagatttga ctaagccctt
     5101 tgaactcttt gtcgacgaga agcagggcta cgccaaaggt gtcctaacgc aaaaactggg
     5161 accttggcgt cggccggtgg cctacctgtc caaaaagcta gacccagtag cagctgggtg
     5221 gcccccttgc ctacggatgg tagcagccat tgccgtactg acaaaggatg caggcaagct
     5281 aaccatggga cagccactag tcattctggc cccccatgca gtagaggcac tagtcaaaca
     5341 accccccgac cgctggcttt ccaacgcccg gatgactcac tatcaggcct tgcttttgga
     5401 cacggaccgg gtccagttcg gaccggtggt agccctgaac ccggctacgc tgctcccact
     5461 gcctgaggaa gggctgcaac acaactgcct tgatatcctg gccgaagccc acggaacccg
     5521 acccgaccta acggaccagc cgctcccaga cgccgaccac acctggtaca cggatggaag
     5581 cagtctctta caagagggac agcgtaaggc gggagctgcg gtgaccaccg agaccgaggt
     5641 aatctgggct aaagccctgc cagccgggac atccgctcag cgggctgaac tgatagcact
     5701 cacccaggcc ctaaagatgg cagaaggtaa gaagctaaat gtttatactg atagccgtta
     5761 tgcttttgct actgcccata tccatggaga aatatacaga aggcgtgggt ggctcacatc
     5821 agaaggcaaa gagatcaaaa ataaagacga gatcttggcc ctactaaaag ccctctttct
     5881 gcccaaaaga cttagcataa tccattgtcc aggacatcaa aagggacaca gcgccgaggc
     5941 tagaggcaac cggatggctg accaagcggc ccgaaaggca gccatcacag agactccaga
     6001 cacctctacc ctcctcatag aaaattcatc accctctggc ggctcaaaaa gaaccgccga
     6061 cggcagcgag aagcgcaccg ccgacagcca gcacagcacc ccccccaaga ccaagcgcaa
     6121 ggtggaattc gagcccaaga agaagaggaa agtctaaccg gtcatcatca ccatcaccat
     6181 tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     6241 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     6301 taaaatgaga aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     6361 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     6421 gtgggctcta tggcttctga ggcggaaaga accagctggg gctcgatacc gtcgacctct
     6481 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     6541 acaattccac acaacatacg agccggaagc ataaagtgta aagcctaggg tgcctaatga
     6601 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     6661 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     6721 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     6781 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     6841 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     6901 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     6961 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     7021 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     7081 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     7141 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     7201 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     7261 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     7321 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     7381 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     7441 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     7501 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     7561 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     7621 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     7681 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     7741 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     7801 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     7861 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     7921 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     7981 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     8041 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     8101 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     8161 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     8221 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     8281 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     8341 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     8401 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     8461 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     8521 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     8581 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     8641 cgaaaagtgc cacctgacgt cgacggatcg ggagatcgat ctcccgatcc cctagggtcg
     8701 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt
     8761 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg
     8821 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta
     8881 cgggccagat atacgcgtt
//
