LOCUS       sequence-390315-        8217 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A-to-G base editor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8217)
  TITLE     Kohn lab plasmids
REFERENCE   2  (bases 1 to 8217)
  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"
COMMENT     Sequence Label: PCMV-ABE8e-VRER
FEATURES             Location/Qualifiers
     source          1..8217
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(16..35)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        207..586
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        587..790
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     741..761
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     832..851
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        832..850
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             900..920
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1515..3230
                     /codon_start=1
                     /product="N-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(N)
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIE"
     CDS             5658..5678
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5687..5704
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5727..5744)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5733..5957
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6028..6044)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6028..6044)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6041..6063)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6052..6068
                     /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(6076..6082,6083..6100,6101..6106))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6121..6142
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6259..6276)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6430..7018)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6510..6529)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7189..7980,7981..8049))
                     /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     7812..7831
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8050..8154)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 agggtcgact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc
       61 tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca
      121 aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg cgctgcttcg
      181 cgatgtacgg gccagatata cgcgttgaca ttgattattg actagttatt aatagtaatc
      241 aattacgggg tcattagttc atagcccata tattgagttc cgcgttacat aacttacggt
      301 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
      361 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      421 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      481 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
      541 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
      601 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
      661 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
      721 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
      781 aagcagagct ggtttagtga accgtcagat ccgctagaga tccgcggccg ctaatacgac
      841 tcactatagg gagagccgcc accatgaaac ggacagccga cggaagcgag ttcgagtcac
      901 caaagaagaa gcggaaagtc tctgaggtgg agttttccca cgagtactgg atgagacatg
      961 ccctgaccct ggccaagagg gcacgggatg agagggaggt gcctgtggga gccgtgctgg
     1021 tgctgaacaa tagagtgatc ggcgagggct ggaacagagc catcggcctg cacgacccaa
     1081 cagcccatgc cgaaattatg gccctgagac agggcggcct ggtcatgcag aactacagac
     1141 tgattgacgc caccctgtac gtgacattcg agccttgcgt gatgtgcgcc ggcgccatga
     1201 tccactctag gatcggccgc gtggtgtttg gcgtgaggaa ctcaaaaaga ggcgccgcag
     1261 gctccctgat gaacgtgctg aactaccccg gcatgaatca ccgcgtcgaa attaccgagg
     1321 gaatcctggc agatgaatgt gccgccctgc tgtgcgattt ctatcggatg cctagacagg
     1381 tgttcaatgc tcagaagaag gcccagagct ccatcaactc cggaggatct agcggaggct
     1441 cctctggctc tgagacacct ggcacaagcg agagcgcaac acctgaaagc agcgggggca
     1501 gcagcggggg gtcagacaag aagtacagca tcggcctggc catcggcacc aactctgtgg
     1561 gctgggccgt gatcaccgac gagtacaagg tgcccagcaa gaaattcaag gtgctgggca
     1621 acaccgaccg gcacagcatc aagaagaacc tgatcggagc cctgctgttc gacagcggcg
     1681 aaacagccga ggccacccgg ctgaagagaa ccgccagaag aagatacacc agacggaaga
     1741 accggatctg ctatctgcaa gagatcttca gcaacgagat ggccaaggtg gacgacagct
     1801 tcttccacag actggaagag tccttcctgg tggaagagga taagaagcac gagcggcacc
     1861 ccatcttcgg caacatcgtg gacgaggtgg cctaccacga gaagtacccc accatctacc
     1921 acctgagaaa gaaactggtg gacagcaccg acaaggccga cctgcggctg atctatctgg
     1981 ccctggccca catgatcaag ttccggggcc acttcctgat cgagggcgac ctgaaccccg
     2041 acaacagcga cgtggacaag ctgttcatcc agctggtgca gacctacaac cagctgttcg
     2101 aggaaaaccc catcaacgcc agcggcgtgg acgccaaggc catcctgtct gccagactga
     2161 gcaagagcag acggctggaa aatctgatcg cccagctgcc cggcgagaag aagaatggcc
     2221 tgttcggaaa cctgattgcc ctgagcctgg gcctgacccc caacttcaag agcaacttcg
     2281 acctggccga ggatgccaaa ctgcagctga gcaaggacac ctacgacgac gacctggaca
     2341 acctgctggc ccagatcggc gaccagtacg ccgacctgtt tctggccgcc aagaacctgt
     2401 ccgacgccat cctgctgagc gacatcctga gagtgaacac cgagatcacc aaggcccccc
     2461 tgagcgcctc tatgatcaag agatacgacg agcaccacca ggacctgacc ctgctgaaag
     2521 ctctcgtgcg gcagcagctg cctgagaagt acaaagagat tttcttcgac cagagcaaga
     2581 acggctacgc cggctacatt gacggcggag ccagccagga agagttctac aagttcatca
     2641 agcccatcct ggaaaagatg gacggcaccg aggaactgct cgtgaagctg aacagagagg
     2701 acctgctgcg gaagcagcgg accttcgaca acggcagcat cccccaccag atccacctgg
     2761 gagagctgca cgccattctg cggcggcagg aagattttta cccattcctg aaggacaacc
     2821 gggaaaagat cgagaagatc ctgaccttcc gcatccccta ctacgtgggc cctctggcca
     2881 ggggaaacag cagattcgcc tggatgacca gaaagagcga ggaaaccatc accccctgga
     2941 acttcgagga agtggtggac aagggcgctt ccgcccagag cttcatcgag cggatgacca
     3001 acttcgataa gaacctgccc aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt
     3061 acttcaccgt gtataacgag ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc
     3121 ccgccttcct gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc
     3181 ggaaagtgac cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag tgcttcgact
     3241 ccgtggaaat ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca taccacgatc
     3301 tgctgaaaat tatcaaggac aaggacttcc tggacaatga ggaaaacgag gacattctgg
     3361 aagatatcgt gctgaccctg acactgtttg aggacagaga gatgatcgag gaacggctga
     3421 aaacctatgc ccacctgttc gacgacaaag tgatgaagca gctgaagcgg cggagataca
     3481 ccggctgggg caggctgagc cggaagctga tcaacggcat ccgggacaag cagtccggca
     3541 agacaatcct ggatttcctg aagtccgacg gcttcgccaa cagaaacttc atgcagctga
     3601 tccacgacga cagcctgacc tttaaagagg acatccagaa agcccaggtg tccggccagg
     3661 gcgatagcct gcacgagcac attgccaatc tggccggcag ccccgccatt aagaagggca
     3721 tcctgcagac agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg
     3781 agaacatcgt gatcgaaatg gccagagaga accagaccac ccagaaggga cagaagaaca
     3841 gccgcgagag aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc cagatcctga
     3901 aagaacaccc cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc
     3961 agaatgggcg ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg
     4021 atgtggacca tatcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc
     4081 tgaccagaag cgacaagaac cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga
     4141 agaagatgaa gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt
     4201 tcgacaatct gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca
     4261 tcaagagaca gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact
     4321 cccggatgaa cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca
     4381 ccctgaagtc caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg
     4441 agatcaacaa ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc
     4501 tgatcaaaaa gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg
     4561 acgtgcggaa gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact
     4621 tcttctacag caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga
     4681 tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg
     4741 gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa
     4801 agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg
     4861 ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgtgagcc
     4921 ctacagttgc ctattctgtc ctagtagtgg caaaagttga gaagggaaaa tccaagaaac
     4981 tgaagtcagt caaagaatta ttggggataa cgattatgga gcgctcgtct tttgaaaaga
     5041 accccatcga cttccttgag gcgaaaggtt acaaggaagt aaaaaaggat ctcataatta
     5101 aactaccaaa gtatagtctg tttgagttag aaaatggccg aaaacggatg ttggctagcg
     5161 ccagagagct tcaaaagggg aacgaactcg cactaccgtc taaatacgtg aatttcctgt
     5221 atttagcgtc ccattacgag aagttgaaag gttcacctga agataacgaa cagaagcaac
     5281 tttttgttga gcagcacaaa cattatctcg acgaaatcat agagcaaatt tcggaattca
     5341 gtaagagagt catcctagct gatgccaatc tggacaaagt attaagcgca tacaacaagc
     5401 acagggataa acccatacgt gagcaggcgg aaaatattat ccatttgttt actcttacca
     5461 acctcggcgc tccagccgca ttcaagtatt ttgacacaac gatagatcgc aaagagtaca
     5521 gatctaccaa ggaggtgcta gacgcgacac tgattcacca atccatcacg ggattatatg
     5581 aaactcggat agatttgtca cagcttgggg gtgactctgg cggctcaaaa agaaccgccg
     5641 acggcagcga attcgagccc aagaagaaga ggaaagtcta accggtcatc atcaccatca
     5701 ccattgagtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     5761 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     5821 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     5881 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     5941 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcga taccgtcgac
     6001 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     6061 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct agggtgccta
     6121 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     6181 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     6241 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     6301 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     6361 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     6421 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     6481 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     6541 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     6601 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     6661 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     6721 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     6781 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     6841 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     6901 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     6961 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     7021 agatcctttg atcttttcta cggggtctga cactcagtgg aacgaaaact cacgttaagg
     7081 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     7141 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     7201 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     7261 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     7321 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     7381 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     7441 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     7501 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     7561 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     7621 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     7681 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     7741 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     7801 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     7861 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     7921 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     7981 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     8041 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     8101 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     8161 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat cgatctcccg atcccct
//
