LOCUS       sequence-207588-        8121 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses pCMV-dLbCpf1-BE0-Delta-iNLS crRNA in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8121)
  AUTHORS   Li X, Wang Y, Liu Y, Yang B, Wang X, Wei J, Lu Z, Zhang Y, Wu J, 
            Huang X, Yang L, Chen J
  TITLE     Base editing with a Cpf1-cytidine deaminase fusion.
  JOURNAL   Nat Biotechnol. 2018 Mar 19. pii: nbt.4102. doi: 10.1038/nbt.4102.
  PUBMED    29553573
REFERENCE   2  (bases 1 to 8121)
  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 
            Biotechnol. 2018 Mar 19. pii: nbt.4102. doi: 10.1038/nbt.4102."
COMMENT     Sequence Label: pCMV-dLbCpf1-BE0-Delta-iNLS
FEATURES             Location/Qualifiers
     source          1..8121
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        23..402
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        403..606
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     557..577
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     648..667
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        648..666
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             680..1366
                     /codon_start=1
                     /product="cytidine deaminase (C to U editing enzyme) from 
                     rat"
                     /label=APOBEC-1
                     /note="can use ssDNA as a substrate (Komor et al., 2016)"
                     /translation="MSSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWG
                     GRHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFL
                     SRYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSN
                     EAHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHI
                     LWATGLK"
     CDS             5117..5365
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             5378..5398
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5407..5424
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5447..5464)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5453..5677
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5748..5764)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5748..5764)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5761..5783)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5772..5788
                     /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(5796..5802,5803..5820,5821..5826))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5841..5862
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5979..5996)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6150..6738)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6230..6249)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6909..7700,7701..7769))
                     /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     7532..7551
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7770..7874)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7953..7972)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
       61 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
      121 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
      181 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
      241 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
      301 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
      361 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
      421 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
      481 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
      541 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
      601 agagctggtt tagtgaaccg tcagatccgc tagagatccg cggccgctaa tacgactcac
      661 tatagggaga gccgccacca tgagctcaga gactggccca gtggctgtgg accccacatt
      721 gagacggcgg atcgagcccc atgagtttga ggtattcttc gatccgagag agctccgcaa
      781 ggagacctgc ctgctttacg aaattaattg ggggggccgg cactccattt ggcgacatac
      841 atcacagaac actaacaagc acgtcgaagt caacttcatc gagaagttca cgacagaaag
      901 atatttctgt ccgaacacaa ggtgcagcat tacctggttt ctcagctgga gcccatgcgg
      961 cgaatgtagt agggccatca ctgaattcct gtcaaggtat ccccacgtca ctctgtttat
     1021 ttacatcgca aggctgtacc accacgctga cccccgcaat cgacaaggcc tgcgggattt
     1081 gatctcttca ggtgtgacta tccaaattat gactgagcag gagtcaggat actgctggag
     1141 aaactttgtg aattatagcc cgagtaatga agcccactgg cctaggtatc cccatctgtg
     1201 ggtacgactg tacgttcttg aactgtactg catcatactg ggcctgcctc cttgtctcaa
     1261 cattctgaga aggaagcagc cacagctgac attctttacc atcgctcttc agtcttgtca
     1321 ttaccagcga ctgcccccac acattctctg ggccaccggg ttgaaaagcg gcagcgagac
     1381 tcccgggacc tcagagtccg ccacacccga aagtatgtca aagctggaga aattcaccaa
     1441 ctgttatagc ctgtctaaga ccctgcgctt caaggcaatc ccagtgggca agacacaaga
     1501 gaacattgac aacaaacggc tcctggtgga ggatgagaag agggctgaag attacaaggg
     1561 cgttaagaag ctgctggata ggtactatct gtcattcatc aacgatgtcc tccacagtat
     1621 caagctgaag aatctgaaca attacatttc tctgttccgg aagaagacac ggaccgagaa
     1681 ggagaacaaa gagctggaga atctggagat caacctgagg aaagaaatag ctaaggcttt
     1741 caaagggaac gagggttaca agtccctgtt caagaaagac attatcgaga ctattctgcc
     1801 tgagttcctg gacgataaag atgagatcgc cctcgtcaat tccttcaatg ggtttaccac
     1861 agcctttacc ggcttcttcg acaatagaga gaatatgttc tctgaagagg ccaaatccac
     1921 tagcatcgcc tttcgctgca taaacgagaa cctgactagg tacatcagca atatggacat
     1981 ctttgagaaa gtcgatgcca tattcgacaa acatgaggtg caggagatta aggagaagat
     2041 cctgaactca gattacgatg tcgaagattt cttcgaggga gagttcttca acttcgtgct
     2101 cacacaagag ggcattgatg tgtacaatgc aatcattgga gggttcgtga cagagagtgg
     2161 cgagaagata aagggcctga acgagtatat caacctctac aaccagaaaa ccaagcagaa
     2221 actgcctaag ttcaagccac tgtacaaaca agtgctctca gatagggaaa gcctgagctt
     2281 ctacggtgaa gggtatacat cagatgaaga agtgctcgaa gtgttccgca acaccctcaa
     2341 taagaacagt gaaatcttct cttcaatcaa gaagctggag aaactgttca agaatttcga
     2401 tgagtactcc tctgccggaa tctttgtgaa gaatggccct gcaatatcca ctattagcaa
     2461 agacatcttt ggcgagtgga acgttatcag ggataagtgg aatgccgagt acgatgatat
     2521 tcatctcaag aagaaagccg tggttacaga gaaatacgag gatgatagac gcaagagctt
     2581 taagaagatt ggtagcttct ctctcgaaca gctgcaggag tacgccgacg ctgacctgtc
     2641 agtcgtggag aaactcaagg agatcataat ccagaaggtg gatgaaatct acaaagtgta
     2701 tggaagctct gagaaactct tcgatgcaga ctttgttctg gagaagagtc tgaagaagaa
     2761 cgacgcagtg gttgctatca tgaaggacct gctggattct gttaagtctt tcgagaatta
     2821 cattaaggca ttctttggtg aagggaagga gacaaatagg gacgagagct tctatggcga
     2881 ctttgttctg gcctacgaca tcctcctcaa ggttgaccac atctatgacg ctatacggaa
     2941 ttacgttacc cagaagccct atagcaaaga caagttcaag ctgtatttcc agaatccaca
     3001 gtttatgggt gggtgggata aagacaaaga aacagattac agggccacta tcctgcggta
     3061 cggcagcaaa tactatctgg ctatcatgga taagaagtac gccaaatgcc tccagaagat
     3121 cgacaaggac gacgtgaacg gtaactacga gaagatcaat tacaagctcc tgccaggacc
     3181 taacaagatg ctgcccaagg tgttcttctc caagaaatgg atggcctact ataacccaag
     3241 cgaggacatt cagaagatat acaagaatgg gacattcaag aagggcgata tgttcaacct
     3301 caacgactgc cacaagctga ttgatttctt caaggatagc atttctcgct atcccaagtg
     3361 gtctaatgca tacgatttca acttcagcga gactgagaag tacaaagaca tcgctggctt
     3421 ctaccgggag gtggaagagc aaggctataa ggtgtcattc gaatccgctt ctaagaagga
     3481 agtggataag ctcgtggaag agggtaagct gtacatgttc cagatataca acaaagactt
     3541 cagcgataag agccacggca ctccaaacct ccatactatg tatttcaagc tgctgtttga
     3601 cgagaacaac cacggacaga ttaggctgtc aggaggcgca gaactcttca tgcgcagagc
     3661 ttcactgaag aaggaggaac tcgttgtcca cccagccaat agccctatag ccaataagaa
     3721 tccagacaat cctaagaaaa ccactactct gtcttacgat gtgtataagg ataagagatt
     3781 ctctgaagat cagtacgaac tgcacatacc cattgccatt aacaagtgcc ctaagaacat
     3841 cttcaagatt aacacagagg ttagagtgct cctgaaacac gacgataacc cttatgttat
     3901 aggcattgct cgcggagaga gaaacctgct gtacatcgtc gtggtggacg gcaaaggcaa
     3961 catcgtggaa cagtacagtc tcaatgaaat cattaacaat ttcaacggaa tccgcattaa
     4021 gaccgactac cattctctcc tcgacaagaa ggagaaagaa aggttcgaag caagacagaa
     4081 ttggacaagt atagagaata tcaaagaact gaaggctggg tacatctctc aggttgtgca
     4141 caagatatgt gagctggtgg agaagtacga cgctgttatc gccctcgcgg acctgaatag
     4201 cggcttcaag aactccaggg tgaaggtgga gaagcaggtg tatcagaagt tcgagaagat
     4261 gctgatcgac aagctcaact atatggtgga caagaaatcc aatccttgcg ctactggtgg
     4321 agccctgaag ggctatcaaa tcaccaataa gttcgaatct ttcaagtcta tgagcaccca
     4381 gaatggcttc atcttctaca tacccgcatg gctgacatcc aagattgatc cctctaccgg
     4441 atttgttaat ctgctcaaga ctaagtacac ctctattgct gactcaaaga agttcatatc
     4501 atcatttgac cgcatcatgt acgtgccaga agaggacctg ttcgagtttg ccctggatta
     4561 caagaatttc tctcggactg acgccgacta catcaagaag tggaagctct actcttatgg
     4621 taatcggatt cgcatattcc gcaatcccaa gaagaataac gtgttcgatt gggaggaagt
     4681 ttgcctcacc agcgcttaca aggagctgtt caataagtat gggattaact accagcaggg
     4741 cgacataaga gccctgctgt gcgaacaatc tgataaggca ttctattcct ctttcatggc
     4801 actgatgtca ctgatgctgc aaatgcgcaa ttccatcacc ggaagaacag acgtggcctt
     4861 tctgatctct cctgtcaaga actcagatgg catcttctac gattcccgca actatgaagc
     4921 acaggagaat gctatcctgc ctaagaatgc cgatgcaaat ggagcctata acatcgccag
     4981 aaaggtcctc tgggccatag gacaattcaa gaaagctgaa gatgagaagc tggacaaggt
     5041 gaagatcgcc atttcaaaca aagagtggct cgaatatgct cagacctcag tgaagcatgg
     5101 atcatctggt ggttctacta atctgtcaga tattattgaa aaggagaccg gtaagcaact
     5161 ggttatccag gaatccatcc tcatgctccc agaggaggtg gaagaagtca ttgggaacaa
     5221 gccggaaagc gatatactcg tgcacaccgc ctacgacgag agcaccgacg agaatgtcat
     5281 gcttctgact agcgacgccc ctgaatacaa gccttgggct ctggtcatac aggatagcaa
     5341 cggtgagaac aagattaaga tgctctctgg tggttctccc aagaagaaga ggaaagtcta
     5401 accggtcatc atcaccatca ccattgagtt taaacccgct gatcagcctc gactgtgcct
     5461 tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt
     5521 gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg
     5581 tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac
     5641 aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc
     5701 tggggctcga taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     5761 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     5821 tgtaaagcct agggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     5881 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     5941 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     6001 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     6061 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     6121 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     6181 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     6241 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     6301 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     6361 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     6421 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     6481 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     6541 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6601 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6661 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6721 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6781 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6841 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     6901 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     6961 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     7021 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
     7081 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     7141 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     7201 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     7261 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     7321 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     7381 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     7441 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     7501 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     7561 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     7621 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     7681 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     7741 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     7801 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
     7861 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
     7921 cgatctcccg atcccctagg gtcgactctc agtacaatct gctctgatgc cgcatagtta
     7981 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
     8041 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg
     8101 cgttttgcgc tgcttcgcga t
//
