LOCUS       sequence-325934-        8213 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CURE-N: C to U RNA Editing, with high efficiency and low off-target 
            effects.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8213)
  AUTHORS   Huang X, Lv J, Li Y, Mao S, Li Z, Jing Z, Sun Y, Zhang X, Shen S, 
            Wang X, Di M, Ge J, Huang X, Zuo E, Chi T
  TITLE     Programmable C-to-U RNA editing using the human APOBEC3A deaminase.
  JOURNAL   EMBO J. 2020 Nov 16;39(22):e104741. doi: 10.15252/embj.2020104741. 
            Epub 2020 Oct 15.
  PUBMED    33058229
REFERENCE   2  (bases 1 to 8213)
  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: "EMBO J."; 
            date: "2020-11-16"; volume: "39(22):e104741. doi"; pages: " 
            10.15252/embj.2020104741. Epub 2020 Oct 15"
COMMENT     Sequence Label: TC1332
FEATURES             Location/Qualifiers
     source          1..8213
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             959..979
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3938..3964
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             3980..4576
                     /codon_start=1
                     /gene="human APOBEC3A"
                     /gene_synonym="A3A"
                     /product="human cytidine deaminase (C to U editing enzyme) 
                     that efficiently edits methylated cytosines (Wang et al., 
                     2018)"
                     /label=APOBEC3A
                     /note="also known as hA3A"
                     /translation="MEASPASGPRHLMDPHIFTSNFNNGIGRHKTYLCYEVERLDNGTS
                     VKMDQHRGFLHNQAKNLLCGFYGRHAELRFLDLVPSLQLDPAQIYRVTWFISWSPCFSW
                     GCAGEVRAFLQENTHVRLRIFAARIYDDDPLYKEALQMLRDAGAQVSIMTYDEFKHCWD
                     TFVDHQGCPFQPWDGLDEHSQALSGRLRAILQNQGN"
     CDS             4622..4642
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4649..4705
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             4706..5416
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(4851..4869)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5119..5138
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    5459..6047
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5512..5532)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5930..5941)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6050..6066)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6051..6067)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(6068..6085)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6074..6281
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6287..6304)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6458..7046)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6538..6557)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7217..8008,8009..8077))
                     /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     7840..7859
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8078..8182)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttgcca ccatgaagcg caccgccgat ggatccgagt tcgaatcccc
      961 taaaaagaaa agaaaggtga acatccccgc tctggtggaa aaccagaaga agtactttgg
     1021 cacctacagc gtgatggcca tgctgaacgc tcagaccgtg ctggaccaca tccagaaggt
     1081 ggccgatatt gagggcgagc agaacgagaa caacgagaat ctgtggtttc accccgtgat
     1141 gagccacctg tacaacgcca agaacggcta cgacaagcag cccgagaaaa ccatgttcat
     1201 catcgagcgg ctgcagagct acttcccatt cctgaagatc atggccgaga accagagaga
     1261 gtacagcaac ggcaagtaca agcagaaccg cgtggaagtg aacagcaacg acatcttcga
     1321 ggtgctgaag cgcgccttcg gcgtgctgaa gatgtacagg gacctgacca acgcatacaa
     1381 gacctacgag gaaaagctga acgacggctg cgagttcctg accagcacag agcaacctct
     1441 gagcggcatg atcaacaact actacacagt ggccctgcgg aacatgaacg agagatacgg
     1501 ctacaagaca gaggacctgg ccttcatcca ggacaagcgg ttcaagttcg tgaaggacgc
     1561 ctacggcaag aaaaagtccc aagtgaatac cggattcttc ctgagcctgc aggactacaa
     1621 cggcgacaca cagaagaagc tgcacctgag cggagtggga atcgccctgc tgatctgcct
     1681 gttcctggac aagcagtaca tcaacatctt tctgagcagg ctgcccatct tctccagcta
     1741 caatgcccag agcgaggaac ggcggatcat catcagatcc ttcggcatca acagcatcaa
     1801 gctgcccaag gaccggatcc acagcgagaa gtccaacaag agcgtggcca tggatatgct
     1861 caacgaagtg aagcggtgcc ccgacgagct gttcacaaca ctgtctgccg agaagcagtc
     1921 ccggttcaga atcatcagcg acgaccacaa tgaagtgctg atgaagcgga gcagcgacag
     1981 attcgtgcct ctgctgctgc agtatatcga ttacggcaag ctgttcgacc acatcaggtt
     2041 ccacgtgaac atgggcaagc tgagatacct gctgaaggcc gacaagacct gcatcgacgg
     2101 ccagaccaga gtcagagtga tcgagcagcc cctgaacggc ttcggcagac tggaagaggc
     2161 cgagacaatg cggaagcaag agaacggcac cttcggcaac agcggcatcc ggatcagaga
     2221 cttcgagaac atgaagcggg acgacgccaa tcctgccaac tatccctaca tcgtggacac
     2281 ctacacacac tacatcctgg aaaacaacaa ggtcgagatg tttatcaacg acaaagagga
     2341 cagcgcccca ctgctgcccg tgatcgagga tgatagatac gtggtcaaga caatccccag
     2401 ctgccggatg agcaccctgg aaattccagc catggccttc cacatgtttc tgttcggcag
     2461 caagaaaacc gagaagctga tcgtggacgt gcacaaccgg tacaagagac tgttccaggc
     2521 catgcagaaa gaagaagtga ccgccgagaa tatcgccagc ttcggaatcg ccgagagcga
     2581 cctgcctcag aagatcctgg atctgatcag cggcaatgcc cacggcaagg atgtggacgc
     2641 cttcatcaga ctgaccgtgg acgacatgct gaccgacacc gagcggagaa tcaagagatt
     2701 caaggacgac cggaagtcca ttcggagcgc cgacaacaag atgggaaaga gaggcttcaa
     2761 gcagatctcc acaggcaagc tggccgactt cctggccaag gacatcgtgc tgtttcagcc
     2821 cagcgtgaac gatggcgaga acaagatcac cggcctgaac taccggatca tgcagagcgc
     2881 cattgccgtg tacgatagcg gcgacgatta cgaggccaag cagcagttca agctgatgtt
     2941 cgagaaggcc cggctgatcg gcaagggcac aacagagcct catccatttc tgtacaaggt
     3001 gttcgcccgc agcatccccg ccaatgccgt cgagttctac gagcgctacc tgatcgagcg
     3061 gaagttctac ctgaccggcc tgtccaacga gatcaagaaa ggcaacagag tggatgtgcc
     3121 cttcatccgg cgggaccaga acaagtggaa aacacccgcc atgaagaccc tgggcagaat
     3181 ctacagcgag gatctgcccg tggaactgcc cagacagatg ttcgacaatg agatcaagtc
     3241 ccacctgaag tccctgccac agatggaagg catcgacttc aacaatgcca acgtgaccta
     3301 tctgatcgcc gagtacatga agagagtgct ggacgacgac ttccagacct tctaccagtg
     3361 gaaccgcaac taccggtaca tggacatgct taagggcgag tacgacagaa agggctccct
     3421 gcagcactgc ttcaccagcg tggaagagag agaaggcctc tggaaagagc gggcctccag
     3481 aacagagcgg tacagaaagc aggccagcaa caagatccgc agcaaccggc agatgagaaa
     3541 cgccagcagc gaagagatcg agacaatcct ggataagcgg ctgagcaaca gccggaacga
     3601 gtaccagaaa agcgagaaag tgatccggcg ctacagagtg caggatgccc tgctgtttct
     3661 gctggccaaa aagaccctga ccgaactggc cgatttcgac ggcgagaggt tcaaactgaa
     3721 agaaatcatg cccgacgccg agaagggaat cctgagcgag atcatgccca tgagcttcac
     3781 cttcgagaaa ggcggcaaga agtacaccat caccagcgag ggcatgaagc tgaagaacta
     3841 cggcgacttc tttgtgctgg ctagcgacaa gaggatcggc aacctgctgg aactcgtggg
     3901 cagcgacatc gtgtccaaag aggatggatc ccttcaactg cctccacttg aaagactgac
     3961 actgggatcc ggctcgagca tggaagccag cccagcatcc gggcccagac acttgatgga
     4021 tccacacata ttcacttcca actttaacaa tggcattgga aggcataaga cctacctgtg
     4081 ctacgaagtg gagcgcctgg acaatggcac ctcggtcaag atggaccagc acaggggctt
     4141 tctacacaac caggctaaga atcttctctg tggcttttac ggccgccatg cggagctgcg
     4201 cttcttggac ctggttcctt ctttgcagtt ggacccggcc cagatctaca gggtcacttg
     4261 gttcatctcc tggagcccct gcttctcctg gggctgtgcc ggggaagtgc gtgcgttcct
     4321 tcaggagaac acacacgtga gactgcgtat cttcgctgcc cgcatctatg atgacgaccc
     4381 cctatataag gaggcactgc aaatgctgcg ggatgctggg gcccaagtct ccatcatgac
     4441 ctacgatgaa tttaagcact gctgggacac ctttgtggac caccagggat gtcccttcca
     4501 gccctgggat ggactagatg agcacagcca agccctgagt gggaggctgc gggccattct
     4561 ccagaatcag ggaaacgcgg ccggatccaa aagaactgcg gatggatctg agtttgagag
     4621 tccaaaaaag aagaggaagg tcggatccgc tactaacttc agcctgctga agcaggctgg
     4681 agacgtggag gagaaccctg gacctatggt gagcaagggc gaggaggata acatggccat
     4741 catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt
     4801 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa
     4861 ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta
     4921 cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt
     4981 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt
     5041 gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac
     5101 caacttcccc tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc
     5161 cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct
     5221 gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt
     5281 gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga
     5341 ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga
     5401 cgagctgtac aagtaaggcc gcgactctag agtcgacctg caggcatgca agcttgataa
     5461 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     5521 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     5581 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     5641 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     5701 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     5761 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     5821 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     5881 ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     5941 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     6001 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgtcg
     6061 acctcgacct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     6121 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     6181 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     6241 aagggggagg attgggaaga caatagcagg catgctgggg agcttcctcg ctcactgact
     6301 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     6361 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     6421 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     6481 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     6541 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     6601 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     6661 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     6721 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     6781 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     6841 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     6901 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     6961 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     7021 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     7081 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     7141 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     7201 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     7261 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     7321 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     7381 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     7441 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     7501 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     7561 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     7621 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     7681 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     7741 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     7801 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     7861 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     7921 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     7981 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     8041 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     8101 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     8161 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
