LOCUS       sequence-325775-        8156 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CURE-C2: C to U RNA Editing, with high efficiency and high 
            off-target effects.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8156)
  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 8156)
  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: TC1681
FEATURES             Location/Qualifiers
     source          1..8156
                     /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             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             4592..4648
                     /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             4649..5359
                     /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(4794..4812)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5062..5081
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    5402..5990
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5455..5475)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5873..5884)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5993..6009)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5994..6010)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(6011..6028)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6017..6224
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6230..6247)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6401..6989)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6481..6500)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7160..7951,7952..8020))
                     /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     7783..7802
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8021..8125)
                     /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 taaagctgct gcaaaggtga 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 ccgctggatc cgctactaac ttcagcctgc tgaagcaggc
     4621 tggagacgtg gaggagaacc ctggacctat ggtgagcaag ggcgaggagg ataacatggc
     4681 catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga
     4741 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
     4801 gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat
     4861 gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc
     4921 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac
     4981 cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg
     5041 caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     5101 ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa
     5161 gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc
     5221 cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct cccacaacga
     5281 ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat
     5341 ggacgagctg tacaagtaag gccgcgactc tagagtcgac ctgcaggcat gcaagcttga
     5401 taatcaacct ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc
     5461 tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg
     5521 tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt
     5581 gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac
     5641 tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc
     5701 tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     5761 gttgggcact gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct
     5821 cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct
     5881 caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct
     5941 tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg
     6001 tcgacctcga cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc
     6061 gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa
     6121 attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac
     6181 agcaaggggg aggattggga agacaatagc aggcatgctg gggagcttcc tcgctcactg
     6241 actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa
     6301 tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc
     6361 aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc
     6421 ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat
     6481 aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc
     6541 cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct
     6601 cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg
     6661 aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc
     6721 cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga
     6781 ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa
     6841 gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta
     6901 gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc
     6961 agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg
     7021 acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga
     7081 tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg
     7141 agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct
     7201 gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg
     7261 agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc
     7321 cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa
     7381 ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc
     7441 cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt
     7501 cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc
     7561 ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt
     7621 tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc
     7681 catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt
     7741 gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata
     7801 gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga
     7861 tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag
     7921 catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa
     7981 aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt
     8041 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     8101 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtc
//
