LOCUS       sequence-325776-        8183 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CURE-X: C to U RNA Editing, with low off-target effects but also 
            lower efficiency.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8183)
  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 8183)
  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: TC1614
FEATURES             Location/Qualifiers
     source          1..8183
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(31..135)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(210..229)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        401..780
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        781..984
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     935..955
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1029..1048
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1029..1047
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1125..1145
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2868..2888
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2901..3497
                     /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             3537..3557
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4743..4763
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4785..4841
                     /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             4842..5552
                     /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(4987..5005)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5255..5274
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    5595..6183
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5648..5668)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6066..6077)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(6186..6202)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6187..6203)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(6204..6221)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6210..6417
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(6423..6440)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6594..7182)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6674..6693)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7353..8144,8145..8183,1..30))
                     /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     7976..7995
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
       61 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
      121 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga
      181 tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
      241 cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa
      301 gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt
      361 tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt
      421 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      481 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      541 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      601 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      661 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
      721 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
      781 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
      841 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      901 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      961 tgggaggtct atataagcag agctctctgg ctaactagag aacccactgc ttactggctt
     1021 atcgaaatta atacgactca ctatagggag acccaagctg gctagcgttt aaacttaagc
     1081 ttgccaccat gaaacggact gctgacggca gcgaatttga gagccccaag aagaagagga
     1141 aagtcggatc catcgagaag aagaagagct tcgccaaggg catgggagtg aagagcaccc
     1201 tggtgtccgg ctctaaggtg tacatgacca catttgctga gggaagcgac gccaggctgg
     1261 agaagatcgt ggagggcgat agcatcagat ccgtgaacga gggagaggct ttcagcgccg
     1321 agatggctga caagaacgct ggctacaaga tcggaaacgc caagttttcc cacccaaagg
     1381 gctacgccgt ggtggctaac aacccactgt acaccggacc agtgcagcag gacatgctgg
     1441 gactgaagga gacactggag aagaggtact tcggcgagtc cgccgacgga aacgataaca
     1501 tctgcatcca ggtcatccac aacatcctgg atatcgagaa gatcctggct gagtacatca
     1561 caaacgccgc ttacgccgtg aacaacatct ccggcctgga caaggatatc atcggcttcg
     1621 gaaagttttc taccgtgtac acatacgacg agttcaagga tccagagcac caccgggccg
     1681 cttttaacaa caacgacaag ctgatcaacg ccatcaaggc tcagtacgac gagttcgata
     1741 actttctgga taaccccagg ctgggctact tcggacaggc tttcttttct aaggagggca
     1801 gaaactacat catcaactac ggaaacgagt gttacgacat cctggccctg ctgagcggac
     1861 tggcccactg ggtggtggcc aacaacgagg aggagtctcg gatcagccgc acctggctgt
     1921 acaacctgga caagaacctg gataacgagt acatctccac actgaactac ctgtacgaca
     1981 ggatcaccaa cgagctgaca aacagcttct ccaagaactc tgccgctaac gtgaactaca
     2041 tcgctgagac cctgggcatc aacccagctg agttcgctga gcagtacttc agattttcca
     2101 tcatgaagga gcagaagaac ctgggcttca acatcacaaa gctgagagaa gtgatgctgg
     2161 acagaaagga tatgtccgag atcaggaaga accacaaggt gttcgattct atcagaacca
     2221 aggtgtacac aatgatggac tttgtgatct acaggtacta catcgaggag gatgccaagg
     2281 tggccgctgc caacaagagc ctgcccgaca acgagaagtc tctgagcgag aaggatatct
     2341 tcgtgatcaa cctgagaggc tcctttaacg acgatcagaa ggacgctctg tactacgatg
     2401 aggccaacag gatctggaga aagctggaga acatcatgca caacatcaag gagttccggg
     2461 gaaacaagac ccgcgagtac aagaagaagg acgctccaag gctgcctagg atcctgcctg
     2521 ctggaaggga cgtgagcgcc ttcagcaagc tgatgtacgc cctgacaatg tttctggacg
     2581 gaaaggagat caacgatctg ctgaccacac tgatcaacaa gttcgacaac atccagtctt
     2641 ttctgaaagt gatgcctctg atcggcgtga acgctaagtt cgtggaggag tacgccttct
     2701 ttaaggacag cgccaagatc gctgatgagc tgcggctgat caagtccttt gccaggatgg
     2761 gagagccaat cgctgacgct aggagagcta tgtacatcga tgccatccgg atcctgggaa
     2821 ccaacggtac cagcaagagg acagcagacg ggtccgaatt cgaatccccc aagaaaaaga
     2881 ggaaggtggg tcccgggtct atggaagcca gcccagcatc cgggcccaga cacttgatgg
     2941 atccacacat attcacttcc aactttaaca atggcattgg aaggcataag acctacctgt
     3001 gctacgaagt ggagcgcctg gacaatggca cctcggtcaa gatggaccag cacaggggct
     3061 ttctacacaa ccaggctaag aatcttctct gtggctttta cggccgccat gcggagctgc
     3121 gcttcttgga cctggttcct tctttgcagt tggacccggc ccagatctac agggtcactt
     3181 ggttcatctc ctggagcccc tgcttctcct ggggctgtgc cggggaagtg cgtgcgttcc
     3241 ttcaggagaa cacacacgtg agactgcgta tcttcgctgc ccgcatctat gatgacgacc
     3301 ccctatataa ggaggcactg caaatgctgc gggatgctgg ggcccaagtc tccatcatga
     3361 cctacgatga atttaagcac tgctgggaca cctttgtgga ccaccaggga tgtcccttcc
     3421 agccctggga tggactagat gagcacagcc aagccctgag tgggaggctg cgggccattc
     3481 tccagaatca gggaaacgga tccaaaagaa ctgcggatgg atctgagttt gagagtccaa
     3541 aaaagaagag gaaggtcggt tctatgcgca acttcatcat caacaacgtg atcagcaaca
     3601 agcggtttca ctacctgatc agatacggcg acccagctca cctgcacgag atcgctaaga
     3661 acgaggccgt ggtgaagttc gtgctgggac ggatcgccga tatccagaag aagcagggcc
     3721 agaacggaaa gaaccagatc gaccgctact acgagacctg catcggcaag gataagggaa
     3781 agtccgtgtc tgagaaggtg gacgctctga ccaagatcat cacaggcatg aactacgacc
     3841 agttcgataa gaagagatct gtgatcgagg acaccggaag ggagaacgcc gagagagaga
     3901 agtttaagaa gatcatcagc ctgtacctga cagtgatcta ccacatcctg aagaacatcg
     3961 tgaacatcaa cgctagatac gtgatcggct tccactgcgt ggagcgcgat gcccagctgt
     4021 acaaggagaa gggatacgac atcaacctga agaagctgga ggagaagggc tttagctccg
     4081 tgaccaagct gtgcgctgga atcgacgaga cagcccccga caagaggaag gatgtggaga
     4141 aggagatggc cgagagagct aaggagagca tcgactccct ggagtctgct aaccctaagc
     4201 tgtacgccaa ctacatcaag tactccgatg agaagaaggc cgaggagttc accaggcaga
     4261 tcaacagaga gaaggccaag accgctctga acgcctacct gaggaacaca aagtggaacg
     4321 tgatcatccg ggaggacctg ctgcgcatcg ataacaagac ctgtacactg ttcgctaaca
     4381 aggctgtggc cctggaggtg gctcgctacg tgcacgccta catcaacgac atcgccgagg
     4441 tgaactccta ctttcagctg taccactaca tcatgcagag gatcatcatg aacgagagat
     4501 acgagaagtc tagcggcaag gtgtctgagt acttcgacgc cgtgaacgat gagaagaagt
     4561 acaacgatag actgctgaag ctgctgtgcg tgcctttcgg atactgtatc ccacggttta
     4621 agaacctgag catcgaggcc ctgttcgacc gcaacgaggc tgccaagttt gataaggaga
     4681 agaagaaggt gagcggcaac tccggttcta agcgcaccgc cgatggatcc gagttcgaat
     4741 cccctaaaaa gaaaagaaag gtgggatccg cggccgctgg atccgctact aacttcagcc
     4801 tgctgaagca ggctggagac gtggaggaga accctggacc tatggtgagc aagggcgagg
     4861 aggataacat ggccatcatc aaggagttca tgcgcttcaa ggtgcacatg gagggctccg
     4921 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggcaccc
     4981 agaccgccaa gctgaaggtg accaagggtg gccccctgcc cttcgcctgg gacatcctgt
     5041 cccctcagtt catgtacggc tccaaggcct acgtgaagca ccccgccgac atccccgact
     5101 acttgaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
     5161 gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggcgagttc atctacaagg
     5221 tgaagctgcg cggcaccaac ttcccctccg acggccccgt aatgcagaag aagaccatgg
     5281 gctgggaggc ctcctccgag cggatgtacc ccgaggacgg cgccctgaag ggcgagatca
     5341 agcagaggct gaagctgaag gacggcggcc actacgacgc tgaggtcaag accacctaca
     5401 aggccaagaa gcccgtgcag ctgcccggcg cctacaacgt caacatcaag ttggacatca
     5461 cctcccacaa cgaggactac accatcgtgg aacagtacga acgcgccgag ggccgccact
     5521 ccaccggcgg catggacgag ctgtacaagt aaggccgcga ctctagagtc gacctgcagg
     5581 catgcaagct tgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc
     5641 ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg
     5701 ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc
     5761 tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg
     5821 acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg
     5881 ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga
     5941 caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct
     6001 ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc ttctgctacg
     6061 tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc
     6121 ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc
     6181 cgcatcgata ccgtcgacct cgacctcgac tgtgccttct agttgccagc catctgttgt
     6241 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     6301 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     6361 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggagct
     6421 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     6481 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     6541 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     6601 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     6661 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     6721 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     6781 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     6841 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     6901 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     6961 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     7021 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     7081 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     7141 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     7201 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     7261 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     7321 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     7381 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     7441 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     7501 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     7561 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     7621 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     7681 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     7741 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     7801 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     7861 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     7921 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     7981 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     8041 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     8101 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     8161 caaaatgccg caaaaaaggg aat
//
