LOCUS       sequence-273617-        5054 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of engineered BFP variant that undergoes conversion to 
            GFP after targeted modification with a cytidine deaminase-based DNA 
            base editor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5054)
  AUTHORS   Standage-Beier K, Tekel SJ, Brookhouser N, Schwarz G, Nguyen T, Wang
            X, Brafman DA
  TITLE     A transient reporter for editing enrichment (TREE) in human cells.
  JOURNAL   Nucleic Acids Res. 2019 Aug 20. pii: 5552066. doi: 
            10.1093/nar/gkz713.
  PUBMED    31428784
REFERENCE   2  (bases 1 to 5054)
  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: "Nucleic 
            Acids Res. 2019 Aug 20. pii: 5552066. doi: 10.1093/nar/gkz713."
COMMENT     Sequence Label: pEF-BFP
FEATURES             Location/Qualifiers
     source          1..5054
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        28..1210
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          259..1201
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1158..1178
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     regulatory      1256..1265
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1262..1264,1265..1267,1268..1981)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTHGVQCFGRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1307..1328)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1568..1587)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1915..1936
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2068..2087)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    2133..2188
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(2187..2206)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(2549..2565)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2549..2565)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2562..2584)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2573..2589
                     /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(2597..2603,2604..2621,2622..2627))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2642..2663
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2722..2917
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2768..2903
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    2923..3057
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2960..2979)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3014..3033
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3125..3142)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3296..3884)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3376..3395)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4055..4846,4847..4915))
                     /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     4678..4697
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4916..5020)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttgattattg actagatcat cgcgtgaggc tccggtgccc gtcagtgggc agagcgcaca
       61 tcgcccacag tccccgagaa gttgggggga ggggtcggca attgaaccgg tgcctagaga
      121 aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag
      181 ggtgggggag aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg
      241 tttgccgcca gaacacaggt aagtgccgtg tgtggttccc gcgggcctgg cctctttacg
      301 ggttatggcc cttgcgtgcc ttgaattact tccacgcccc tggctgcagt acgtgattct
      361 tgatcccgag cttcgggttg gaagtgggtg ggagagttcg aggccttgcg cttaaggagc
      421 cccttcgcct cgtgcttgag ttgaggcctg gcctgggcgc tggggccgcc gcgtgcgaat
      481 ctggtggcac cttcgcgcct gtctcgctgc tttcgataag tctctagcca tttaaaattt
      541 ttgatgacct gctgcgacgc tttttttctg gcaagatagt cttgtaaatg cgggccaaga
      601 tctgcacact ggtatttcgg tttttggggc cgcgggcggc gacggggccc gtgcgtccca
      661 gcgcacatgt tcggcgaggc ggggcctgcg agcgcggcca ccgagaatcg gacgggggta
      721 gtctcaagct ggccggcctg ctctggtgcc tggcctcgcg ccgccgtgta tcgccccgcc
      781 ctgggcggca aggctggccc ggtcggcacc agttgcgtga gcggaaagat ggccgcttcc
      841 cggccctgct gcagggagct caaaatggag gacgcggcgc tcgggagagc gggcgggtga
      901 gtcacccaca caaaggaaaa gggcctttcc gtcctcagcc gtcgcttcat gtgactccac
      961 ggagtaccgg gcgccgtcca ggcacctcga ttagttctcg agcttttgga gtacgtcgtc
     1021 tttaggttgg ggggaggggt tttatgcgat ggagtttccc cacactgagt gggtggagac
     1081 tgaagttagg ccagcttggc acttgatgta attctccttg gaatttgccc tttttgagtt
     1141 tggatcttgg ttcattctca agcctcagac agtggttcaa agtttttttc ttccatttca
     1201 ggtgtcgtga ggaattctgc agtcgacggt accgcgggcc cgggatccac cggtcgccac
     1261 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     1321 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     1381 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     1441 cctcgtgacc accctgaccc acggcgtgca gtgcttcggc cgctaccccg accacatgaa
     1501 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     1561 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     1621 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     1681 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
     1741 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     1801 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     1861 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     1921 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta
     1981 aagcggccgc actcctcagg tgcaggctgc ctatcagaag gtggtggctg gtgtggccaa
     2041 tgccctggct cacaaatacc actgagatct ttttccctct gccaaaaatt atggggacat
     2101 catgaagccc cttgagcatc tgacttctgg ctaataaagg aaatttattt tcattgcaat
     2161 agtgtgttgg aattttttgt gtctctcact cggaaggaca tatgggaggg caaatcattt
     2221 aaaacatcag aatgagtatt tggtttagag tttggcaaca tatgcccata tgctggctgc
     2281 catgaacaaa ggttggctat aaagaggtca tcagtatatg aaacagcccc ctgctgtcca
     2341 ttccttattc catagaaaag ccttgacttg aggttagatt ttttttatat tttgttttgt
     2401 gttatttttt tctttaacat ccctaaaatt ttccttacat gttttactag ccagattttt
     2461 cctcctctcc tgactactcc cagtcatagc tgtccctctt ctcttatgga gatccctcga
     2521 cctgcagccc aagcttggcg taatcatggt catagctgtt tcctgtgtga aattgttatc
     2581 cgctcacaat tccacacaac atacgagccg gaagcataaa gtgtaaagcc tggggtgcct
     2641 aatgagtgag ctaactcaca ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa
     2701 acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca accatagtcc cgcccctaac
     2761 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     2821 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     2881 gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct
     2941 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     3001 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctggatccgc
     3061 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     3121 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     3181 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     3241 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     3301 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     3361 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     3421 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     3481 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     3541 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     3601 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     3661 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     3721 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
     3781 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     3841 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     3901 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     3961 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     4021 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     4081 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     4141 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     4201 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     4261 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     4321 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     4381 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     4441 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     4501 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     4561 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     4621 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     4681 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     4741 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     4801 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     4861 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     4921 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     4981 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     5041 cacctgggtc gaca
//
