LOCUS       sequence-485489-        7932 bp    DNA     circular SYN 08-SEP-2025
DEFINITION  Proximity-triggered Protein Trans-Splicing to generate the splice 
            product AML1-ETO, Extended Figure 3 in Nature Chemical Biology 20.10
            (2024): 1353-1360..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7932)
  AUTHORS   Lee G, Muir TW
  TITLE     Distinct phases of cellular signaling revealed by time-resolved 
            protein synthesis.
  JOURNAL   bioRxiv [Preprint]. 2023 Jul 10:2023.07.10.548208. doi: 
            10.1101/2023.07.10.548208.
  PUBMED    37503273
REFERENCE   2  (bases 1 to 7932)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "bioRxiv 
            [Preprint]. 2023 Jul 10:2023.07.10.548208. doi: 
            10.1101/2023.07.10.548208."
COMMENT     Sequence Label: pcDNA3.1-NES-CAGEc-ETOc (Npu)
FEATURES             Location/Qualifiers
     source          1..7932
                     /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             988..1011
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1021..1044
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1054..1077
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1114..1389
                     /codon_start=1
                     /product="T2098L mutant of FKBP-rapamycin binding domain of
                     human FRAP"
                     /label=FRB*
                     /note="binds synthetic ligands such as AP21967"
                     /translation="ILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTL
                     KETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLLQAWDLYYHVFRRIS"
     CDS             1573..1593
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1594..1614
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3472..3498
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(3523..3540)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3529..3753
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3799..4227
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3886..3905)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4096..4117
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4236..4256)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4241..4570
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4421..4556
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4483..4502
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4637..5431
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4691..4710)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5301..5320
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5605..5726
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5642..5661)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5696..5715
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5775..5791)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5775..5791)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5788..5810)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5799..5815
                     /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(5823..5829,5830..5847,5848..5853))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5868..5889
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6006..6023)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6177..6765)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6257..6276)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6936..7727,7728..7796))
                     /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     7559..7578
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7797..7901)
                     /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 gccatggctc tccaaaaaaa gttggaagag cttgaactgg acgaacaggg aggaagtgga
      961 ggtagtctcg agggtaccat gggttcagac tataaagatg atgatgataa aggtggctcc
     1021 gactacaagg atgacgatga caagggaggt agtgactaca aagacgatga cgacaagggc
     1081 ggaggaggaa gtggaggtag tggaagagta gcgatcctct ggcatgagat gtggcatgaa
     1141 ggcctggaag aggcatctcg tttgtacttt ggggaaagga acgtgaaagg catgtttgag
     1201 gtgctggagc ccttgcatgc tatgatggaa cggggccccc agactctgaa ggaaacatcc
     1261 tttaatcagg cctatggtcg agatttaatg gaggcccaag agtggtgcag gaagtacatg
     1321 aaatcaggga atgtcaagga cctcctccaa gcctgggacc tctattatca tgtgttccga
     1381 cgaatctcag gaaatggaaa tggaggagaa caagaggtgt ttgagtattg tttggaagat
     1441 ggttcattga ttcgggcaac aaaagaccat aagtttatga ctgttgatgg tcaaatgttg
     1501 ccaattgatg aaatatttga acgtgaattg gatttgatgc gggttgataa tttgccgaat
     1561 ggaagtggag gagagaacct gtattttcag ggcgagaacc tgtattttca gggcggaagt
     1621 ggaggaatca aaatagccac acgtaaatat ttaggcaaac aaaatgtcta tgacattgga
     1681 gttgagcgcg accataattt tgcactcaaa aatggcttca tagcttctaa ttgctttaat
     1741 cgtactgaga agcactccac aatgccagac tcacctgtgg atgtgaagac gcaatctagg
     1801 ctgactcctc caacaatgcc acctccccca actactcaag gagctccaag aaccagttca
     1861 tttacaccga caacgttaac taatggcacg agccattctc ctacagcctt gaatggcgcc
     1921 ccctcaccac ccaatggctt cagcaatggg ccttcctctt cttcctcctc ctctctggct
     1981 aatcaacagc tgcccccagc ctgtggtgcc aggcaactca gcaagctgaa aaggttcctt
     2041 actaccctgc agcagtttgg caatgacatt tcacccgaga taggagaaag agttcgcacc
     2101 ctcgttctgg gactagtgaa ctccactttg acaattgaag aatttcattc caaactgcaa
     2161 gaagctacta acttcccact gagacctttt gtcatcccat ttttgaaggc caacttgccc
     2221 ctgctgcagc gtgagctcct ccactgcgca agactggcca aacagaaccc tgcccagtac
     2281 ctcgcccagc atgaacagct gcttctggat gccagcacca cctcacctgt tgactcctca
     2341 gagctgcttc tcgatgtgaa cgaaaacggg aagaggcgaa ctccagacag aaccaaagaa
     2401 aatggctttg acagagagcc tttgcactca gaacatccaa gcaagcgacc atgcactatt
     2461 agcccaggcc agcggtacag tccaaataac ggcttatcct accagcccaa tggcctgcct
     2521 caccctaccc cacctccacc tcagcattac cgtttggatg atatggccat tgcccaccac
     2581 tacagggact cctatcgaca ccccagccac agggacctca gggacagaaa cagacctatg
     2641 gggttgcatg gcacacgtca agaagaaatg attgatcaca gactaacaga cagagaatgg
     2701 gcagaagagt ggaaacatct tgaccatctg ttaaactgca taatggacat ggtagaaaaa
     2761 acaaggcgat ctctcaccgt actaaggcgg tgtcaagaag cagaccggga agaattgaat
     2821 tactggatcc ggcggtacag tgacgccgag gacttaaaaa aaggtggcgg cagtagcagc
     2881 agccactcta ggcagcagag tcccgtcaac ccagacccag ttgcactaga cgcgcatcgg
     2941 gaattccttc acaggcctgc gtctggatac gtgccagagg agatctggaa gaaagctgag
     3001 gaggccgtca atgaggtgaa gcgccaggcg atgacggagc tgcagaaggc cgtgtctgag
     3061 gcggagcgga aagcccacga catgatcaca acagagaggg ccaagatgga gcgcacggtc
     3121 gccgaggcca aacggcaggc ggcggaggac gcactggcag ttatcaatca gcaggaggat
     3181 tcaagcgaga gttgctggaa ttgtggccgt aaagcgagtg aaacctgcag tggctgtaac
     3241 acagcccgat actgtggctc attttgccag cacaaagact gggagaagca ccatcacatc
     3301 tgtggacaga ccctgcaggc ccagcagcag ggagacacac ctgcagtcag ctcctctgtc
     3361 acgcccaaca gcggggctgg gagcccgatg gacacaccac cagcagccac tccgaggtca
     3421 accaccccgg gaaccccttc caccatagag acaacccctc gctctcgact ctacccgtac
     3481 gacgttccgg actacgcttc ttgagtttaa accgctgatc agcctcgact gtgccttcta
     3541 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     3601 ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc
     3661 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     3721 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     3781 gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     3841 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     3901 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     3961 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     4021 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     4081 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     4141 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc
     4201 tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg
     4261 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     4321 aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     4381 caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc
     4441 cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg cagaggccga
     4501 ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg
     4561 cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc aagagacagg
     4621 atgaggatcg tttcgcatga ttgaacaaga tggattgcac gcaggttctc cggccgcttg
     4681 ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct ctgatgccgc
     4741 cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg acctgtccgg
     4801 tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca cgacgggcgt
     4861 tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc tgctattggg
     4921 cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga aagtatccat
     4981 catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc cattcgacca
     5041 ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc ttgtcgatca
     5101 ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg ccaggctcaa
     5161 ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa
     5221 tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc tgggtgtggc
     5281 ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc ttggcggcga
     5341 atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc agcgcatcgc
     5401 cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga aatgaccgac
     5461 caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt ctatgaaagg
     5521 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
     5581 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
     5641 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
     5701 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
     5761 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     5821 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     5881 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     5941 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     6001 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
     6061 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
     6121 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
     6181 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
     6241 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
     6301 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
     6361 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
     6421 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
     6481 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
     6541 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
     6601 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
     6661 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
     6721 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
     6781 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
     6841 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
     6901 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
     6961 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
     7021 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
     7081 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
     7141 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
     7201 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
     7261 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
     7321 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
     7381 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
     7441 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
     7501 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
     7561 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
     7621 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     7681 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     7741 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     7801 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     7861 tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg
     7921 ccacctgacg tc
//
