LOCUS       sequence-323738-        5138 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express Venus fused to the N-terminus of the Bcl-2 family protein, 
            tBid with mutation in BH3 domain to disrupt binding anti-apoptotic 
            proteins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5138)
  AUTHORS   Osterlund EJ, Hirmiz N, Pemberton JM, Nougarede A, Liu Q, Leber B, 
            Fang Q, Andrews DW
  TITLE     Efficacy and specificity of inhibitors of BCL-2 family protein 
            interactions assessed by affinity measurements in live cells.
  JOURNAL   Sci Adv. 2022 Apr 22;8(16):eabm7375. doi: 10.1126/sciadv.abm7375. 
            Epub 2022 Apr 20.
  PUBMED    35442739
REFERENCE   2  (bases 1 to 5138)
  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: "Sci Adv."; 
            date: "2022-04-22"; volume: "8(16):eabm7375. doi"; pages: " 
            10.1126/sciadv.abm7375. Epub 2022 Apr 20"
COMMENT     Sequence Label: Venus-tBid-4E-pEGFP-C1
FEATURES             Location/Qualifiers
     source          1..5138
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     ERVTAAGITLGMDELYK"
     primer_bind     complement(658..679)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(919..938)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    1750..1802
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    1926..2047
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1963..1982)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2017..2036
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2054..2509)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2191..2212)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2403..2422
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2536..2640
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2642..2999
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2642..2833
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2665..2685)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2850..2985
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3034..3828
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3034..3828
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3088..3107)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3698..3717
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4016..4035)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4060..4107
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4436..5024
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4925..4944
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg
      661 gtcgagctgg acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc
      721 gatgccacct acggcaagct gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgggc tacggcctgc agtgcttcgc ccgctacccc
      841 gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag
      901 cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag
      961 ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac
     1021 atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat caccgccgac
     1081 aagcagaaga acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcggc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag ctaccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga acgcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagg gaggctccgg cagccaggcc agccgctcct tcaaccaagg aagaatagag
     1381 ccagattctg aaagtcagga agaaatcatc cacaacgaag ccagacatga agcccaagaa
     1441 ggcgatgagg aagaccacaa catccagccc acactggtga gacagctagc cgcacagttc
     1501 atgaatggca gcctgtcgga ggaagacaaa aggaactgcc tggccaaagc ccttgatgag
     1561 gtgaagacag ccttccccag agacatggag aacgacaagg ccatgctgat aatgaccatg
     1621 ctgttggcca aaaaagtggc cagtcacgca ccatctttgc tccgtgatgt cttccacacg
     1681 actgtcaact ttattaacca gaacctattc tcctatgtga ggaacttggt tagaaacgag
     1741 atggactaac tcgagctcaa gcttcgaatt ctgcagtcga cggtaccgcg ggcccgggat
     1801 ccaccggatc tagataactg atcataatca gccataccac atttgtagag gttttacttg
     1861 ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg
     1921 ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     1981 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     2041 tatcttaacg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt
     2101 taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa
     2161 gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag
     2221 aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt
     2281 gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac
     2341 cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag
     2401 gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg
     2461 cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact
     2521 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     2581 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     2641 cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     2701 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     2761 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     2821 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     2881 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
     2941 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag
     3001 atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca
     3061 ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc
     3121 ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc
     3181 aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg
     3241 ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg
     3301 gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct
     3361 gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct
     3421 acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa
     3481 gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa
     3541 ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc
     3601 gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt
     3661 ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct
     3721 gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc
     3781 gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg
     3841 ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg
     3901 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     3961 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa
     4021 cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata
     4081 aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact
     4141 ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc
     4201 caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag
     4261 gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt
     4321 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     4381 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     4441 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     4501 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     4561 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     4621 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     4681 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     4741 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     4801 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     4861 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     4921 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     4981 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     5041 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     5101 cccctgattc tgtggataac cgtattaccg ccatgcat
//
