LOCUS       sequence-329004-        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        51..354
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        355..558
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     509..529
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      597..606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(603..605,606..608,609..1319)
                     /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(648..669)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(909..928)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    1740..1792
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    1916..2037
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1953..1972)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(2044..2499)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2181..2202)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2393..2412
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2526..2630
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2632..2989
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2632..2823
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2655..2675)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2840..2975
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3024..3818
                     /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             3024..3818
                     /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(3078..3097)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3688..3707
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4006..4025)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4050..4097
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4426..5014
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4915..4934
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
       61 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
      121 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
      181 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
      241 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
      301 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtgatg
      361 cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg atttccaagt
      421 ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg ggactttcca
      481 aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt acggtgggag
      541 gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta ccggtcgcca
      601 ccatggtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
      661 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
      721 acggcaagct gaccctgaag ctgatctgca ccaccggcaa gctgcccgtg ccctggccca
      781 ccctcgtgac caccctgggc tacggcctgc agtgcttcgc ccgctacccc gaccacatga
      841 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
      901 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
      961 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     1021 acaagctgga gtacaactac aacagccaca acgtctatat caccgccgac aagcagaaga
     1081 acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcggc gtgcagctcg
     1141 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     1201 actacctgag ctaccagtcc gccctgagca aagaccccaa cgagaagcgc gatcacatgg
     1261 tcctgctgga acgcgtgacc gccgccggga tcactctcgg catggacgag ctgtacaagg
     1321 gaggctccgg cagccaggcc agccgctcct tcaaccaagg aagaatagag ccagattctg
     1381 aaagtcagga agaaatcatc cacaacgaag ccagacatga agcccaagaa ggcgatgagg
     1441 aagaccacaa catccagccc acactggtga gacagctagc cgcacagttc atgaatggca
     1501 gcctgtcgga ggaagacaaa aggaactgcc tggccaaagc ccttgatgag gtgaagacag
     1561 ccttccccag agacatggag aacgacaagg ccatgctgat aatgaccatg ctgttggcca
     1621 aaaaagtggc cagtcacgca ccatctttgc tccgtgatgt cttccacacg actgtcaact
     1681 ttattaacca gaacctattc tcctatgtga ggaacttggt tagaaacgag atggactaac
     1741 tcgagctcaa gcttcgaatt ctgcagtcga cggtaccgcg ggcccgggat ccaccggatc
     1801 tagataactg atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa
     1861 cctcccacac ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt
     1921 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     1981 agcatttttt tcactgcatt ctagttgtgg tttgtcaaaa ctcatcaatg tatcttaagg
     2041 cgtaaattgt aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct
     2101 cattttttaa ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg
     2161 agatagggtt gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact
     2221 ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac
     2281 cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga
     2341 gcccccgatt tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga
     2401 aagcgaaagg agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca
     2461 ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa
     2521 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     2581 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg
     2641 aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     2701 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     2761 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     2821 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     2881 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct
     2941 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag
     3001 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     3061 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     3121 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     3181 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     3241 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     3301 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     3361 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     3421 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     3481 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     3541 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     3601 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     3661 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     3721 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     3781 gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat
     3841 gaccgaccaa gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta
     3901 tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg
     3961 ggatctcatg ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga
     4021 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     4081 tgttgggtcg tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac
     4141 cccaccgaga ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc
     4201 cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat
     4261 agcctcaggt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     4321 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     4381 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     4441 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     4501 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     4561 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtatc
     4621 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     4681 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     4741 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     4801 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     4861 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     4921 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     4981 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     5041 gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc
     5101 tgtggataac cgtattaccg ccatgcatta gttattaa
//
