LOCUS       sequence-329013-        5468 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of mCerulean3 linked to Venus by 31 amino acid 
            linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5468)
  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 5468)
  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: mCerulean3-31aa-Venus-pEGFP-C3
FEATURES             Location/Qualifiers
     source          1..5468
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        142..445
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        446..649
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     600..620
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      688..697
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(694..696,697..699,700..1410)
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      1498..1507
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1504..1506,1507..1509,1510..2220)
                     /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"
     polyA_signal    2337..2458
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2374..2393)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2428..2447
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2465..2920)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2602..2623)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2814..2833
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2947..3051
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3053..3410
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3053..3244
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3076..3096)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3261..3396
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3445..4239
                     /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             3445..4239
                     /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(3499..3518)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4109..4128
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4427..4446)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4471..4518
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4847..5435
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5336..5355
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gccttttgct ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat
       61 aaccgtatta ccgccatgca ttagttatta atagtaatca attacggggt cattagttca
      121 tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc
      181 gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat
      241 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
      301 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
      361 cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
      421 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
      481 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      541 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      601 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagagctg gtttagtgaa
      661 ccgtcagatc cgctagcgct accggtcgcc accatggtga gcaagggcga ggagctgttc
      721 accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc
      781 gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gttcatctgc
      841 accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgag ctggggcgtg
      901 cagtgcttcg cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg
      961 cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc
     1021 cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc
     1081 gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacgc catccacggc
     1141 aacgtctata tcaccgccga caagcagaag aacggcatca aggccaactt cggcctcaac
     1201 tgcaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa cacccccatc
     1261 ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc caagctgagc
     1321 aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg
     1381 atcactctcg gcatggacga gctgtacaag tccggactca gatctcgagg tagtgggcta
     1441 agatctcggg cacaagcttc aaattctgcc gtcgatgctc gaggaggagg accggtcgcc
     1501 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
     1561 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
     1621 tacggcaagc tgaccctgaa gctgatctgc accaccggca agctgcccgt gccctggccc
     1681 accctcgtga ccaccctggg ctacggcctg cagtgcttcg cccgctaccc cgaccacatg
     1741 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
     1801 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
     1861 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1921 cacaagctgg agtacaacta caacagccac aacgtctata tcaccgccga caagcagaag
     1981 aacggcatca aggccaactt caagatccgc cacaacatcg aggacggcgg cgtgcagctc
     2041 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     2101 cactacctga gctaccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
     2161 gtcctgctgg aacgcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     2221 tccggactca gatcataatc agccatacca catttgtaga ggttttactt gctttaaaaa
     2281 acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt gttgttaact
     2341 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2401 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttaac
     2461 gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     2521 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     2581 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     2641 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     2701 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     2761 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     2821 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     2881 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac ttttcgggga
     2941 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     3001 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tcctgaggcg
     3061 gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3121 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3181 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     3241 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3301 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     3361 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa
     3421 gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg
     3481 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     3541 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     3601 ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg
     3661 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     3721 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     3781 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     3841 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     3901 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     3961 aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4021 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4081 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     4141 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     4201 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     4261 tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc gccgccttct
     4321 atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg
     4381 gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa acacggaagg
     4441 agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg
     4501 gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata
     4561 ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc
     4621 ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca
     4681 tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
     4741 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
     4801 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     4861 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     4921 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     4981 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     5041 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     5101 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     5161 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     5221 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     5281 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     5341 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     5401 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     5461 ggttcctg
//
