LOCUS       sequence-329015-        5414 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For expression of mCerulean3 linked to Venus by 13 amino acid 
            linker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5414)
  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 5414)
  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: mCerulean3-13aa-Venus-pEGFP-C3
FEATURES             Location/Qualifiers
     source          1..5414
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        52..355
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        356..559
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     510..530
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      598..607
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(604..606,607..609,610..1320)
                     /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      1354..1363
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1360..1362,1363..1365,1366..2076)
                     /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    2193..2314
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2230..2249)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2284..2303
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2321..2776)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2458..2479)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2670..2689
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2803..2907
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2909..3266
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2909..3100
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2932..2952)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3117..3252
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3301..4095
                     /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             3301..4095
                     /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(3355..3374)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3965..3984
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4283..4302)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4327..4374
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4703..5291
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5192..5211
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 atagtaatca attacggggt cattagttca tagcccatat atggagttcc gcgttacata
       61 acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat
      121 aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga
      181 gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc
      241 ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt acatgacctt
      301 atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta ccatggtgat
      361 gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg gatttccaag
      421 tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac gggactttcc
      481 aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg tacggtggga
      541 ggtctatata agcagagctg gtttagtgaa ccgtcagatc cgctagcgct accggtcgcc
      601 accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
      661 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
      721 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
      781 accctcgtga ccaccctgag ctggggcgtg cagtgcttcg cccgctaccc cgaccacatg
      841 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
      901 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
      961 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
     1021 cacaagctgg agtacaacgc catccacggc aacgtctata tcaccgccga caagcagaag
     1081 aacggcatca aggccaactt cggcctcaac tgcaacatcg aggacggcag cgtgcagctc
     1141 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
     1201 cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg cgatcacatg
     1261 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
     1321 tccggactca gatctcgagg aggaggaccg gtcgccacca tggtgagcaa gggcgaggag
     1381 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     1441 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagctg
     1501 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgggctac
     1561 ggcctgcagt gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1621 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1681 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1741 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1801 agccacaacg tctatatcac cgccgacaag cagaagaacg gcatcaaggc caacttcaag
     1861 atccgccaca acatcgagga cggcggcgtg cagctcgccg accactacca gcagaacacc
     1921 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccgcc
     1981 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggaacg cgtgaccgcc
     2041 gccgggatca ctctcggcat ggacgagctg tacaagtccg gactcagatc ataatcagcc
     2101 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc
     2161 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
     2221 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta
     2281 gttgtggttt gtccaaactc atcaatgtat cttaacgcgt aaattgtaag cgttaatatt
     2341 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     2401 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     2461 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     2521 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     2581 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     2641 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     2701 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     2761 ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     2821 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     2881 cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg tggaatgtgt
     2941 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3001 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3061 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3121 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3181 tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag tgaggaggct
     3241 tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg atcgtttcgc
     3301 atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc
     3361 ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca
     3421 gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg
     3481 caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg
     3541 ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag
     3601 gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg
     3661 cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc
     3721 atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa
     3781 gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag catgcccgac
     3841 ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat
     3901 ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac
     3961 atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc
     4021 ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt
     4081 gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc
     4141 tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg
     4201 ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg
     4261 cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa ggaacccgcg
     4321 ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt gttcataaac
     4381 gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc
     4441 aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag
     4501 ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac tcatatatac
     4561 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     4621 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     4681 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     4741 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     4801 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     4861 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     4921 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     4981 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     5041 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     5101 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     5161 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     5221 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     5281 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     5341 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcca
     5401 tgcattagtt atta
//
