LOCUS       sequence-390149-        4932 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian cell expression of EGFP-tagged Ubiquitin.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4932)
  AUTHORS   Harding O, Holzer E, Riley JF, Martens S, Holzbaur ELF
  TITLE     Damaged mitochondria recruit the effector NEMO to activate NF-kappaB
            signaling.
  JOURNAL   Mol Cell. 2023 Sep 7;83(17):3188-3204.e7. doi: 
            10.1016/j.molcel.2023.08.005.
  PUBMED    37683611
REFERENCE   2  (bases 1 to 4932)
  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"; doi: 
            "10.1016/j.molcel.2023.08"; journalName: "Mol Cell"; date: 
            "2023-09-7- 7"; volume: "83"; issue: "17"; pages: "3188-3204.e7"
COMMENT     Sequence Label: EGFP-Ubiquitin
FEATURES             Location/Qualifiers
     source          1..4932
                     /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="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(613..615,616..618,619..1329)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     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"
     primer_bind     1266..1287
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1366..1593
                     /codon_start=1
                     /product="human ubiquitin"
                     /label=ubiquitin
                     /translation="MQIFVKTLTGKTITLEVEPSDTIENVKAKIQDKEGIPPDQQRLIF
                     AGKQLEDGRTLSDYNIQKESTLHLVLRLRGG"
     polyA_signal    1741..1862
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1778..1797)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1832..1851
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1869..2324)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2006..2027)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2218..2237
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2351..2455
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        2457..2648
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2480..2500)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2645..2779
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2828..3622
                     /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             2828..3622
                     /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(2882..2901)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3492..3511
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3810..3829)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3854..3901
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4230..4818
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4719..4738
                     /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 ttcatctgca ccaccggcaa gctgcccgtg
      781 ccctggccca ccctcgtgac caccctgacc tacggcgtgc agtgcttcag 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 catggccgac
     1081 aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga ggacggcagc
     1141 gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg
     1201 cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa cgagaagcgc
     1261 gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg catggacgag
     1321 ctgtacaagt ccggactcag atctcgagct caagcttcgg gccgcatgca gatcttcgtg
     1381 aagactctga ctggtaagac catcaccctc gaggttgagc ccagtgacac cattgagaat
     1441 gtcaaggcaa agatccaaga taaggaaggc atccctcctg accagcagag gctgatcttt
     1501 gctggaaaac agctggaaga tgggcgcacc ctgtctgact acaacatcca gaaagagtcc
     1561 accctgcacc tggtactccg tctcagaggt gggtgaggta ccgcgggccc gggatccacc
     1621 ggatctagat aactgatcat aatcagccat accacatttg tagaggtttt acttgcttta
     1681 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     1741 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     1801 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     1861 taacgcgtaa attgtaagcg ttaatatttt gttaaaattc gcgttaaatt tttgttaaat
     1921 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     1981 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     2041 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     2101 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     2161 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     2221 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     2281 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtcaggtg gcacttttcg
     2341 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     2401 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtcctga
     2461 ggcggaaaga accagctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     2521 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     2581 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     2641 atagtcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     2701 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     2761 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     2821 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     2881 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     2941 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3001 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3061 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3121 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3181 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3241 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3301 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     3361 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     3421 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     3481 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     3541 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     3601 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     3661 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     3721 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     3781 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     3841 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     3901 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     3961 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     4021 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     4081 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4141 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     4201 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     4261 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     4321 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     4381 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     4441 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     4501 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     4561 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     4621 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     4681 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     4741 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     4801 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     4861 tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt
     4921 accgccatgc at
//
