LOCUS       sequence-458398-        6343 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses control plasmid for ER-mitochondrial linkers, tagged with 
            EBFP2, in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6343)
  AUTHORS   Dematteis G, Tapella L, Casali C, Talmon M, Tonelli E, Reano S, 
            Ariotti A, Pessolano E, Malecka J, Chrostek G, Kulkoviene G, 
            Umbrasas D, Distasi C, Grilli M, Ladds G, Filigheddu N, Fresu LG, 
            Mikoshiba K, Matute C, Ramos-Gonzalez P, Jekabsone A, Cali T, Brini 
            M, Biggiogera M, Cavaliere F, Miggiano R, Genazzani AA, Lim D
  TITLE     ER-mitochondria distance is a critical parameter for efficient 
            mitochondrial Ca(2+) uptake and oxidative metabolism.
  JOURNAL   Commun Biol. 2024 Oct 10;7(1):1294. doi: 10.1038/s42003-024-06933-9.
  PUBMED    39390051
REFERENCE   2  (bases 1 to 6343)
  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: "Commun 
            Biol."; date: "2024-10-10"; volume: "7(1):1294. doi"; pages: " 
            10.1038/s42003-024-06933-9"
COMMENT     Sequence Label: pcDNA3.1-ER-EBFP2
FEATURES             Location/Qualifiers
     source          1..6343
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1934..1951)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1940..2164
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2210..2638
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2297..2316)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2507..2528
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2647..2667)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2652..2981
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2832..2967
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2894..2913
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3048..3842
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3102..3121)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3712..3731
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4016..4137
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4053..4072)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4107..4126
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4186..4202)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4186..4202)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4199..4221)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4210..4226
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4234..4240,4241..4258,4259..4264))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4279..4300
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4417..4434)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4588..5176)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4668..4687)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5347..6138,6139..6207))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5970..5989
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6208..6312)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 atggtgtcca agggagagga gctgttcacc ggagtggtgc caatcctggt ggagctggac
      961 ggcgatgtga atggccacaa gttttctgtg cggggagagg gagagggaga cgcaaccaac
     1021 ggcaagctga cactgaagtt catctgcacc acaggcaagc tgcccgtgcc ttggccaacc
     1081 ctggtgacca cactgagcca cggcgtgcag tgttttgcca gatacccaga ccacatgaag
     1141 cagcacgatt tctttaagtc cgccatgcca gagggatacg tgcaggagcg caccatcttc
     1201 tttaaggacg atggcaccta taagacacgg gccgaggtga agttcgaggg cgatacactg
     1261 gtgaacagaa tcgagctgaa gggcgtggac ttcaaggagg atggcaatat cctgggccac
     1321 aagctggagt acaactttaa ttcccacaac atctatatca tggccgtgaa gcagaagaac
     1381 ggcatcaagg tgaacttcaa gatcaggcac aatgtggagg acggctctgt gcagctggcc
     1441 gatcactacc agcagaacac ccctatcgga gacggacccg tgctgctgcc tgatagccac
     1501 tatctgtcca cacagtctgt gctgagcaag gacccaaatg agaagagaga tcacatggtg
     1561 ctgctggagt tcaggaccgc agcaggaatc acactgggaa tggacgagct gtacaagtcc
     1621 ggactgcgct ctcgggccca ggccagcaac tccatggaga aggagaggaa gcggagagag
     1681 gaggatgagc agaggcgccg gaaggaggag gaggagagaa ggatgaagct ggagatggag
     1741 gccaagcgca agcaggaaga agaggagaga aagaagaggg aggacgatga gaagaggatc
     1801 caggcagaga gccgggccca ggcctctaat agccgggtga tggtgtatat cggcatcgcc
     1861 atcttcctgt ttgtgggcct gtttatgaag taatgactcg agtctagagg gcccgtttaa
     1921 acccgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     1981 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     2041 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     2101 gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct
     2161 atggcttctg aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt
     2221 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     2281 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     2341 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     2401 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
     2461 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     2521 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
     2581 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaattaa
     2641 ttctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
     2701 gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc ccaggctccc
     2761 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata gtcccgcccc
     2821 taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg ccccatggct
     2881 gactaatttt ttttatttat gcagaggccg aggccgcctc tgcctctgag ctattccaga
     2941 agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagctcccg ggagcttgta
     3001 tatccatttt cggatctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     3061 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     3121 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     3181 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
     3241 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     3301 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     3361 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     3421 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     3481 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     3541 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
     3601 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     3661 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     3721 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     3781 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     3841 gagcgggact ctggggttcg aaatgaccga ccaagcgacg cccaacctgc catcacgaga
     3901 tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt tccgggacgc
     3961 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     4021 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     4081 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     4141 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     4201 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     4261 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     4321 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     4381 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4441 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4501 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4561 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4621 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4681 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4741 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4801 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4861 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4921 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     4981 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5041 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5101 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5161 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5221 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5281 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5341 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5401 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5461 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5521 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5581 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5641 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5701 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5761 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5821 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5881 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     5941 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6001 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6061 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6121 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6181 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6241 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6301 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtc
//
