LOCUS       sequence-458402-        6688 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stabilizes the ER-mitochondrial distance at 20 nm; tagged with mRFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6688)
  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 6688)
  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: pcDNA3.1-20nmEML-mRFP
FEATURES             Location/Qualifiers
     source          1..6688
                     /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"
     CDS             1246..1917
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKMR
                     LKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHST
                     GA"
     primer_bind     complement(2279..2296)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2285..2509
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2555..2983
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2642..2661)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2852..2873
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2992..3012)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2997..3326
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3177..3312
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3239..3258
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3393..4187
                     /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(3447..3466)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4057..4076
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4361..4482
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4398..4417)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4452..4471
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4531..4547)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4531..4547)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4544..4566)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4555..4571
                     /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(4579..4585,4586..4603,4604..4609))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4624..4645
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4762..4779)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4933..5521)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5013..5032)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5692..6483,6484..6552))
                     /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     6315..6334
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6553..6657)
                     /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 atggcgattc aactgcgtag cctgtttccg ctggcgctgc cgggtctgct ggcgctgctg
      961 ggttggtggt ggttctttag ccgtaagaag gacccgaccc gtagcgcgaa caaacagcaa
     1021 gaggaagagg cggagcgtct gcgtcgtatc caggaagaga tggagaagga acgtaaacgt
     1081 cgtgaagagg atgaacaacg tcgtcgtaag gaagaggaag agcgtcgtat gaaactggag
     1141 atggaagcga agcgtaaaca ggaagaggaa gagcgtaaga aacgtgaaga cgatgagaag
     1201 cgtatccaag cggagcgtaa cagcgacccg ccggtggcga ccctggcgag cagcgaggat
     1261 gttatcaagg aattcatgcg tttcaaagtg cgtatggagg gcagcgttaa cggtcacgag
     1321 ttcgaaattg agggtgaagg cgagggtcgt ccgtacgaag gtacccagac cgcgaagctg
     1381 aaagtgacca aaggtggccc gctgccgttt gcgtgggaca tcctgagccc gcagtttcaa
     1441 tacggcagca aggcgtatgt taaacacccg gcggacattc cggattatct gaagctgagc
     1501 ttcccggagg gttttaaatg ggaacgtgtg atgaactttg aggatggtgg cgtggttacc
     1561 gttacccagg acagcagcct gcaagatggc gaattcatct acaaggtgaa actgcgtggc
     1621 accaactttc cgagcgacgg tccggttatg cagaagaaaa ccatgggttg ggaggcgagc
     1681 accgaacgta tgtatccgga ggatggcgcg ctgaagggtg aaattaaaat gcgtctgaag
     1741 ctgaaagacg gtggccacta cgatgcggag gtgaagacca cctatatggc gaagaaaccg
     1801 gttcaactgc cgggcgcgta caagaccgac atcaaactgg atattaccag ccacaacgaa
     1861 gactacacca tcgtggaaca gtatgagcgt gcggaaggtc gtcacagcac cggtgcgagc
     1921 ggtctgcgta gccgtgcgca agcgagcaac agcaaacagc aagaagagga agcggagcgt
     1981 ctgcgccgca ttcaggagga aatggaaaag gagcgtaaac gtcgtgagga agacgagcaa
     2041 cgtcgtcgta aggaggaaga ggaacgtcgt atgaaactgg aaatggaggc gaagcgtaaa
     2101 caggaggaag aggagcgtaa gaaacgtgag gacgatgaaa agcgtatcca ggcggagagc
     2161 cgtgcgcagg cgagcaacag ccgtgttatg gtgtatattg gtattgcgat ttttctgttt
     2221 gttggtctgt ttatgaagta actcgagtct agagggcccg tttaaacccg ctgatcagcc
     2281 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     2341 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     2401 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     2461 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     2521 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     2581 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     2641 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     2701 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     2761 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     2821 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     2881 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     2941 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt
     3001 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3061 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3121 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3181 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3241 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     3301 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat
     3361 ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     3421 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     3481 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     3541 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
     3601 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     3661 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     3721 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     3781 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     3841 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     3901 tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg
     3961 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     4021 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     4081 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     4141 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     4201 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     4261 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     4321 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     4381 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4441 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     4501 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     4561 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4621 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4681 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4741 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4801 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4861 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4921 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4981 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5041 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5101 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5161 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5221 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5281 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5341 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5401 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5461 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     5521 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     5581 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5641 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     5701 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     5761 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     5821 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     5881 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     5941 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6001 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6061 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6121 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6181 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6241 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6301 ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg
     6361 aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat ccagttcgat
     6421 gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca gcgtttctgg
     6481 gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga cacggaaatg
     6541 ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg gttattgtct
     6601 catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg ttccgcgcac
     6661 atttccccga aaagtgccac ctgacgtc
//
