LOCUS       sequence-458400-        6892 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Stabilizes the ER-mitochondrial distance at 30 nm; tagged with mRFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6892)
  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 6892)
  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: pcDNA3.1-30nmEML-mRFP
FEATURES             Location/Qualifiers
     source          1..6892
                     /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             1450..2121
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="ASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAKL
                     KVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGVV
                     TVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKMR
                     LKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHST
                     GA"
     primer_bind     complement(1577..1595)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(2483..2500)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2489..2713
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2759..3187
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2846..2865)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3056..3077
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3196..3216)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3201..3530
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3381..3516
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3443..3462
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3597..4391
                     /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(3651..3670)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4261..4280
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4565..4686
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4602..4621)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4656..4675
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4735..4751)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4735..4751)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4748..4770)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4759..4775
                     /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(4783..4789,4790..4807,4808..4813))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4828..4849
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4966..4983)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5137..5725)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5217..5236)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5896..6687,6688..6756))
                     /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     6519..6538
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6757..6861)
                     /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 atggctattc agctgcgctc actgtttcct ctggctctgc ctggactgct ggccctgctg
      961 gggtggtggt ggttcttttc ccgcaaaaag gaccccaccc gctccgccaa caagcagcag
     1021 gaagaggagg cagagcggct gcggagaatc caggaggaga tggagaagga gagaaagagg
     1081 cgcgaggagg atgagcagcg gagaaggaaa gaggaggagg agcgccggat gaagctggag
     1141 atggaggcca agcggaagca ggaggaggag gagagaaaga agagggagga cgatgagaag
     1201 agaatccagg ccgagaaaca gcaggaagag gaggccgaga ggctgagaag gattcaggaa
     1261 gagatggaga aggaaaggaa gaggagggag gaggacgagc agagaaggcg caaggaggag
     1321 gaggagcgga gaatgaaact ggaaatggaa gctaaaagaa aacaggaaga ggaggagcgc
     1381 aagaagcggg aagatgatga gaagaggatc caggcagaga ggaacagcga cccacctgtg
     1441 gccacactgg ccagctccga ggatgtgatc aaggagttca tgcgctttaa agtgcggatg
     1501 gagggctccg tgaatggcca cgagttcgag atcgagggag agggagaggg cagaccctac
     1561 gagggcaccc agacagccaa gctgaaggtg accaagggag gaccactgcc cttcgcctgg
     1621 gacatcctgt ctcctcagtt tcagtacggc agcaaggcct atgtgaagca ccctgccgac
     1681 atcccagatt atctgaagct gtctttccca gagggcttta agtgggagag ggtgatgaac
     1741 tttgaggatg gcggcgtggt gaccgtgaca caggactcta gcctgcagga tggcgagttc
     1801 atctacaagg tgaagctgcg cggcacaaat tttccctccg acggccctgt gatgcagaag
     1861 aagaccatgg gctgggaggc ctctacagag aggatgtatc ctgaggatgg cgccctgaag
     1921 ggcgagatca agatgcgcct gaagctgaag gacggcggcc actacgatgc cgaggtgaag
     1981 accacatata tggccaagaa gccagtgcag ctgccaggag cctacaagac cgacatcaag
     2041 ctggatatca caagccacaa cgaggactac accatcgtgg agcagtatga gagagccgag
     2101 ggcaggcaca gcaccggagc ctccggcctg cgctctcggg cacaggcctc caattctaaa
     2161 cagcaggaag aagaggcaga gcggctgagg cgcattcagg aagaaatgga aaaagagaga
     2221 aagcggagag aggaggatga gcagaggcgc cggaaagaag aggaggagag aagaatgaaa
     2281 ctggagatgg aagctaaaag aaagcaggaa gaagaagaaa gaaagaagag ggaagatgac
     2341 gaaaagagaa tccaggcaga gagcagggca caggcctcca attcaagggt gatggtctac
     2401 attgggattg ctatttttct gtttgtcgga ctgtttatga agtgactcga gtctagaggg
     2461 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt
     2521 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     2581 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     2641 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     2701 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac
     2761 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     2821 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     2881 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     2941 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca
     3001 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3061 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa
     3121 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     3181 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3241 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3301 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     3361 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3421 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc
     3481 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg
     3541 gagcttgtat atccattttc ggatctgatc aagagacagg atgaggatcg tttcgcatga
     3601 ttgaacaaga tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct
     3661 atgactgggc acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc
     3721 aggggcgccc ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg
     3781 acgaggcagc gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg
     3841 acgttgtcac tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc
     3901 tcctgtcatc tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc
     3961 ggctgcatac gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg
     4021 agcgagcacg tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc
     4081 atcaggggct cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg
     4141 aggatctcgt cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc
     4201 gcttttctgg attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag
     4261 cgttggctac ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg
     4321 tgctttacgg tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg
     4381 agttcttctg agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc
     4441 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4501 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4561 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4621 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     4681 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     4741 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     4801 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     4861 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     4921 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     4981 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5041 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5101 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5161 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5221 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5281 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5341 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5401 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5461 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     5521 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5581 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     5641 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     5701 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     5761 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     5821 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     5881 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     5941 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6001 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6061 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6121 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6181 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6241 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6301 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6361 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6421 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6481 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     6541 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6601 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     6661 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     6721 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     6781 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     6841 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg tc
//
