LOCUS       sequence-323265-        7733 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Detect the long Peroxisomes-mitochondria-Endoplasmic reticulum 
            contact .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7733)
  AUTHORS   Vallese F, Catoni C, Cieri D, Barazzuol L, Ramirez O, Calore V, 
            Bonora M, Giamogante F, Pinton P, Brini M, Cali T
  TITLE     An expanded palette of improved SPLICS reporters detects multiple 
            organelle contacts in vitro and in vivo.
  JOURNAL   Nat Commun. 2020 Nov 27;11(1):6069. doi: 10.1038/s41467-020-19892-6.
  PUBMED    33247103
REFERENCE   2  (bases 1 to 7733)
  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: "Nat 
            Commun."; date: "2020-11-27"; volume: "11(1):6069. doi"; pages: " 
            10.1038/s41467-020-19892-6"
COMMENT     Sequence Label: SPLICS Mt-ER- Po Long P2A
FEATURES             Location/Qualifiers
     source          1..7733
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(49..68)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        240..619
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        620..823
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     774..794
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     868..887
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        868..886
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             925..972
                     /codon_start=1
                     /product="11th beta-strand of superfolder GFP"
                     /label=GFP11
                     /note="associates with GFP(1-10), consisting of the first 
                     10 beta-strands, to reconstitute fluorescent GFP (Cabantous
                     et al., 2005; Kamiyama et al., 2016)"
                     /translation="RDHMVLHEYVNAAGIT"
     CDS             1489..1545
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2335..2391
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     primer_bind     complement(3329..3346)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3335..3559
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3605..4033
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3692..3711)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3902..3923
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4042..4062)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4047..4376
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4227..4362
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4289..4308
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4443..5237
                     /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(4497..4516)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5107..5126
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5411..5532
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5448..5467)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5502..5521
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5581..5597)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5581..5597)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5594..5616)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5605..5621
                     /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(5629..5635,5636..5653,5654..5659))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5674..5695
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5812..5829)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5983..6571)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6063..6082)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6742..7533,7534..7602))
                     /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     7365..7384
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7603..7707)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
       61 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
      121 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa
      181 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg
      241 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
      301 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
      361 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
      421 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
      481 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
      541 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
      601 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
      661 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
      721 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
      781 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga
      841 acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg
      901 ctagcgttta aacttaagct tatgcgggac cacatggtgc tgcacgagta cgtgaacgcc
      961 gccggcatca ccggaggcga tggaggctcc ggaggaggct ctaagctgaa caattctggc
     1021 gcccctcaca gggagaagag gggaggagag accgacgagt tctccaatgt gcggagaggc
     1081 cggggccaca gaatgcagca cctgtctgag ggcacaaagg gaaggcaagt gggctctgga
     1141 ggcgacggag agagatgggg cagcgatagg ggctctcgcg gcagcctgaa cgagcagatc
     1201 gccctggtgc tgatgaggct gcaggaggat atgcagaatg tgctgcagcg cctgcagaag
     1261 ctggagaccc tgacagccct gcaggccaag agctccacca gcacactgca gaccgcacca
     1321 cagcctacaa gccagaggcc atcctggtgg ccctttgaga tgtcccctgg cgtgctgaca
     1381 ttcgccatca tctggccctt catcgcccag tggctggtgt atctgtacta tcagaggcgc
     1441 cggagaaagc tgaacggatc cctgaaggag ctggcatgcg gcagcggagc aaccaacttc
     1501 tccctgctga agcaggcagg cgacgtggag gagaatcccg gccctcggtc tgaattcatg
     1561 gtgggccgca attccgcaat cgcagcagga gtgtgcggcg ccctgttcat cggctactgt
     1621 atctattttg accggaagcg gagaagcgat ccaaattcca agggcgagga gctgttcaca
     1681 ggcgtggtgc ccatcctggt ggagctggac ggcgatgtga acggccacaa gttttctgtg
     1741 cggggagagg gagagggcga cgcaaccatc ggcaagctga cactgaagtt catctgcacc
     1801 acaggcaagc tgcccgtgcc ttggccaacc ctggtgacca cactgacata cggcgtgcag
     1861 tgttttagca ggtatcctga ccacatgaag cgccacgatt tctttaagtc cgccatgcca
     1921 gagggctacg tgcaggagcg gaccatctct ttcaaggacg atggcaagta taagaccaga
     1981 gccgtggtga agtttgaggg cgacacactg gtgaacagga tcgagctgaa gggcaccgac
     2041 ttcaaggagg atggcaatat cctgggccac aagctggagt acaactttaa tagccacaac
     2101 gtgtatatca cagccgataa gcagaagaac ggcatcaagg ccaatttcac cgtgagacac
     2161 aatgtggagg acggctccgt gcagctggcc gatcactacc agcagaacac accaatcggc
     2221 gacggccctg tgctgctgcc tgataatcac tatctgtcta cccagacagt gctgagcaag
     2281 gaccccaacg agaagggcac cctcgagctg aaggagctgg catgcggcag cggagcaacc
     2341 aacttctccc tgctgaagca ggcaggcgac gtggaggaga atcccggccc tcggtctggg
     2401 cccatgagca agggagagga gctgttcacc ggagtggtgc ctatcctggt ggagctggac
     2461 ggcgatgtga atggccacaa gttttccgtg cgcggagagg gagagggcga cgcaacaatc
     2521 ggcaagctga ccctgaagtt catctgcacc acaggcaagc tgcccgtgcc ttggccaacc
     2581 ctggtgacca cactgggcta cggcctgcag tgttttgccc ggtatccaga ccacatgaag
     2641 agacacgatt tctttaagtc cgccatgccc gagggctacg tgcaggagag gacaatctct
     2701 ttcaaggacg atggcaagta taagacacgc gccgtggtga agtttgaggg cgacaccctg
     2761 gtgaaccgga tcgagctgaa gggcacagac ttcaaggagg atggcaatat cctgggccac
     2821 aagctggagt acaactttaa ttcccacaac gtgtatatca ccgccgataa gcagaagaac
     2881 ggcatcaagg ccaatttcac agtgcggcac aatgtggagg acggctctgt gcagctggcc
     2941 gatcactacc agcagaacac ccctatcggc gacggccctg tgctgctgcc tgataatcac
     3001 tacctgtctt atcagaccgt gctgagcaag gacccaaacg agaagggcac aggaggcgat
     3061 ggaggcagcg gaggaggctc caagctgaga gtgttcctgg ccctgcccat catcatggtg
     3121 gtggcctttt ctatgtgcat catctgtctg ctgatggcag gcgacacctg gacagagacc
     3181 ctggcctacg tgctgttttg gggcgtggcc agcatcggca cattctttat catcctgtat
     3241 aacggcaagg acttcgtgga tgcccccagg ctggtgcaga aggagaagat cgattgatct
     3301 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     3361 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3421 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     3481 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     3541 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     3601 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3661 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3721 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3781 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3841 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3901 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3961 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4021 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     4081 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     4141 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4201 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4261 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     4321 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     4381 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     4441 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     4501 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     4561 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     4621 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     4681 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     4741 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     4801 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     4861 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     4921 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     4981 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     5041 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     5101 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     5161 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     5221 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     5281 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     5341 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     5401 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5461 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5521 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     5581 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     5641 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     5701 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5761 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     5821 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     5881 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     5941 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6001 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6061 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     6121 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     6181 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     6241 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     6301 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     6361 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     6421 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     6481 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     6541 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     6601 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     6661 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     6721 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     6781 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     6841 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     6901 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
     6961 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
     7021 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
     7081 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
     7141 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
     7201 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
     7261 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
     7321 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
     7381 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
     7441 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
     7501 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
     7561 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
     7621 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
     7681 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctg
//
