LOCUS       sequence-323258-        7397 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Detect the short Peroxisomes-mitochondria-Endoplasmic reticulum 
            contact .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7397)
  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 7397)
  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 Short P2A
FEATURES             Location/Qualifiers
     source          1..7397
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(73..92)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        264..643
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        644..847
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     798..818
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     892..911
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        892..910
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             949..996
                     /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             1177..1233
                     /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             2023..2079
                     /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(3017..3034)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3023..3247
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3293..3721
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3380..3399)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3590..3611
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3730..3750)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3735..4064
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3915..4050
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3977..3996
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4131..4925
                     /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(4185..4204)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4795..4814
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5099..5220
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5136..5155)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5190..5209
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5269..5285)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5269..5285)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5282..5304)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5293..5309
                     /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(5317..5323,5324..5341,5342..5347))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5362..5383
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5500..5517)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5671..6259)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5751..5770)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6430..7221,7222..7290))
                     /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     7053..7072
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7291..7395)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
       61 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
      121 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
      181 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
      241 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
      301 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      361 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      421 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      481 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      541 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      601 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      661 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
      721 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
      781 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
      841 cagagctctc tggctaacta gagaacccac tgcttactgg cttatcgaaa ttaatacgac
      901 tcactatagg gagacccaag ctggctagcg tttaaactta agcttatgcg ggaccacatg
      961 gtgctgcacg agtacgtgaa cgccgccggc atcaccggag gcgatggagg cagcggagga
     1021 ggctccaagc tgtggtggcc cttcgagatg tctcctggcg tgctgacatt cgccatcatc
     1081 tggcccttta tcgcccagtg gctggtgtat ctgtactatc agcggagaag gcgcaagctg
     1141 aatggatccc tgaaggagct ggcatgcggc agcggagcaa ccaacttctc cctgctgaag
     1201 caggcaggcg acgtggagga gaatcccggc cctcggtctg aattcatggt gggccgcaat
     1261 tccgcaatcg cagcaggagt gtgcggcgcc ctgttcatcg gctactgtat ctattttgac
     1321 cggaagcgga gaagcgatcc aaattccaag ggcgaggagc tgttcacagg cgtggtgccc
     1381 atcctggtgg agctggacgg cgatgtgaac ggccacaagt tttctgtgcg gggagaggga
     1441 gagggcgacg caaccatcgg caagctgaca ctgaagttca tctgcaccac aggcaagctg
     1501 cccgtgcctt ggccaaccct ggtgaccaca ctgacatacg gcgtgcagtg ttttagcagg
     1561 tatcctgacc acatgaagcg ccacgatttc tttaagtccg ccatgccaga gggctacgtg
     1621 caggagcgga ccatctcttt caaggacgat ggcaagtata agaccagagc cgtggtgaag
     1681 tttgagggcg acacactggt gaacaggatc gagctgaagg gcaccgactt caaggaggat
     1741 ggcaatatcc tgggccacaa gctggagtac aactttaata gccacaacgt gtatatcaca
     1801 gccgataagc agaagaacgg catcaaggcc aatttcaccg tgagacacaa tgtggaggac
     1861 ggctccgtgc agctggccga tcactaccag cagaacacac caatcggcga cggccctgtg
     1921 ctgctgcctg ataatcacta tctgtctacc cagacagtgc tgagcaagga ccccaacgag
     1981 aagggcaccc tcgagctgaa ggagctggca tgcggcagcg gagcaaccaa cttctccctg
     2041 ctgaagcagg caggcgacgt ggaggagaat cccggccctc ggtctgggcc catgagcaag
     2101 ggagaggagc tgttcaccgg agtggtgcct atcctggtgg agctggacgg cgatgtgaat
     2161 ggccacaagt tttccgtgcg cggagaggga gagggcgacg caacaatcgg caagctgacc
     2221 ctgaagttca tctgcaccac aggcaagctg cccgtgcctt ggccaaccct ggtgaccaca
     2281 ctgggctacg gcctgcagtg ttttgcccgg tatccagacc acatgaagag acacgatttc
     2341 tttaagtccg ccatgcccga gggctacgtg caggagagga caatctcttt caaggacgat
     2401 ggcaagtata agacacgcgc cgtggtgaag tttgagggcg acaccctggt gaaccggatc
     2461 gagctgaagg gcacagactt caaggaggat ggcaatatcc tgggccacaa gctggagtac
     2521 aactttaatt cccacaacgt gtatatcacc gccgataagc agaagaacgg catcaaggcc
     2581 aatttcacag tgcggcacaa tgtggaggac ggctctgtgc agctggccga tcactaccag
     2641 cagaacaccc ctatcggcga cggccctgtg ctgctgcctg ataatcacta cctgtcttat
     2701 cagaccgtgc tgagcaagga cccaaacgag aagggcacag gaggcgatgg aggcagcgga
     2761 ggaggctcca agctgagagt gttcctggcc ctgcccatca tcatggtggt ggccttttct
     2821 atgtgcatca tctgtctgct gatggcaggc gacacctgga cagagaccct ggcctacgtg
     2881 ctgttttggg gcgtggccag catcggcaca ttctttatca tcctgtataa cggcaaggac
     2941 ttcgtggatg cccccaggct ggtgcagaag gagaagatcg attgatctag agggcccgtt
     3001 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc
     3061 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat
     3121 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg
     3181 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc
     3241 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     3301 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     3361 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     3421 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     3481 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     3541 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     3601 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     3661 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     3721 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     3781 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     3841 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     3901 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     3961 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     4021 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     4081 gtatatccat tttcggatct gatcaagaga caggatgagg atcgtttcgc atgattgaac
     4141 aagatggatt gcacgcaggt tctccggccg cttgggtgga gaggctattc ggctatgact
     4201 gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt ccggctgtca gcgcaggggc
     4261 gcccggttct ttttgtcaag accgacctgt ccggtgccct gaatgaactg caggacgagg
     4321 cagcgcggct atcgtggctg gccacgacgg gcgttccttg cgcagctgtg ctcgacgttg
     4381 tcactgaagc gggaagggac tggctgctat tgggcgaagt gccggggcag gatctcctgt
     4441 catctcacct tgctcctgcc gagaaagtat ccatcatggc tgatgcaatg cggcggctgc
     4501 atacgcttga tccggctacc tgcccattcg accaccaagc gaaacatcgc atcgagcgag
     4561 cacgtactcg gatggaagcc ggtcttgtcg atcaggatga tctggacgaa gagcatcagg
     4621 ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg catgcccgac ggcgaggatc
     4681 tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat ggtggaaaat ggccgctttt
     4741 ctggattcat cgactgtggc cggctgggtg tggcggaccg ctatcaggac atagcgttgg
     4801 ctacccgtga tattgctgaa gagcttggcg gcgaatgggc tgaccgcttc ctcgtgcttt
     4861 acggtatcgc cgctcccgat tcgcagcgca tcgccttcta tcgccttctt gacgagttct
     4921 tctgagcggg actctggggt tcgaaatgac cgaccaagcg acgcccaacc tgccatcacg
     4981 agatttcgat tccaccgccg ccttctatga aaggttgggc ttcggaatcg ttttccggga
     5041 cgccggctgg atgatcctcc agcgcgggga tctcatgctg gagttcttcg cccaccccaa
     5101 cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa
     5161 taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta
     5221 tcatgtctgt ataccgtcga cctctagcta gagcttggcg taatcatggt catagctgtt
     5281 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     5341 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     5401 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     5461 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     5521 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     5581 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     5641 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     5701 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     5761 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     5821 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     5881 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5941 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     6001 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     6061 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     6121 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     6181 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     6241 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     6301 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     6361 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     6421 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     6481 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     6541 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     6601 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6661 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6721 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6781 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6841 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6901 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6961 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     7021 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     7081 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7141 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7201 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7261 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7321 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     7381 aataggggtt ccgcgca
//
