LOCUS       sequence-426328-        7508 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Detection of ER-mito contact site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7508)
  AUTHORS   Li X, Gamuyao R, Wu ML, Cho WJ, King SV, Petersen RA, Stabley DR, 
            Lindow C, Climer LK, Shirinifard A, Ferrara F, Throm RE, Robinson 
            CG, Zhou Y, Carisey AF, Tebo AG, Chang CL
  TITLE     A fluorogenic complementation tool kit for interrogating lipid 
            droplet-organelle interaction.
  JOURNAL   J Cell Biol. 2024 Sep 2;223(9):e202311126. doi: 
            10.1083/jcb.202311126. Epub 2024 Jul 1.
  PUBMED    38949658
REFERENCE   2  (bases 1 to 7508)
  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: "J Cell 
            Biol."; date: "2024-09-2"; volume: "223(9):e202311126. doi"; pages: 
            " 10.1083/jcb.202311126. Epub 2024 Jul 1"
COMMENT     Sequence Label: FAB(ER-mito)
FEATURES             Location/Qualifiers
     source          1..7508
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      154..163
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    join(190..198,199..201,202..776)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(363..382)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     592..609
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     complement(846..865)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          848..1077
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     CDS             complement(1182..1472)
                     /codon_start=1
                     /gene="SEC61B"
                     /product="human Sec61-beta subunit of the translocon in the
                     endoplasmic reticulum"
                     /label=Sec61-beta
                     /translation="MPGPTPSGTNVGSSGRSPSKAVAARAAGSTVRQRKNASCGTRSAG
                     RTTSAGTGGMWRFYTEDSPGLKVGPVPVLVMSLLFIASVFMLHIWGKYTRS"
     CDS             complement(1518..1559)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     regulatory      1898..1907
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2288..2297
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2324..2527)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2353..2373)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2528..2831
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3006..3594)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3086..3105)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3923..3970
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3995..4014
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(4202..4996)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(4202..4996)
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4313..4332)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4923..4942
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(5031..5388)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5045..5180
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5197..5388
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5345..5365
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        complement(5390..5494)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5521..5976
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5608..5627)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5818..5839
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    5983..6104
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5994..6013)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6048..6067
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6457..7347)
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     regulatory      7344..7353
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
ORIGIN
        1 cggcaccttc tagaattcta cctccggcac ttccaccagc actgcctcca gctgagaccg
       61 gtagggtgga acctccagaa ccgccgctgc ctccggagcc gcctccggtt tcggtttcac
      121 tagtccgttt cacaaagacc caatagctgt cacccatggt ggctagcgga tctgacggcg
      181 cgcctcccgg gttgtggcaa gcttatcatc gtgtttttca aaggaaaacc acgtccccgt
      241 ggttcggggg gcctagacgt ttttttaacc tcgactaaac acatgtaaag catgtgcacc
      301 gaggccccag atcagatccc atacaatggg gtaccttctg ggcatccttc agccccttgt
      361 tgaatacgct tgaggagagc catttgactc tttccacaac tatccaactc acaacgtggc
      421 actggggttg tgccgccttt gcaggtgtat cttatacacg tggcttttgg ccgcagaggc
      481 acctgtcgcc aggtgggggg ttccgctgcc tgcaaagggt cgctacagac gttgtttgtc
      541 ttcaagaagc ttccagagga actgcttcct tcacgacatt caacagacct tgcattcctt
      601 tggcgagagg ggaaagaccc ctaggaatgc tcgtcaagaa gacagggcca ggtttccggg
      661 ccctcacatt gccaaaagac ggcaatatgg tggaaaataa catatagaca aacgcacacc
      721 ggccttattc caagcggctt cggccagtaa cgttaggggg gggggaggga gagggggggg
      781 gggagagggg cggaattggc cgccctagat gcatgctcga cctgcagttg gacctgggag
      841 tggacacctg tggagagaaa ggcaaagtgg atgtcattgt cactcaagtg tatggccaga
      901 tctcaagcct gccacacctc aagcttgaca acaaaaagat tgtcttttct gaccagatgg
      961 acgcggccac cctcaaaggc atcaccgcgg gccaggtgaa tatcaaatcc tcctcgtttt
     1021 tggaaactga caatcttagc gcagaagtca tgcccgcttt tgagagggag tactcacccc
     1081 aacagctggc cctcgcagac agcgaattaa ttccagcaca ctggatcagt tatctatgcg
     1141 gccgcttagt cctcggccat gtaccgtcga ctgcagaatt cctacgaacg agtgtacttg
     1201 ccccaaatgt gcaacataaa tacagaagcg atgaacagaa gactcataac caatactgga
     1261 acagggccaa ctttgagccc aggtgaatct tctgtgtaga atcgccacat ccccccggtg
     1321 cctgccgagg ttgtgcggcc tgcactcctt gtcccacagc tggcattttt cctctgccgg
     1381 acagtggatc ccgccgcccg ggcggccact gctttgctgg gagagcgccc tgaggatccc
     1441 acgttagtgc cactgggggt cggaccaggc atagatctga gtccggactt gtacagctcg
     1501 tccatgccat taagtttggt gctgtccagg cccagcaggg ggttggggat gggcttgcca
     1561 ccgctcccac tacctccgct gccacttgag attcgtcgga acacatgata atagaggtcc
     1621 caggcttggg tgaggtcctt gacattccct gatttcatgt acttcctgca ccactcttgg
     1681 gcctccatta aatctcgacc ataggcctga ttaaaggatg tttccttcag agtctggggg
     1741 ccccgttcca tcatagcatg caagggctcc agcacctcaa acatgccttt cacgttcctt
     1801 tccccaaagt acaaacgaga tgcctcttcc aggccttcat gccacatctc gccccctcct
     1861 gatccccctc cgccgctgcc ctcctcgccc ttgctcacca tggtggcgac cggtagagat
     1921 ccccctccgc cgctgccgcc tcctccggag gtcatggcct tcttcatgtg caccttcacc
     1981 ttggtggggc ctctgctggt ggggatcatc cactcgaaca tggtgttcag gtcgcccttc
     2041 tgcacgccct ccttgaatct gccctggaac tccttgctct gggtgccggg ggccacgtcg
     2101 gtgaagaagt tcttgccgat cacggtcttg gggtctctgc cggtgatggt gccctcggcg
     2161 gcgttgtagt ggatgatctt gccgttgccg tccagctgga tggcgccgaa ggccagcttg
     2221 tccagggcct tgtcgtccat cttggccagg ctgttctcga tgtcgtcgcc gccgaatctc
     2281 acggtctcca tggtggctag cggatctgac ggttcactaa accagctctg cttatataga
     2341 cctcccaccg tacacgccta ccgcccattt gcgtcaatgg ggcggagttg ttacgacatt
     2401 ttggaaagtc ccgttgattt tggtgccaaa acaaactccc attgacgtca atggggtgga
     2461 gacttggaaa tccccgtgag tcaaaccgct atccacgccc attgatgtac tgccaaaacc
     2521 gcatcaccat ggtaatagcg atgactaata cgtagatgta ctgccaagta ggaaagtccc
     2581 ataaggtcat gtactgggca taatgccagg cgggccattt accgtcattg acgtcaatag
     2641 ggggcgtact tggcatatga tacacttgat gtactgccaa gtgggcagtt taccgtaaat
     2701 actccaccca ttgacgtcaa tggaaagtcc ctattggcgt tactatggga acatacgtca
     2761 ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc aggcgggcca tttaccgtaa
     2821 gttatgtaac gcggaactcc atatatgggc tatgaactaa tgaccccgta attgattact
     2881 attaataact aatgcatggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     2941 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3001 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3061 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3121 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3181 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3241 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     3301 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     3361 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     3421 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     3481 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     3541 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     3601 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     3661 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     3721 tttaaatcaa tctaaagtat atatgagtaa cctgaggcta tggcagggcc tgccgccccg
     3781 acgttggctg cgagccctgg gccttcaccc gaacttgggg ggtggggtgg ggaaaaggaa
     3841 gaaacgcggg cgtattggcc ccaatggggt ctcggtgggg tatcgacaga gtgccagccc
     3901 tgggaccgaa ccccgcgttt atgaacaaac gacccaacac cgtgcgtttt attctgtctt
     3961 tttattgccg tcatagcgcg ggttccttcc ggtattgtct ccttccgtgt ttcagttagc
     4021 ctccccctag ggtgggcgaa gaactccagc atgagatccc cgcgctggag gatcatccag
     4081 ccggcgtccc ggaaaacgat tccgaagccc aacctttcat agaaggcggc ggtggaatcg
     4141 aaatctcgtg atggcaggtt gggcgtcgct tggtcggtca tttcgaaccc cagagtcccg
     4201 ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga
     4261 taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac
     4321 gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga
     4381 atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca
     4441 cgacgagatc ctcgccgtcg ggcatgctcg ccttgagcct ggcgaacagt tcggctggcg
     4501 cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag
     4561 tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa
     4621 gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt
     4681 gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt
     4741 cagtgacaac gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc
     4801 gcgctgcctc gtcttgcagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa
     4861 ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt
     4921 gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc
     4981 catcttgttc aatcatgcga aacgatcctc atcctgtctc ttgatcgatc tttgcaaaag
     5041 cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc cgaggcggcc
     5101 tcggcctctg cataaataaa aaaaattagt cagccatggg gcggagaatg ggcggaactg
     5161 ggcggagtta ggggcgggat gggcggagtt aggggcggga ctatggttgc tgactaattg
     5221 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acacctggtt
     5281 gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt
     5341 ccacacccta actgacacac attccacagc tggttctttc cgcctcagga ctcttccttt
     5401 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     5461 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     5521 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     5581 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     5641 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     5701 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     5761 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     5821 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     5881 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     5941 acgcgaattt taacaaaata ttaacgctta caatttacgc gttaagatac attgatgagt
     6001 ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa atttgtgatg
     6061 ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac aacaattgca
     6121 ttcattttat gtttcaggtt cagggggagg tgtgggaggt tttttaaagc aagtaaaacc
     6181 tctacaaatg tggtatggct gattatgatc agttatctag agttaattaa ttggccctcg
     6241 agttacttgt accgatccgg tggatcctta tcaaagttgt tgccggtatc tcatgaaagc
     6301 ccaggctgct accatggtga gggcaaacac tggcaccagc accataaata tgggaatacc
     6361 acttgggtcc ccttcacctc gatgtcctat aggtccattc tgcacctgca aagaattcga
     6421 agcttgagct cgagatctga gtccggactt gtacatgccg gaaatctcga gcgtcgacag
     6481 ccagcgcgcg atctcgctgc cgatcaggtc cgggttgtct tcttgcagca gattcagacc
     6541 cgggccgatg tccacagcct tgcagttagg caggcttttg gccaggcgag cggcttcggc
     6601 cggtgggatc agaacgcctg gggtgcccca gaacagcagc ttcgggacag gggactggtg
     6661 cagccagtcc atgtattctt cgaccagcgc gacgatgttc gctggctcac cggcgattgg
     6721 cagctcgttt gggaagcgcc acagtggctc gcggtcaaca ggattcagga acggctcgcg
     6781 gtaatggtcc atctcgactt cagtcagcgg gcggacgaca cccatcggca gcgtaccctc
     6841 gataaaaacg ttctgatcga tgatcagctt gcggccgacg tcggtggtgc ggaaggcctg
     6901 gaaggtctcg cgggcaaatt ctggccattc gtcccaggtc gggatagggc ggatgaactc
     6961 cataaatgca atacctttga cgcgctctgg attgcgcttg gcccagtgga aacccagagc
     7021 ggagccccag tcgtgaatga ccaggacgac ctcttccaga cccagggctt cgatgaaggc
     7081 atccatgaag cggacgtggt cgtcgaagaa ataacccagg tctggtttgt cggatttgcc
     7141 cataccgatc aggtctggag caatgcagcg atgggtcggt gcaacatgcg ggatgatgtt
     7201 gcgccacacg taggaggagg tcgggttacc gtgcaggaac agcacagggg tgccatcgcg
     7261 cggaccaaca tcgacgtagt gcatgcgctc gcccaggact tccacataat gggggtcgaa
     7321 tggaaagcca gtaccgattt ctgccatggt ggcgaccggt gtcgaacctc ccccacccga
     7381 tccgccacct ccagatccac ctccgcctga accgccacca cctgatgctc tagctgcagc
     7441 ctccctggct gcagcttctc tagcagctgc ctcacgtgca gctgcttccc tagcggcagc
     7501 ctctcttg
//
