LOCUS       sequence-426359-        7608 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Detection of ER-lipid droplet contact site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7608)
  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 7608)
  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-LD)
FEATURES             Location/Qualifiers
     source          1..7608
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      226..235
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      616..625
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(674..693)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(675..693)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(738..941)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(767..787)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        942..1321
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     1493..1512
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        1587..1691
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1692..1760,1761..2552)
                     /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     complement(1910..1929)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2723..3311
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3212..3231
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3465..3482
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3599..3620
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3635..3640,3641..3658,3659..3665)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3673..3689
                     /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)."
     primer_bind     3678..3700
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3697..3713
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3697..3713
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    3815..3936
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     3880..3899
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5093..5983)
                     /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      5980..5989
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     misc_feature    join(6321..6329,6330..6332,6333..6907)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(6494..6513)
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     primer_bind     6723..6740
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     complement(6977..6996)
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     intron          6979..7208
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     CDS             complement(7454..7495)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
ORIGIN
        1 actcttgggc ctccattaaa tctcgaccat aggcctgatt aaaggatgtt tccttcagag
       61 tctgggggcc ccgttccatc atagcatgca agggctccag cacctcaaac atgcctttca
      121 cgttcctttc cccaaagtac aaacgagatg cctcttccag gccttcatgc cacatctcgc
      181 cccctcctga tccccctccg ccgctgccct cctcgccctt gctcaccatg gtggcgaccg
      241 gtagagatcc ccctccgccg ctgccgcctc ctccggaggt catggccttc ttcatgtgca
      301 ccttcacctt ggtggggcct ctgctggtgg ggatcatcca ctcgaacatg gtgttcaggt
      361 cgcccttctg cacgccctcc ttgaatctgc cctggaactc cttgctctgg gtgccggggg
      421 ccacgtcggt gaagaagttc ttgccgatca cggtcttggg gtctctgccg gtgatggtgc
      481 cctcggcggc gttgtagtgg atgatcttgc cgttgccgtc cagctggatg gcgccgaagg
      541 ccagcttgtc cagggccttg tcgtccatct tggccaggct gttctcgatg tcgtcgccgc
      601 cgaatctcac ggtctccatg gtggctagct aggccgcaga tatcgatccg agctcggtac
      661 caagcttggg tctccctata gtgagtcgta ttaatttcga taagccagta agcagtgggt
      721 tctctagtta gccagagagc tctgcttata tagacctccc accgtacacg cctaccgccc
      781 atttgcgtca atggggcgga gttgttacga cattttggaa agtcccgttg attttggtgc
      841 caaaacaaac tcccattgac gtcaatgggg tggagacttg gaaatccccg tgagtcaaac
      901 cgctatccac gcccattgat gtactgccaa aaccgcatca ccatggtaat agcgatgact
      961 aatacgtaga tgtactgcca agtaggaaag tcccataagg tcatgtactg ggcataatgc
     1021 caggcgggcc atttaccgtc attgacgtca atagggggcg tacttggcat atgatacact
     1081 tgatgtactg ccaagtgggc agtttaccgt aaatactcca cccattgacg tcaatggaaa
     1141 gtccctattg gcgttactat gggaacatac gtcattattg acgtcaatgg gcgggggtcg
     1201 ttgggcggtc agccaggcgg gccatttacc gtaagttatg taacgcggaa ctccatatat
     1261 gggctatgaa ctaatgaccc cgtaattgat tactattaat aactagtcaa taatcaatgt
     1321 caacgcgtat atctggcccg tacatcgcga agcagcgcaa aacgcctaac cctaagcaga
     1381 ttcttcatgc aattgtcggt caagccttgc cttgttgtag cttaaatttt gctcgcgcac
     1441 tactcagcga cctccaacac acaagcaggg agcagatact ggcttaacta tgcggcatca
     1501 gagcagattg tactgagagt gcaccatagg ggatcgggag atctcccgat ccgtcgacgt
     1561 caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac
     1621 attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa
     1681 aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat
     1741 tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc
     1801 agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga
     1861 gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg
     1921 cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc
     1981 agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag
     2041 taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc
     2101 tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg
     2161 taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg
     2221 acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac
     2281 ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac
     2341 cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg
     2401 agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg
     2461 tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg
     2521 agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac
     2581 tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg
     2641 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     2701 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     2761 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     2821 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt
     2881 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     2941 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     3001 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     3061 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     3121 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     3181 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     3241 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     3301 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     3361 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     3421 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     3481 aggaagcgga agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt
     3541 aatgcagctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta
     3601 atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta
     3661 tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt
     3721 acgccaagct ctagctagag gtcgacggta tacagacatg ataagataca ttgatgagtt
     3781 tggacaaacc acaactagaa tgcactcgac gcgttaagat acattgatga gtttggacaa
     3841 accacaacta gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct
     3901 ttatttgtaa ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt
     3961 atgtttcagg ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa
     4021 tgtggtatgg ctgattatga tcagttatct agagttaatt aattggccct cgagttactt
     4081 gtaccgatcc ggtggatcct taggagcgct tggctgccat tagcgcccgg gagaatctct
     4141 ggcaaagcca tacgaacagc aagccgaggt ggaatgcaac cagcctgagt aacgcgaaac
     4201 cgacgaagag cgggtaacta aagtaataga ggttccgctt gtgaggagat tcagaatttg
     4261 cgcttctctt cgcagccatt aaggctcgtg aaaatcgctg gcacaaccag acgaataata
     4321 atcccagatg aaatgctaca agacgtaata gtgcgaaccc tacgaagaga ggatatgaga
     4381 aatagtacag attccgctta tgaggtgatt cgctatttgc ggaacgctta gctgccatca
     4441 aggctcgcga gaagcgctgg cagagccaga cgaatagtag gcctagatgg aaggccacta
     4501 gccggagtag agcaaagcct acaaaaagag ggtaagagaa gtagtataag ttccgcttat
     4561 gcggggattc gctattggca ctcctcttag ccgccattaa agctcgcgaa aatctttggc
     4621 aaagccacac gaacagaaga ccaagatgaa acgcaactag ccgaaggaga gcgaagccaa
     4681 cgaacaacgg gtaagagaag taatagaggt tgcgtttatg aggcgattcg ctatttgccg
     4741 acctctttgc tgccattaac gctcgcgaga accgttgaca taaccagaca aacaacaaac
     4801 ctagatgaaa cgcgaccagc cgaagtagcg cgaaacccac gaacaaggga tagctgaagt
     4861 aatataagtt cctcttatgg ggcgattcac tattagccga acgtttcgca gccatcagcg
     4921 ctctagaaaa gcgttgacac aaccaaacaa acagtaatcc aagatgaaaa gcaaccaatc
     4981 gaagtagagc aaacccgaca aaaagggggt aagagaagta gtacaggttg cgtttgtgcg
     5041 gggattcgga attcgatgct tgagctcgag atctgagtcc ggacttgtac atgccggaaa
     5101 tctcgagcgt cgacagccag cgcgcgatct cgctgccgat caggtccggg ttgtcttctt
     5161 gcagcagatt cagacccggg ccgatgtcca cagccttgca gttaggcagg cttttggcca
     5221 ggcgagcggc ttcggccggt gggatcagaa cgcctggggt gccccagaac agcagcttcg
     5281 ggacagggga ctggtgcagc cagtccatgt attcttcgac cagcgcgacg atgttcgctg
     5341 gctcaccggc gattggcagc tcgtttggga agcgccacag tggctcgcgg tcaacaggat
     5401 tcaggaacgg ctcgcggtaa tggtccatct cgacttcagt cagcgggcgg acgacaccca
     5461 tcggcagcgt accctcgata aaaacgttct gatcgatgat cagcttgcgg ccgacgtcgg
     5521 tggtgcggaa ggcctggaag gtctcgcggg caaattctgg ccattcgtcc caggtcggga
     5581 tagggcggat gaactccata aatgcaatac ctttgacgcg ctctggattg cgcttggccc
     5641 agtggaaacc cagagcggag ccccagtcgt gaatgaccag gacgacctct tccagaccca
     5701 gggcttcgat gaaggcatcc atgaagcgga cgtggtcgtc gaagaaataa cccaggtctg
     5761 gtttgtcgga tttgcccata ccgatcaggt ctggagcaat gcagcgatgg gtcggtgcaa
     5821 catgcgggat gatgttgcgc cacacgtagg aggaggtcgg gttaccgtgc aggaacagca
     5881 caggggtgcc atcgcgcgga ccaacatcga cgtagtgcat gcgctcgccc aggacttcca
     5941 cataatgggg gtcgaatgga aagccagtac cgatttctgc catggtggcg accggtgtcg
     6001 aacctccccc acccgatccg ccacctccag atccacctcc gcctgaaccg ccaccacctg
     6061 atgctctagc tgcagcctcc ctggctgcag cttctctagc agctgcctca cgtgcagctg
     6121 cttccctagc ggcagcctct cttgcggcac cttctagaat tctacctccg gcacttccac
     6181 cagcactgcc tccagctgag accggtaggg tggaacctcc agaaccgccg ctgcctccgg
     6241 agccgcctcc ggtttcggtt tcactagtgg acacccgttt cacaaagacc caatagctgt
     6301 cacccatggc gcgcctcccg ggttgtggca agcttatcat cgtgtttttc aaaggaaaac
     6361 cacgtccccg tggttcgggg ggcctagacg tttttttaac ctcgactaaa cacatgtaaa
     6421 gcatgtgcac cgaggcccca gatcagatcc catacaatgg ggtaccttct gggcatcctt
     6481 cagccccttg ttgaatacgc ttgaggagag ccatttgact ctttccacaa ctatccaact
     6541 cacaacgtgg cactggggtt gtgccgcctt tgcaggtgta tcttatacac gtggcttttg
     6601 gccgcagagg cacctgtcgc caggtggggg gttccgctgc ctgcaaaggg tcgctacaga
     6661 cgttgtttgt cttcaagaag cttccagagg aactgcttcc ttcacgacat tcaacagacc
     6721 ttgcattcct ttggcgagag gggaaagacc cctaggaatg ctcgtcaaga agacagggcc
     6781 aggtttccgg gccctcacat tgccaaaaga cggcaatatg gtggaaaata acatatagac
     6841 aaacgcacac cggccttatt ccaagcggct tcggccagta acgttagggg ggggggaggg
     6901 agaggggggg ggggagaggg gcggaattgg ccgccctaga tgcatgctcg acctgcagtt
     6961 ggacctggga gtggacacct gtggagagaa aggcaaagtg gatgtcattg tcactcaagt
     7021 gtatggccag atctcaagcc tgccacacct caagcttgac aacaaaaaga ttgtcttttc
     7081 tgaccagatg gacgcggcca ccctcaaagg catcaccgcg ggccaggtga atatcaaatc
     7141 ctcctcgttt ttggaaactg acaatcttag cgcagaagtc atgcccgctt ttgagaggga
     7201 gtactcaccc caacagctgg ccctcgcaga cagcgaatta attccagcac actggatcag
     7261 ttatctatgc ggccgcttag tcctcggcca tgtacaggcg gtacatcagg gccaccacca
     7321 gggcggagat ggcggggatc acccagttgg tccaccagga ggagttggac tccacggtgg
     7381 tgatcgagga attcgaagct tgagctcgag atctgagtcc ggacttgtac cgctcgtcca
     7441 tgccattaag tttggtgctg tccaggccca gcagggggtt ggggatgggc ttgccaccgc
     7501 tcccactacc tccgctgcca cttgagattc gtcggaacac atgataatag aggtcccagg
     7561 cttgggtgag gtccttgaca ttccctgatt tcatgtactt cctgcacc
//
