LOCUS       sequence-491572-        6776 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of LAMP1-iLID-Halo fusion for light-induced 
            recruitment to late endosomes/lysosomes..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6776)
  AUTHORS   Choi H, Liao YC, Yoon YJ, Grimm J, Wang N, Lavis LD, Singer RH, 
            Lippincott-Schwartz J
  TITLE     Secretome translation shaped by lysosomes and lunapark-marked ER 
            junctions.
  JOURNAL   Nature. 2025 Nov 5. doi: 10.1038/s41586-025-09718-0.
  PUBMED    41193816
REFERENCE   2  (bases 1 to 6776)
  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: "Nature. 
            2025 Nov 5. doi: 10.1038/s41586-025-09718-0."
COMMENT     Sequence Label: pCMV-LAMP1-iLID-Halo
FEATURES             Location/Qualifiers
     source          1..6776
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        764..1067
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1068..1271
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1222..1242
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1322..1331
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1326..1335
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2633..3070
                     /codon_start=1
                     /product="improved light-inducible dimer when used with 
                     SspB nano or SspB micro (Guntas et al., 2015)"
                     /label=iLID
                     /translation="LATTLERIEKNFVITDPRLPDNPIIFASDSFLQLTEYSREEILGR
                     NCRFLQGPETDRATVRKIRDAIDNQTEVTVQLINYTKSGKKFWNVFHLQPMRDYKGDVQ
                     YFIGVQLDGTERLHGAAEREAVCLIKKTAFQIAEAANDENYF"
     CDS             3251..4141
                     /codon_start=1
                     /product="modified bacterial dehalogenase that forms 
                     covalent bonds with chloroalkane derivatives"
                     /label=HaloTag(R)
                     /translation="MAEIGTGFPFDPHYVEVLGERMHYVDVGPRDGTPVLFLHGNPTSS
                     YVWRNIIPHVAPTHRCIAPDLIGMGKSDKPDLGYFFDDHVRFMDAFIEALGLEEVVLVI
                     HDWGSALGFHWAKRNPERVKGIAFMEFIRPIPTWDEWPEFARETFQAFRTTDVGRKLII
                     DQNVFIEGTLPMGVVRPLTEVEMDHYREPFLNPVDREPLWRFPNELPIAGEPANIVALV
                     EEYMDWLHQSPVPKLLFWGTPGVLIPPAEAARLAKSLPNCKAVDIGPGLNLLQEDNPDL
                     IGSEIARWLSTLEISG"
     polyA_signal    4267..4388
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4304..4323)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4358..4377
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4395..4850)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4532..4553)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4744..4763
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4877..4981
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4983..5340
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4983..5174
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(5006..5026)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5191..5326
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5375..6169
                     /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             5375..6169
                     /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(5429..5448)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6039..6058
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6357..6376)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6401..6448
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat
      721 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      781 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      841 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      901 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      961 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
     1021 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     1081 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     1141 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     1201 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     1261 taagcagagc tggtttagtg aaccgtcaga tccgctagcg ctaccggact cagatctcga
     1321 ggccaccatg gcggcccccg gcagcgcccg gcggccgctg ctcctgttgc tgctggcagg
     1381 ccttgcacac agcgccccag cactgttcga ggtgaaagac aacaacggca cagcgtgtat
     1441 aatggccagc ttctctgcct cctttctgac cacctatgag gctggacatg tttctaaggt
     1501 ctcgaatatg accctgccag cctctgcaga agtcctgaag aatagcagct cttgtggtga
     1561 aaagaatgct tctgagccca ccctcgcaat cacctttgga gaaggatatt tactgaaact
     1621 caccttcaca aaaaacacaa cacgttacag tgtccagcac atgtatttca catataacct
     1681 gtcagacaca caattctttc ccaatgccag ctccaaaggg cccgacactg tggattccac
     1741 aactgacatc aaggcagaca tcaacaaaac ataccgatgt gtcagcgaca tcagggtcta
     1801 catgaagaat gtgaccattg tgctctggga cgctactatc caggcctacc tgccgagtag
     1861 caacttcagc aaggaagaga cacgctgccc acaggatcaa ccttccccaa ctactgggcc
     1921 acccagcccc tcaccaccac ttgtgcccac aaaccccagt gtgtccaagt acaatgtgac
     1981 tggtgacaat ggaacctgcc tgctggcctc tatggcactg caactcaaca tcacctacat
     2041 gaagaaggac aacacgactg tgaccagagc attcaacatc aacccaagtg acaaatatag
     2101 tgggacttgc ggtgcccagt tggtgaccct gaaggtgggg aacaagagca gagtcctgga
     2161 gctgcagttt gggatgaatg ccacttctag cctgtttttc ctgcaaggag ttcagttgaa
     2221 catgactctt cctgatgcca tagagcccac gttcagcacc tccaactatt ccctgaaagc
     2281 tcttcaggcc agtgtcggca actcatacaa gtgcaacagt gaggagcaca tctttgtcag
     2341 caaggcgctc gccctcaatg tcttcagcgt gcaagtccag gctttcaggg tagaaagtga
     2401 caggtttggg tctgtggaag agtgtgtaca ggacggtaac aacatgctga tccccattgc
     2461 tgtgggcggg gccctggcag ggctggtcct catcgtcctc atcgcctacc tcatcggcag
     2521 gaagaggagt cacgcgggct atcagaccat ctcggaattc ggctccaccg gctccacagg
     2581 ctccaccggc gcggtcgacg gtggaggaag tggcggaagt ggcggggagt ttctggcaac
     2641 cacactggaa cggatcgaga aaaatttcgt gattactgat ccgagactgc ctgacaaccc
     2701 aatcattttt gcgagcgatt ccttcctgca gctgacagaa tattctcggg aagagatcct
     2761 ggggcgcaat tgccgttttc tgcagggacc cgagacagac cgtgccactg ttcggaaaat
     2821 cagagatgct attgacaacc agactgaagt gaccgttcag ctgatcaatt ataccaagag
     2881 cggcaagaag ttctggaacg tgttccacct gcagccgatg cgcgattata agggcgacgt
     2941 ccagtacttc attggcgtgc agctggatgg caccgaacgt cttcatggcg ccgctgagcg
     3001 tgaggcggtc tgcctgatca aaaagacagc ctttcagatt gctgaggcag cgaacgacga
     3061 aaattacttt ggaagcggga gtgggagcaa actaattgct aaaagtgcag aagacgaaaa
     3121 agcgaaggaa gaaccaggga accataggat cgtaattctt gcaatgttag ctattggcgt
     3181 gttctcttta ggggcgttta tcaaaattat tcaattaaga aaaaataatg gaggaagtgg
     3241 cggaagtggc atggcagaaa tcggtactgg ctttccattc gacccccatt atgtggaagt
     3301 cctgggcgag cgcatgcact acgtcgatgt tggtccgcgc gatggcaccc ctgtgctgtt
     3361 cctgcacggt aacccgacct cctcctacgt gtggcgcaac atcatcccgc atgttgcacc
     3421 gacccatcgc tgcattgctc cagacctgat cggtatgggc aaatccgaca aaccagacct
     3481 gggttatttc ttcgacgacc acgtccgctt catggatgcc ttcatcgaag ccctgggtct
     3541 ggaagaggtc gtcctggtca ttcacgactg gggctccgct ctgggtttcc actgggccaa
     3601 gcgcaatcca gagcgcgtca aaggtattgc atttatggag ttcatccgcc ctatcccgac
     3661 ctgggacgaa tggccagaat ttgcccgcga gaccttccag gccttccgca ccaccgacgt
     3721 cggccgcaag ctgatcatcg atcagaacgt ttttatcgag ggtacgctgc cgatgggtgt
     3781 cgtccgcccg ctgactgaag tcgagatgga ccattaccgc gagccgttcc tgaatcctgt
     3841 tgaccgcgag ccactgtggc gcttcccaaa cgagctgcca atcgccggtg agccagcgaa
     3901 catcgtcgcg ctggtcgaag aatacatgga ctggctgcac cagtcccctg tcccgaagct
     3961 gctgttctgg ggcaccccag gcgttctgat cccaccggcc gaagccgctc gcctggccaa
     4021 aagcctgcct aactgcaagg ctgtggacat cggcccgggt ctgaatctgc tgcaagaaga
     4081 caacccggac ctgatcggca gcgagatcgc gcgctggctg tcgacgctcg agatttccgg
     4141 ctaaagcggc cgcgactcta gatcataatc agccatacca catttgtaga ggttttactt
     4201 gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa tgcaattgtt
     4261 gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     4321 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     4381 gtatcttaag gcgtaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg
     4441 ttaaatcagc tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa
     4501 agaatagacc gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa
     4561 gaacgtggac tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg
     4621 tgaaccatca ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa
     4681 ccctaaaggg agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa
     4741 ggaagggaag aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct
     4801 gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt caggtggcac
     4861 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     4921 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     4981 tcctgaggcg gaaagaacca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca
     5041 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt
     5101 ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca
     5161 gcaaccatag tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc
     5221 cattctccgc cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg
     5281 gcctctgagc tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa
     5341 gatcgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     5401 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     5461 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     5521 caagaccgac ctgtccggtg ccctgaatga actgcaagac gaggcagcgc ggctatcgtg
     5581 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     5641 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     5701 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     5761 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     5821 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     5881 actgttcgcc aggctcaagg cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg
     5941 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     6001 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     6061 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     6121 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     6181 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     6241 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     6301 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ctagggggag gctaactgaa
     6361 acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa aagacagaat
     6421 aaaacgcacg gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac
     6481 tctgtcgata ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc
     6541 ccaccccacc ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca
     6601 ggccctgcca tagcctcagg ttactcatat atactttaga ttgatttaaa acttcatttt
     6661 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     6721 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttc
//
