LOCUS       sequence-247518-        6075 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a chimera of LAMP1, SBP tag and GFP (fluorescent tag).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6075)
  AUTHORS   Guardia CM, De Pace R, Sen A, Saric A, Jarnik M, Kolin DA, Kunwar A,
            Bonifacino JS
  TITLE     Reversible association with motor proteins (RAMP): A 
            streptavidin-based method to manipulate organelle positioning.
  JOURNAL   PLoS Biol. 2019 May 17;17(5):e3000279. doi: 
            10.1371/journal.pbio.3000279. eCollection 2019 May.
  PUBMED    31100061
REFERENCE   2  (bases 1 to 6075)
  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: "PLoS 
            Biol."; date: "2019-05-17"; volume: "17(5):e3000279. doi"; pages: " 
            10.1371/journal.pbio.3000279. eCollection 2019 May"
COMMENT     Sequence Label: LAMP1-SBP-GFP
FEATURES             Location/Qualifiers
     source          1..6075
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      620..629
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      624..633
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(626..688,689..1846)
                     /codon_start=1
                     /product="rat lysosome-associated membrane glycoprotein 1"
                     /label=LAMP1
                     /translation="MAAPGARRPLLLLLLAGLAHSAPALFEVKDNNGTACIMASFSASF
                     LTTYEAGHVSKVSNMTLPASAEVLKNSSSCGEKNASEPTLAITFGEGYLLKLTFTKNTT
                     RYSVQHMYFTYNLSDTQFFPNASSKGPDTVDSTTDIKADINKTYRCVSDIRVYMKNVTI
                     VLWDATIQAYLPSSNFSKEETRCPQDQPSPTTGPPSPSPPLVPTNPSVSKYNVTGDNGT
                     CLLASMALQLNITYMKKDNTTVTRAFNINPSDKYSGTCGAQLVTLKVGNKSRVLELQFG
                     MNATSSLFFLQGVQLNMTLPDAIEPTFSTSNYSLKALQASVGNSYKCNSEEHIFVSKAL
                     ALNVFSVQVQAFRVESDRFGSVEECVQDGNNMLIPIAVGGALAGLVLIVLIAYLIGRKR
                     SHAGYQTI"
     CDS             1895..2005
                     /codon_start=1
                     /product="streptavidin-binding peptide"
                     /label=SBP
                     /note="selected from a peptide library; binds streptavidin 
                     with nanomolar affinity (Keefe et al., 2001)"
                     /translation="DEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     regulatory      2015..2024
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2021..2023,2024..2026,2027..2740)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2066..2087)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2327..2346)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2674..2695
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2863..2984
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2900..2919)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2954..2973
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2991..3446)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3128..3149)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3340..3359
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3473..3577
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3579..3936
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3579..3770
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3602..3622)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3787..3922
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3971..4765
                     /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             3971..4765
                     /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(4025..4044)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4635..4654
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4953..4972)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4997..5044
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5373..5961
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5862..5881
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggactcag atctcgagcg ccaccatggc ggccccgggc gcccggcggc cgctgctcct
      661 gttgctgctg gcaggccttg cacacagcgc cccagcactg ttcgaggtga aagacaacaa
      721 cggcacagcg tgtataatgg ccagcttctc tgcctccttt ctgaccacct atgaggctgg
      781 acatgtttct aaggtctcga atatgaccct gccagcctct gcagaagtcc tgaagaatag
      841 cagctcttgt ggtgaaaaga atgcttctga gcccaccctc gcaatcacct ttggagaagg
      901 atatttactg aaactcacct tcacaaaaaa cacaacacgt tacagtgtcc agcacatgta
      961 tttcacatat aacctgtcag acacacaatt ctttcccaat gccagctcca aagggcccga
     1021 cactgtggat tccacaactg acatcaaggc agacatcaac aaaacatacc gatgtgtcag
     1081 cgacatcagg gtctacatga agaatgtgac cattgtgctc tgggacgcta ctatccaggc
     1141 ctacctgccg agtagcaact tcagcaagga agagacacgc tgcccacagg atcaaccttc
     1201 cccaactact gggccaccca gcccctcacc accacttgtg cccacaaacc ccagtgtgtc
     1261 caagtacaat gtgactggtg acaatggaac ctgcctgctg gcctctatgg cactgcaact
     1321 caacatcacc tacatgaaga aggacaacac gactgtgacc agagcattca acatcaaccc
     1381 aagtgacaaa tatagtggga cttgcggtgc ccagttggtg accctgaagg tggggaacaa
     1441 gagcagagtc ctggagctgc agtttgggat gaatgccact tctagcctgt ttttcctgca
     1501 aggagttcag ttgaacatga ctcttcctga tgccatagag cccacgttca gcacctccaa
     1561 ctattccctg aaagctcttc aggccagtgt cggcaactca tacaagtgca acagtgagga
     1621 gcacatcttt gtcagcaagg cgctcgccct caatgtcttc agcgtgcaag tccaggcttt
     1681 cagggtagaa agtgacaggt ttgggtctgt ggaagagtgt gtacaggacg gtaacaacat
     1741 gctgatcccc attgctgtgg gcggggccct ggcagggctg gtcctcatcg tcctcatcgc
     1801 ctacctcatc ggcaggaaga ggagtcacgc gggctatcag accatctcgg aattcggctc
     1861 caccggctcc accggctcca ccggcgcgga tccggacgag aagaccactg gttggcgagg
     1921 tggacacgtt gttgaaggac tggctgggga acttgaacaa cttcgtgcac gactggagca
     1981 tcacccacaa ggtcaacgtg aaccacgacc ggtcgccacc atggtgagca agggcgagga
     2041 gctgttcacc ggggtggtgc ccatcctggt cgagctggac ggcgacgtaa acggccacaa
     2101 gttcagcgtg tccggcgagg gcgagggcga tgccacctac ggcaagctga ccctgaagtt
     2161 catctgcacc accggcaagc tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta
     2221 cggcgtgcag tgcttcagcc gctaccccga ccacatgaag cagcacgact tcttcaagtc
     2281 cgccatgccc gaaggctacg tccaggagcg caccatcttc ttcaaggacg acggcaacta
     2341 caagacccgc gccgaggtga agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa
     2401 gggcatcgac ttcaaggagg acggcaacat cctggggcac aagctggagt acaactacaa
     2461 cagccacaac gtctatatca tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa
     2521 gatccgccac aacatcgagg acggcagcgt gcagctcgcc gaccactacc agcagaacac
     2581 ccccatcggc gacggccccg tgctgctgcc cgacaaccac tacctgagca cccagtccaa
     2641 gctgagcaaa gaccccaacg agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc
     2701 cgccgggatc actctcggca tggacgagct gtacaagtaa agcggccgcg actctagatc
     2761 ataatcagcc ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc
     2821 cccctgaacc tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct
     2881 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     2941 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttaaggcgt aaattgtaag
     3001 cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca
     3061 ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag
     3121 tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg
     3181 gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt
     3241 tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag
     3301 agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc
     3361 gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc
     3421 gcttaatgcg ccgctacagg gcgcgtcagg tggcactttt cggggaaatg tgcgcggaac
     3481 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc
     3541 ctgataaatg cttcaataat attgaaaaag gaagagtcct gaggcggaaa gaaccagctg
     3601 tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     3661 caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca
     3721 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     3781 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     3841 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     3901 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     3961 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     4021 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     4081 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     4141 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     4201 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     4261 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     4321 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     4381 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     4441 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     4501 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     4561 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     4621 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     4681 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     4741 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     4801 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     4861 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     4921 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     4981 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     5041 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     5101 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     5161 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     5221 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     5281 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     5341 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     5401 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     5461 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     5521 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     5581 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     5641 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     5701 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     5761 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     5821 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     5881 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     5941 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     6001 tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt
     6061 attaccgcca tgcat
//
