LOCUS       sequence-439253-        7369 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Synchronize the trafficking of LIMP2 from the ER..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7369)
  AUTHORS   Ecard J, Lian YL, Divoux S, Gouveia Z, Vigne E, Perez F, Boncompain 
            G
  TITLE     Lysosomal membrane proteins LAMP1 and LIMP2 are segregated in the 
            Golgi apparatus independently of their clathrin adaptor binding 
            motif.
  JOURNAL   Mol Biol Cell. 2024 Mar 1;35(3):ar42. doi: 10.1091/mbc.E23-06-0251. 
            Epub 2024 Jan 17.
  PUBMED    38231876
REFERENCE   2  (bases 1 to 7369)
  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: "Mol Biol 
            Cell."; date: "2024-03-1"; volume: "35(3):ar42. doi"; pages: " 
            10.1091/mbc.E23-06-0251. Epub 2024 Jan 17"
COMMENT     Sequence Label: Mini-Ii-Str_SBP-EGFP-LIMP2
FEATURES             Location/Qualifiers
     source          1..7369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     intron          1656..1885
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     1868..1887
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    1943..2515
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2109..2126)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2336..2355
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2586..2699
                     /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="MDEKTTGWRGGHVVEGLAGELEQLRARLEHHPQGQREP"
     CDS             join(2709..2711,2712..3422)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2709..2711,2712..3422)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2751..2772)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3012..3031)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3359..3380
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5042..5163
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5079..5098)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5133..5152
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5212..5228)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5212..5228)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5225..5247)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5236..5252
                     /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(5260..5266,5267..5284,5285..5290))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5305..5326
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5443..5460)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5614..6202)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5694..5713)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6373..7164,7165..7233))
                     /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     6996..7015
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7234..7338)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gcttaaggcc tgttaaccgg tcgtacgtct ccggattcga attaattcgg atccgtgatg
      961 cacaggagga gagccagggc ctgtcgggaa gatcaaaagc cagtcatcga tgatcagcgc
     1021 gaccttatct ccaacaatga gcaactgccc atgctgggcc ggcgccctgg ggccccggag
     1081 agcaagtgca gccgcctcga ggaccctagc aaagactcaa aagctcaggt gtccgctgcc
     1141 gaggctggca ttactggaac atggtacaat cagctcggga gcacctttat tgtgactgct
     1201 ggagccgatg gagccctcac cggaacatac gaatctgctg tgggaaacgc cgaatcacgg
     1261 tacgtcctca ctggccgata cgatagtgcc cctgccaccg acggatctgg gactgccctg
     1321 ggatggactg tcgcttggaa aaacaactac cggaatgctc attctgccac aacatggagt
     1381 ggacagtacg tgggaggcgc tgaggctaga atcaatacac agtggctgct cacatctggc
     1441 acaaccgagg caaatgcttg gaaatccacc ctggtgggac atgacacatt caccaaagtg
     1501 aaaccctccg ccgcttcaat cgatgccgcc aaaaaagccg gagtcaacaa cggcaatcct
     1561 ctggatgccg tccagcagta aggatccgcg gccgcataga taactgatcc agtgtgctgg
     1621 aattaattcg ctgtctgcga gggccagctg ttggggtgag tactccctct caaaagcggg
     1681 catgacttct gcgctaagat tgtcagtttc caaaaacgag gaggatttga tattcacctg
     1741 gcccgcggtg atgcctttga gggtggccgc gtccatctgg tcagaaaaga caatcttttt
     1801 gttgtcaagc ttgaggtgtg gcaggcttga gatctggcca tacacttgag tgacaatgac
     1861 atccactttg cctttctctc cacaggtgtc cactcccagg tccaactgca ggtcgagcat
     1921 gcatctaggg cggccaattc cgcccctctc cccccccccc cctaacgtta ctggccgaag
     1981 ccgcttggaa taaggccggt gtgcgtttgt ctatatgtta ttttccacca tattgccgtc
     2041 ttttggcaat gtgagggccc ggaaacctgg ccctgtcttc ttgacgagca ttcctagggg
     2101 tctttcccct ctcgccaaag gaatgcaagg tctgttgaat gtcgtgaagg aagcagttcc
     2161 tctggaagct tcttgaagac aaacaacgtc tgtagcgacc ctttgcaggc agcggaaccc
     2221 cccacctggc gacaggtgcc tctgcggcca aaagccacgt gtataagata cacctgcaaa
     2281 ggcggcacaa ccccagtgcc acgttgtgag ttggatagtt gtggaaagag tcaaatggct
     2341 ctcctcaagc gtattcaaca aggggctgaa ggatgcccag aaggtacccc attgtatggg
     2401 atctgatctg gggcctcggt gcacatgctt tacatgtgtt tagtcgaggt taaaaaaacg
     2461 tctaggcccc ccgaaccacg gggacgtggt tttcctttga aaaacacgat gataagcttg
     2521 ccacaaccca caaggagacg accttccatg accgagtaca agcggcgcgc caaattgcga
     2581 attccatgga cgagaagacc actggttggc gaggtggaca cgttgttgaa ggactggctg
     2641 gggaacttga acaacttcgt gcacgactgg agcatcaccc acaaggtcaa cgtgaaccac
     2701 ctgcaggtgt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     2761 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     2821 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     2881 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     2941 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     3001 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     3061 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     3121 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     3181 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     3241 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     3301 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     3361 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca
     3421 agggccggcc aggccgatgc tgcttctaca cggcggggac gttgtccctg ctcctgctgg
     3481 tgaccagcgt cacgctgctg gtggcccggg tcttccagaa ggctgtagac cagagtatcg
     3541 agaagaaaat tgtgttaagg aatggtactg aggcatttga ctcctgggag aagccccctc
     3601 tgcctgtgta tactcagttc tatttcttca atgtcaccaa tccagaggag atcctcagag
     3661 gggagacccc tcgggtggaa gaagtggggc catacaccta cagggaactc agaaacaaag
     3721 caaatattca atttggagat aatggaacaa caatatctgc tgttagcaac aaggcctatg
     3781 tttttgaacg agaccaatct gttggagacc ctaaaattga cttaattaga acattaaata
     3841 ttcctgtatt gactgtcata gagtggtccc aggtgcactt cctcagggag atcatcgagg
     3901 ccatgttgaa agcctatcag cagaagctct ttgtgactca cacagttgac gaattgctct
     3961 ggggctacaa agatgaaatc ttgtccctta tccatgtttt caggcccgat atctctccct
     4021 attttggcct attctatgag aaaaatggga ctaatgatgg agactatgtt tttctaactg
     4081 gagaagacag ttaccttaac tttacaaaaa ttgtggaatg gaatgggaaa acgtcacttg
     4141 actggtggat aacagacaag tgcaatatga ttaatggaac agatggagat tcttttcacc
     4201 cactaataac caaagatgag gtcctttatg tcttcccatc tgacttttgc aggtcagtgt
     4261 atattacttt cagtgactat gagagtgtac agggactgcc tgcctttcgg tataaagttc
     4321 ctgcagaaat attagccaat acgtcagaca atgccggctt ctgtatacct gagggaaact
     4381 gcctgggctc aggagttctg aatgtcagca tctgcaagaa tggtgcaccc atcattatgt
     4441 ctttcccaca cttttaccaa gcagatgaga ggtttgtttc tgccatagaa ggcatgcacc
     4501 caaatcagga agaccatgag acatttgtgg acattaatcc tttgactgga ataatcctaa
     4561 aagcagccaa gaggttccaa atcaacattt atgtcaaaaa attagatgac tttgttgaaa
     4621 cgggagacat tagaaccatg gttttcccag tgatgtacct caatgagagt gttcacattg
     4681 ataaagagac ggcgagtcga ctgaagtcta tgattaacac tactttgatc atcaccaaca
     4741 taccctacat catcatggcg ctgggtgtgt tctttggttt ggtttttacc tggcttgcat
     4801 gcaaaggaca gggatccatg gatgagggaa cagcggatga aagagcaccc ctcattcgaa
     4861 cctaaattaa ttaagcggcc gcttcgaaat gaccgaccaa gcgacgccca acctgccatc
     4921 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     4981 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     5041 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
     5101 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
     5161 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct
     5221 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat
     5281 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc
     5341 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     5401 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
     5461 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     5521 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     5581 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     5641 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     5701 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     5761 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     5821 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     5881 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     5941 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     6001 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     6061 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     6121 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     6181 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     6241 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     6301 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     6361 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     6421 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     6481 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     6541 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     6601 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     6661 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     6721 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     6781 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     6841 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     6901 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     6961 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     7021 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     7081 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     7141 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     7201 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     7261 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     7321 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtc
//
