LOCUS       sequence-439255-        7244 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 7244)
  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 7244)
  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: Str-Endo-KKXX_SBP-EGFP-LIMP2
FEATURES             Location/Qualifiers
     source          1..7244
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(48..67)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        239..618
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..822
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     773..793
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     867..886
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        867..885
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1417..1443
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1417..1443
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     intron          1545..1774
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     primer_bind     1757..1776
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     misc_feature    join(1847..2421,2422..2424,2425..2433)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(2014..2031)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     2241..2260
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             2461..2574
                     /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(2584..2586,2587..3297)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2584..2586,2587..3297)
                     /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(2626..2647)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2887..2906)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3234..3255
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    4868..4989
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4905..4924)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(5091..5107)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5091..5107)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5104..5126)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5115..5131
                     /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(5139..5145,5146..5163,5164..5169))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5184..5205
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5322..5339)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5493..6081)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5573..5592)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6252..7043,7044..7112))
                     /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     6875..6894
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7113..7217)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct
       61 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
      121 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
      181 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
      241 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
      301 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
      361 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
      421 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
      481 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
      541 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta
      601 gtcatcgcta ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg
      661 tttgactcac ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg
      721 caccaaaatc aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg
      781 ggcggtaggc gtgtacggtg ggaggtctat ataagcagag ctctctggct aactagagaa
      841 cccactgctt actggcttat cgaaattaat acgactcact atagggagac ccaagcttgg
      901 taccgagctc ggatcgatat ctgaaccatg gaccccagca aggacagcaa ggcccaggtg
      961 agcgccgccg aggccggcat caccggcacc tggtacaacc agctgggcag caccttcatc
     1021 gtgaccgccg gcgccgacgg cgccctgacc ggcacctacg agagcgccgt gggcaacgcc
     1081 gagagcagat acgtgctgac cggcagatac gacagcgccc ccgccaccga cggcagcggc
     1141 accgccctgg gctggaccgt ggcctggaag aacaactaca gaaacgccca cagcgccacc
     1201 acctggagcg gccagtacgt gggcggcgcc gaggccagaa tcaacaccca gtggctgctg
     1261 accagcggca ccaccgaggc caacgcctgg aagagcaccc tggtgggcca cgacaccttc
     1321 accaaggtga agcccagcgc cgccagcatc gacgccgcca agaaggccgg cgtgaacaac
     1381 ggcaaccccc tggacgccgt gcagcagggc ggatcctacc cctacgacgt gcccgactac
     1441 gccgccggct accagaccat caagaagacc aactgagcgg ccgcatagat aactgatcca
     1501 gtgtgctgga attaattcgc tgtctgcgag ggccagctgt tggggtgagt actccctctc
     1561 aaaagcgggc atgacttctg cgctaagatt gtcagtttcc aaaaacgagg aggatttgat
     1621 attcacctgg cccgcggtga tgcctttgag ggtggccgcg tccatctggt cagaaaagac
     1681 aatctttttg ttgtcaagct tgaggtgtgg caggcttgag atctggccat acacttgagt
     1741 gacaatgaca tccactttgc ctttctctcc acaggtgtcc actcccaggt ccaactgcag
     1801 gtcgagcatg catctagggc ggccaattcc gcccctctcc cccccccccc tctccctccc
     1861 ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata
     1921 tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg
     1981 tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg caaggtctgt
     2041 tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca acgtctgtag
     2101 cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc ggccaaaagc
     2161 cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga
     2221 tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg ctgaaggatg
     2281 cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca tgctttacat
     2341 gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc
     2401 tttgaaaaac acgatgataa gcttgccaca acccgggagg cgcgccaaat tgcgaattcc
     2461 atggacgaga agaccactgg ttggcgaggt ggacacgttg ttgaaggact ggctggggaa
     2521 cttgaacaac ttcgtgcacg actggagcat cacccacaag gtcaacgtga accacctgca
     2581 ggtgtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     2641 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     2701 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     2761 ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc gctaccccga ccacatgaag
     2821 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     2881 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     2941 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     3001 aagctggagt acaactacaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     3061 ggcatcaagg tgaacttcaa gatccgccac aacatcgagg acggcagcgt gcagctcgcc
     3121 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     3181 tacctgagca cccagtccgc cctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     3241 ctgctggagt tcgtgaccgc cgccgggatc actctcggca tggacgagct gtacaagggc
     3301 cggccaggcc gatgctgctt ctacacggcg gggacgttgt ccctgctcct gctggtgacc
     3361 agcgtcacgc tgctggtggc ccgggtcttc cagaaggctg tagaccagag tatcgagaag
     3421 aaaattgtgt taaggaatgg tactgaggca tttgactcct gggagaagcc ccctctgcct
     3481 gtgtatactc agttctattt cttcaatgtc accaatccag aggagatcct cagaggggag
     3541 acccctcggg tggaagaagt ggggccatac acctacaggg aactcagaaa caaagcaaat
     3601 attcaatttg gagataatgg aacaacaata tctgctgtta gcaacaaggc ctatgttttt
     3661 gaacgagacc aatctgttgg agaccctaaa attgacttaa ttagaacatt aaatattcct
     3721 gtattgactg tcatagagtg gtcccaggtg cacttcctca gggagatcat cgaggccatg
     3781 ttgaaagcct atcagcagaa gctctttgtg actcacacag ttgacgaatt gctctggggc
     3841 tacaaagatg aaatcttgtc ccttatccat gttttcaggc ccgatatctc tccctatttt
     3901 ggcctattct atgagaaaaa tgggactaat gatggagact atgtttttct aactggagaa
     3961 gacagttacc ttaactttac aaaaattgtg gaatggaatg ggaaaacgtc acttgactgg
     4021 tggataacag acaagtgcaa tatgattaat ggaacagatg gagattcttt tcacccacta
     4081 ataaccaaag atgaggtcct ttatgtcttc ccatctgact tttgcaggtc agtgtatatt
     4141 actttcagtg actatgagag tgtacaggga ctgcctgcct ttcggtataa agttcctgca
     4201 gaaatattag ccaatacgtc agacaatgcc ggcttctgta tacctgaggg aaactgcctg
     4261 ggctcaggag ttctgaatgt cagcatctgc aagaatggtg cacccatcat tatgtctttc
     4321 ccacactttt accaagcaga tgagaggttt gtttctgcca tagaaggcat gcacccaaat
     4381 caggaagacc atgagacatt tgtggacatt aatcctttga ctggaataat cctaaaagca
     4441 gccaagaggt tccaaatcaa catttatgtc aaaaaattag atgactttgt tgaaacggga
     4501 gacattagaa ccatggtttt cccagtgatg tacctcaatg agagtgttca cattgataaa
     4561 gagacggcga gtcgactgaa gtctatgatt aacactactt tgatcatcac caacataccc
     4621 tacatcatca tggcgctggg tgtgttcttt ggtttggttt ttacctggct tgcatgcaaa
     4681 ggacagggat ccatggatga gggaacagcg gatgaaagag cacccctcat tcgaacctaa
     4741 attaattaac tctagataac tgatcataat cagccatacc acatttgtag aggttttact
     4801 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     4861 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     4921 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4981 tgtatcttaa cgcgtcgagt gcattctagt tgtggtttgt ccaaactcat caatgtatct
     5041 tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg
     5101 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     5161 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     5221 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     5281 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     5341 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     5401 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     5461 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     5521 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     5581 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     5641 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     5701 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     5761 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     5821 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     5881 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     5941 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     6001 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     6061 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     6121 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     6181 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     6241 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     6301 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     6361 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     6421 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     6481 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     6541 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     6601 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     6661 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     6721 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     6781 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     6841 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     6901 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     6961 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     7021 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     7081 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     7141 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     7201 caaatagggg ttccgcgcac atttccccga aaagtgccac ctga
//
