LOCUS       sequence-383968-        5315 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses myrlysin in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5315)
  AUTHORS   Pu J, Schindler C, Jia R, Jarnik M, Backlund P, Bonifacino JS
  TITLE     BORC, a multisubunit complex that regulates lysosome positioning.
  JOURNAL   Dev Cell. 2015 Apr 20;33(2):176-88. doi: 
            10.1016/j.devcel.2015.02.011.
  PUBMED    25898167
REFERENCE   2  (bases 1 to 5315)
  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"; doi: 
            "10.1016/j.devcel.2015.02"; journalName: "Dev Cell"; date: 
            "2015-04-20- 20"; volume: "33"; issue: "2"; pages: "176-88"
COMMENT     Sequence Label: GFP-myrlysin/BORCS5
FEATURES             Location/Qualifiers
     source          1..5315
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        98..401
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        402..605
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     556..576
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      1292..1301
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1298..1300,1301..1303,1304..2017)
                     /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(1343..1364)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1604..1623)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1951..1972
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    2140..2261
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2177..2196)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2231..2250
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2268..2723)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2405..2426)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2617..2636
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2750..2854
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2856..3213
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2856..3047
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2879..2899)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3064..3199
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3248..4042
                     /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             3248..4042
                     /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(3302..3321)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3912..3931
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4230..4249)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4274..4321
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4650..5238
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5139..5158
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccctgattct gtggataacc gtattaccgc catgcattag ttattaatag taatcaatta
       61 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      121 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      181 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      241 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      301 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      361 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      421 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      481 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      541 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      601 gagctggttt agtgaaccgt cagatccgct agcgctaccg gactcagatc tcgagctcaa
      661 gcttcgaatt cgcatgggca gtgagcagag ctccgaggcc gagagccgac ccaacgatct
      721 gaactcctca gtgactcctt caccagccaa gcatagagcc aagatggatg atattgtggt
      781 tgtagctcag ggctcccagg cctcacggaa cgtcagcaac gatcccgatg tcatcaagtt
      841 gcaagagatt ccaaccttcc agcccctttt gaaagggcta ttgagtggcc agacttcccc
      901 aacaaatgcc aaattggaga aactggactc tcagcaggtg ttgcagctct gcctccgata
      961 tcaagatcac ctgcatcagt gtgcagaggc cgttgctttt gaccagaatg ctttggttaa
     1021 acgaatcaaa gagatggatc tgtctgtaga aactctgttc agcttcatgc aggagcgcca
     1081 gaaaagatac gccaagtatg ccgagcagat ccagaaagtg aacgagatgt ccgccatcct
     1141 ccgccgcata cagatgggca tcgaccagac tgtgcccctg ctggacaggc tcaacagcat
     1201 gctgcccgag ggcgagcggc tggagccctt cagcatgaag cccgaccgcg agctcaggct
     1261 gaaggtaccg cgggcccggg atccaccggt cgccaccatg gtgagcaagg gcgaggagct
     1321 gttcaccggg gtggtgccca tcctggtcga gctggacggc gacgtaaacg gccacaagtt
     1381 cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc aagctgaccc tgaagttcat
     1441 ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc gtgaccaccc tgacctacgg
     1501 cgtgcagtgc ttcagccgct accccgacca catgaagcag cacgacttct tcaagtccgc
     1561 catgcccgaa ggctacgtcc aggagcgcac catcttcttc aaggacgacg gcaactacaa
     1621 gacccgcgcc gaggtgaagt tcgagggcga caccctggtg aaccgcatcg agctgaaggg
     1681 catcgacttc aaggaggacg gcaacatcct ggggcacaag ctggagtaca actacaacag
     1741 ccacaacgtc tatatcatgg ccgacaagca gaagaacggc atcaaggtga acttcaagat
     1801 ccgccacaac atcgaggacg gcagcgtgca gctcgccgac cactaccagc agaacacccc
     1861 catcggcgac ggccccgtgc tgctgcccga caaccactac ctgagcaccc agtccaagct
     1921 gagcaaagac cccaacgaga agcgcgatca catggtcctg ctggagttcg tgaccgccgc
     1981 cgggatcact ctcggcatgg acgagctgta caagtaaagc ggccgcgact ctagatcata
     2041 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     2101 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     2161 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     2221 cattctagtt gtggtttgtc caaactcatc aatgtatctt aaggcgtaaa ttgtaagcgt
     2281 taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata
     2341 ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt
     2401 tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg
     2461 aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt
     2521 ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc
     2581 ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg
     2641 cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct
     2701 taatgcgccg ctacagggcg cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     2761 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     2821 ataaatgctt caataatatt gaaaaaggaa gagtcctgag gcggaaagaa ccagctgtgg
     2881 aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa
     2941 agcatgcatc tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc
     3001 agaagtatgc aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg
     3061 cccatcccgc ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt
     3121 ttttttattt atgcagaggc cgaggccgcc tcggcctctg agctattcca gaagtagtga
     3181 ggaggctttt ttggaggcct aggcttttgc aaagatcgat caagagacag gatgaggatc
     3241 gtttcgcatg attgaacaag atggattgca cgcaggttct ccggccgctt gggtggagag
     3301 gctattcggc tatgactggg cacaacagac aatcggctgc tctgatgccg ccgtgttccg
     3361 gctgtcagcg caggggcgcc cggttctttt tgtcaagacc gacctgtccg gtgccctgaa
     3421 tgaactgcaa gacgaggcag cgcggctatc gtggctggcc acgacgggcg ttccttgcgc
     3481 agctgtgctc gacgttgtca ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc
     3541 ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag aaagtatcca tcatggctga
     3601 tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc ccattcgacc accaagcgaa
     3661 acatcgcatc gagcgagcac gtactcggat ggaagccggt cttgtcgatc aggatgatct
     3721 ggacgaagag catcaggggc tcgcgccagc cgaactgttc gccaggctca aggcgagcat
     3781 gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc tgcttgccga atatcatggt
     3841 ggaaaatggc cgcttttctg gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta
     3901 tcaggacata gcgttggcta cccgtgatat tgctgaagag cttggcggcg aatgggctga
     3961 ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg
     4021 ccttcttgac gagttcttct gagcgggact ctggggttcg aaatgaccga ccaagcgacg
     4081 cccaacctgc catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc
     4141 ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag
     4201 ttcttcgccc accctagggg gaggctaact gaaacacgga aggagacaat accggaagga
     4261 acccgcgcta tgacggcaat aaaaagacag aataaaacgc acggtgttgg gtcgtttgtt
     4321 cataaacgcg gggttcggtc ccagggctgg cactctgtcg ataccccacc gagaccccat
     4381 tggggccaat acgcccgcgt ttcttccttt tccccacccc accccccaag ttcgggtgaa
     4441 ggcccagggc tcgcagccaa cgtcggggcg gcaggccctg ccatagcctc aggttactca
     4501 tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc
     4561 ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca
     4621 gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc
     4681 tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta
     4741 ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt
     4801 ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc
     4861 gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg
     4921 ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg
     4981 tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag
     5041 ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc
     5101 agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat
     5161 agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg
     5221 gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc
     5281 tggccttttg ctcacatgtt ctttcctgcg ttatc
//
