LOCUS       sequence-477953-        4227 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of EGFP-V5 under control of copper-inducible promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4227)
  AUTHORS   Bosch JA, Beltran PMJ, Cavers C, LaGraff JT, Melanson R, Singh A, 
            Chen W, Hu Y, Tattikota SG, Liu Y, Hashmi Y, Asara JM, Branon T, 
            Ting AY, Carr SA, Perrimon N
  TITLE     Multi-omic mapping of Drosophila protein secretomes reveals 
            tissue-specific origins and inter-organ trafficking
  JOURNAL   bioRxiv 2025.07.09.659702; doi: 
            https://doi.org/10.1101/2025.07.09.659702
REFERENCE   2  (bases 1 to 4227)
  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: "bioRxiv 
            2025.07.09.659702; doi: https://doi.org/10.1101/2025.07.09.659702"
COMMENT     Sequence Label: pMT-EGFP-V5
FEATURES             Location/Qualifiers
     source          1..4227
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        401..827
                     /label=MT promoter
                     /note="Drosophila metallothionein promoter"
     CDS             join(877..879,880..882,883..1593)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(877..879,880..882,883..1593)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(922..943)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1183..1202)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1530..1551
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1618..1659
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             1669..1686
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1709..1726)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1728..1862
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1752..1771)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1806..1825
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(2006..2022)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2006..2022)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2019..2041)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2030..2046
                     /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(2054..2060,2061..2078,2079..2084))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2099..2120
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2237..2254)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2408..2996)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2488..2507)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3167..3958,3959..4027))
                     /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     3790..3809
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4028..4132)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4200..4218
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt aattcgttgc aggacaggat
      421 gtggtgcccg atgtgactag ctctttgctg caggccgtcc tatcctctgg ttccgataag
      481 agacccagaa ctccggcccc ccaccgccca ccgccacccc catacatatg tggtacgcaa
      541 gtaagagtgc ctgcgcatgc cccatgtgcc ccaccaagag ctttgcatcc catacaagtc
      601 cccaaagtgg agaaccgaac caattcttcg cgggcagaac aaaagcttct gcacacgtct
      661 ccactcgaat ttggagccgg ccggcgtgtg caaaagaggt gaatcgaacg aaagacccgt
      721 gtgtaaagcc gcgtttccaa aatgtataaa accgagagca tctggccaat gtgcatcagt
      781 tgtggtcagc agcaaaatca agtgaatcat ctcagtgcaa ctaaaggggg gatctagatc
      841 ggggtaccta ctagtccagt gtggtggaat tccaaaatgg tgagcaaggg cgaggagctg
      901 ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg ccacaagttc
      961 agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct gaagttcatc
     1021 tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct gacctacggc
     1081 gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt caagtccgcc
     1141 atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg caactacaag
     1201 acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga gctgaagggc
     1261 atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa ctacaacagc
     1321 cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa cttcaagatc
     1381 cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca gaacaccccc
     1441 atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca gtccgccctg
     1501 agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc
     1561 gggatcactc tcggcatgga cgagctgtac aagtctctag agggcccgcg gttcgaaggt
     1621 aagcctatcc ctaaccctct cctcggtctc gattctacgc gtaccggtca tcatcaccat
     1681 caccattgag tttaaacccg ctgatcagcc tcgactgtgc cttctaagat ccagacatga
     1741 taagatacat tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta
     1801 tttgtgaaat ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag
     1861 ttaacaacaa caattgcatt cattttatgt ttcaggttca gggggaggtg tgggaggttt
     1921 tttaaagcaa gtaaaacctc tacaaatgtg gtatggctga ttatgatcag tcgacctgca
     1981 ggcatgcaag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
     2041 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     2101 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     2161 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     2221 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     2281 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     2341 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     2401 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     2461 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     2521 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     2581 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     2641 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     2701 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     2761 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     2821 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     2881 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     2941 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     3001 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     3061 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     3121 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     3181 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     3241 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     3301 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     3361 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     3421 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     3481 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     3541 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     3601 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     3661 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     3721 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     3781 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     3841 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     3901 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     3961 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     4021 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     4081 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     4141 cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
     4201 ataggcgtat cacgaggccc tttcgtc
//
