LOCUS       sequence-473408-        6104 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express hVPS4A-NanoLuc(R) Fusion Protein in Mammalian Cells under a 
            CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6104)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6104)
  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: 
            "Unpublished"
COMMENT     Sequence Label: hVPS4A-NanoLuc Vector
FEATURES             Location/Qualifiers
     source          1..6104
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    17..98
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     42..61
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(275..787)
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     promoter        complement(2158..2176)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(2159..2176)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        2296..2599
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2754..2774
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2807..2854
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2886..3680
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2940..2959)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3550..3569
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3838..4426)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3918..3937)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4552..4600
                     /note="synthetic polyadenylation signal"
     CDS             complement(4624..5661)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4637..4645
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     promoter        complement(5692..6049)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5706..5841
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5858..6049
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     6006..6026
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
ORIGIN
        1 aagacgcgtg acgtcggcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
       61 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
      121 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
      181 aatgtggtat ggctgattat gatccgcggc cgcaatacgt cgacgttatc agctgacttc
      241 gtacgagagc ctaggtgttt ctagagtcgc ggccttacgc cagaatgcgt tcgcacagcc
      301 gccagccggt cactccgttg atggttactc ggaacagcag ggagccgtcg gggttgatca
      361 ggcgctcgtc gataattttg ttgccgttcc acagggtccc tgttacagtg atctttttgc
      421 cgtcgaacac ggcgatgcct tcatacggcc gtccgaaata gtcgatcatg ttcggcgtaa
      481 ccccgtcgat taccagtgtg ccatagtgca ggatcacctt aaagtgatga tcatccacag
      541 ggtacaccac cttaaaaatt ttttcgatct ggcccatttg gtcgccgctc agaccttcat
      601 acgggatgat gacatggatg tcgatcttca gcccattttc accgctcagg acaatccttt
      661 ggatcggagt tacggacacc ccgagattct gaaacaaact ggacacacct ccctgttcaa
      721 ggacttggtc caggttgtag ccggctgtct gtcgccagtc cccaacgaaa tcttcgagtg
      781 tgaagacgcc gctcgagccg agagaaacac tctcttgccc aaagtcctct gagaatttct
      841 tcactttcag gaggtcgtct gcattcaccg tgggccgggt ggtggccaga gaccgcagca
      901 tgtccgacat gcaaaccaca ggctctaaga gtttgtcccc agggacatcc atccaagtca
      961 tctccatggc tcctgggtcc cctggtgagc atggagtcag gaggtcatca atcatcatgc
     1021 tggggttggt gcgagagggg ccacagacct ttttgaagtg tgtggccgac tgcaccttcc
     1081 tcacgggctg catgagagag tcccgcacga tgatgctgat gtccgcgccc gagtagcctt
     1141 ccgtcttccg ggccagctcg tggatgtttg catccgtgag gttgtgggga gtgctcccga
     1201 gatgcaaccg gaacatctgg gcgcgggcag cttcctccgg caaggggata taaattcgtt
     1261 tttcaaacct cctcctgatg gccgaatcca acacccatgg gatgtttgtg gctccaagaa
     1321 ccagagtccc atcattgtta ttccccaccc cctgcatctg gaccaagaac tccgttttga
     1381 tcctccgggc ggcctcactc tcattttcat ttcgggaccc gcagagggaa tccacctcat
     1441 cgatgaagat gatggagggc ttgtgctgcc tggccagctc aaacaggttc ttgaccagct
     1501 tttcactctc ccccagccac ttggacatca gatctgagga ggacacagag aagaaggtgg
     1561 agttgttggc ctctgttgcc acggctttgg ccaggtagga tttccctgtg ccagggggtc
     1621 cgaacagcag aatcccccgc cagggggtgc gcttgcctgt gaacaagtgt gggaatttga
     1681 ttggcaaaat gacagcttct ttgagggcct ccttggcccc ctccagcccg gccacgtcgt
     1741 tccaccgtat gttgggcttc tccatcacga cggcacccat cagctgttct tgcagtttct
     1801 ttttctccgg attatcccct tcactgtcac tgtcactgcc cttgccctca ctctggttct
     1861 ctttgactgg cttcttgccg tgtttctctt tgcttcgtaa ataatccttc agcttctcgg
     1921 cccggtctag gtactgcacg cacttggctc gaatgctctc cttggccttg tcgctgtggg
     1981 cctcatactt gatagcgtgg aggaagtact ccaccgcatg ctggtacagc cgcagcgcct
     2041 cctcgtagtt cttggctttg tcctcctctg tggctttcgt caccagatca atggctttct
     2101 ggagggttga cgttgtcatg gcgatcgcta gctgcaaaaa gaacaagtag cttgtattct
     2161 atagtgtcac ctaaatgggc gatctgacgg ttcactaaac cagctctgct tatataggcc
     2221 tccctggcgc gccactagtt attaatagta atcaattacg gggtcattag ttcatagccc
     2281 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     2341 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     2401 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     2461 agtgtatcat atgccaagtc cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     2521 gcattatgcc cagtacatga ccttacggga ctttcctact tggcagtaca tctacgtatt
     2581 agtcatcgct attaccatgg tgatgcggtt ttggcagtac accaatgggc gtggatagcg
     2641 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     2701 gcaccaaaat caacgggact ttccaaaatg tcgtaataac cccgccccgt tgacgcaaat
     2761 gggcggtagg cgtgtacggt gggaggtcaa atcgaattct aatactgttg acaattaatc
     2821 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa cccaggaggc
     2881 agatcatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc
     2941 tattcggcta tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc
     3001 tgtcagcgca ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg
     3061 aactgcagga cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag
     3121 ctgtgctcga cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg
     3181 ggcaggatct cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg
     3241 caatgcggcg gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac
     3301 atcgcatcga gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg
     3361 acgaagagca tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc
     3421 ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg
     3481 aaaatggccg cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc
     3541 aggacatagc gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc
     3601 gcttcctcgt gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc
     3661 ttcttgacga gttcttctga ttcgaaatga ccgaccaagc gacgcccaac cggtatcagc
     3721 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3781 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3841 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3901 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3961 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     4021 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     4081 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     4141 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     4201 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     4261 ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     4321 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     4381 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tttcaagaag atcctttgat
     4441 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4501 gagattatca aaaaggatct tcacctagat ccttttatag tccggaatcg tcacacaaaa
     4561 aaccaacaca cagatgtaat gaaaataaag atattttatt gcggccatcg tgatggctag
     4621 agtctaaact cgacctacct ccttagcgcg cggtcgtgta ctgggccggc ggttgccgct
     4681 gtcggccagc acctcgacgc agccgtcggt ccatacggct gcgctccggc gagcgatttg
     4741 tgtgcgaccg acggtgccgg ccccgctgcg gacgatggca tcgcagcggc cttgagccca
     4801 ggcagcatcg tcgaagttgc cgtccacgag gctctggtag agctgatcca ggccgatgcg
     4861 cagcatgtag gctcgcagac gagggctgcc ggccagctcg ggatgccggc gctcgaagta
     4921 gcgagtctgc tgctccatgc aagccagcca gggccgccag aagaagatgt tggccacctc
     4981 gtactgactg tccccgaaca tagcttcgga ccagtcgatt acggcggtga tgcggccgtt
     5041 gtcggtcagg acgttgttgc tgccgaagtc ggcatggacg aggtggcgca cctcgggaca
     5101 gtcttcggcc cacagcatca gttcgtccag ggcttgagct acgctggcgg acacggtgtc
     5161 gtccatcacg gtctgccagt ggtagacatg gggatcagca atggcgcaaa tgaaatcccg
     5221 ccaggttgtg tactggccga tgccttgggg cccgaatggg ccgaagccgc tggtttgact
     5281 gaggtcggcg gcggcaatag catccatagc ttcggcgaca ggctgtaaca cagcaggcag
     5341 ctctgtttcg gggaggtctt ggagagtaac gccttgggcg cgtctactga tgcagtatgt
     5401 caggctctcg ctgaactcgc cgatgtccaa cacttcgggg atgggtagtg cagcgctggc
     5461 gaagtggcgg tacacgtagc ggtctttgta gaagccatca gcgcagctat tcacccgcag
     5521 tacatagccg cgtccgccga catcgaagct gaaggctcgg ctctcttcgc cctccgacaa
     5581 ctgcatcagg tcgctcacac tgtcgaactt ctcgatgaga aatttttcaa cgctggtagc
     5641 ggtgagttcg ggcttcttca tggtggcgct agtgtcagaa gaatcgagct ttttgcaaaa
     5701 gcctaggcct ccaaaaaagc ctcctcacta cttctggaat agctcagagg ccgaggcggc
     5761 ctcggcctct gcataaataa aaaaaattag tcagccatgg ggcggagaat gggcggaact
     5821 gggcggagtt aggggcggga tgggcggagt taggggcggg actatggttg ctgactaatt
     5881 gagatgcatg ctttgcatac ttctgcctgc tggggagcct ggggactttc cacacctggt
     5941 tgctgactaa ttgagatgca tgctttgcat acttctgcct gctggggagc ctggggactt
     6001 tccacaccct aactgacaca cattccacag ctggttcttt ccgcctcaga aggtacctaa
     6061 ccaagttcct ctttcagagg ttatttcagg ccatggtgct gcgc
//
