LOCUS       sequence-473402-        5740 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express NanoLuc(R)-hVTA1 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 5740)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5740)
  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            "Unpublished"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: NanoLuc-hVTA1 Vector
FEATURES             Location/Qualifiers
     source          1..5740
                     /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(1243..1755)
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     promoter        complement(1794..1812)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1795..1812)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        1932..2235
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2390..2410
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2443..2490
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2522..3316
                     /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(2576..2595)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3186..3205
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3474..4062)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3554..3573)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4188..4236
                     /note="synthetic polyadenylation signal"
     CDS             complement(4260..5297)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4273..4281
                     /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(5328..5685)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5342..5477
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5494..5685
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5642..5662
                     /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 ctagaggatc cccgggtacc gagcccgaat tcgtttaaac
      301 ttctctgcct gtcgtcagta acttgagagc cttttgtaga ttctggacag cagtgcttac
      361 atcttcatac tgcaaagcac tgccagcata tttgcagtac ttctgagctc tagcaaagtc
      421 ttctggggtt agacgaacat ccccctggga aattgtattg aaaagtgcgg gatcaatggc
      481 aggtatagtc tgtggagtag gttggatagt attacttgct acacctgtgc tgtgaggcac
      541 ttctgctggt gtattagctg gagcgtgtgc acccggagga atctgtattc cagtatagtt
      601 gcctgatggc atgttgcttg ggtcataagt tgaagatgat gatggctgag ttggctgagt
      661 gggcagagag gctgctccag catcttcatt ttcttcaata tcattatctt cttcaattcc
      721 aacagggcct gcttgaggag tctccccatt ctttaaacaa ttatggatgt atgttgcctt
      781 ccatctggca tacttcctgt gtttcacatt ttcatcagtg agttctccaa atactgttat
      841 gacatctatc aaaagacttg cagtatagaa ggacttgatc atgtttttgt gaaatcgtcc
      901 agcacgatct tcattgtctg catacaaaaa cattttcaaa gcataattct ccaaatgggc
      961 acagcccact atttcttgag taatagcttc attatcaccc aactgcttct ttagagcttc
     1021 taactgatcc attaactttg ataaaaattt gcgacattca ggagttttac tatcgatctt
     1081 cattccagtc tgcattgcgt ataaacgaca gtaataagcc accacagggt ctcgcttgtc
     1141 atgctcctga gccgtcctca gatgatgctg tatgctcttg aactgtgcgg ggagcggggg
     1201 cagcggtgca agcgcggcca tggcgatcgc gccgctcgag cccgccagaa tgcgttcgca
     1261 cagccgccag ccggtcactc cgttgatggt tactcggaac agcagggagc cgtcggggtt
     1321 gatcaggcgc tcgtcgataa ttttgttgcc gttccacagg gtccctgtta cagtgatctt
     1381 tttgccgtcg aacacggcga tgccttcata cggccgtccg aaatagtcga tcatgttcgg
     1441 cgtaaccccg tcgattacca gtgtgccata gtgcaggatc accttaaagt gatgatcatc
     1501 cacagggtac accaccttaa aaattttttc gatctggccc atttggtcgc cgctcagacc
     1561 ttcatacggg atgatgacat ggatgtcgat cttcagccca ttttcaccgc tcaggacaat
     1621 cctttggatc ggagttacgg acaccccgag attctgaaac aaactggaca cacctccctg
     1681 ttcaaggact tggtccaggt tgtagccggc tgtctgtcgc cagtccccaa cgaaatcttc
     1741 gagtgtgaag accatggtga gcgctagctg caaaaagaac aagtagcttg tattctatag
     1801 tgtcacctaa atgggcgatc tgacggttca ctaaaccagc tctgcttata taggcctccc
     1861 tggcgcgcca ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     1921 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     1981 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     2041 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     2101 tatcatatgc caagtccgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     2161 tatgcccagt acatgacctt acgggacttt cctacttggc agtacatcta cgtattagtc
     2221 atcgctatta ccatggtgat gcggttttgg cagtacacca atgggcgtgg atagcggttt
     2281 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     2341 caaaatcaac gggactttcc aaaatgtcgt aataaccccg ccccgttgac gcaaatgggc
     2401 ggtaggcgtg tacggtggga ggtcaaatcg aattctaata ctgttgacaa ttaatcatcg
     2461 gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccca ggaggcagat
     2521 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     2581 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     2641 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     2701 gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     2761 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     2821 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     2881 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     2941 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     3001 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc gcatgcccga
     3061 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     3121 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     3181 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     3241 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     3301 tgacgagttc ttctgattcg aaatgaccga ccaagcgacg cccaaccggt atcagctcac
     3361 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3421 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3481 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3541 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3601 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3661 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3721 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3781 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3841 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3901 ggctacacta gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3961 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4021 gtttgcaagc agcagattac gcgcagaaaa aaaggatttc aagaagatcc tttgatcttt
     4081 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4141 ttatcaaaaa ggatcttcac ctagatcctt ttatagtccg gaatcgtcac acaaaaaacc
     4201 aacacacaga tgtaatgaaa ataaagatat tttattgcgg ccatcgtgat ggctagagtc
     4261 taaactcgac ctacctcctt agcgcgcggt cgtgtactgg gccggcggtt gccgctgtcg
     4321 gccagcacct cgacgcagcc gtcggtccat acggctgcgc tccggcgagc gatttgtgtg
     4381 cgaccgacgg tgccggcccc gctgcggacg atggcatcgc agcggccttg agcccaggca
     4441 gcatcgtcga agttgccgtc cacgaggctc tggtagagct gatccaggcc gatgcgcagc
     4501 atgtaggctc gcagacgagg gctgccggcc agctcgggat gccggcgctc gaagtagcga
     4561 gtctgctgct ccatgcaagc cagccagggc cgccagaaga agatgttggc cacctcgtac
     4621 tgactgtccc cgaacatagc ttcggaccag tcgattacgg cggtgatgcg gccgttgtcg
     4681 gtcaggacgt tgttgctgcc gaagtcggca tggacgaggt ggcgcacctc gggacagtct
     4741 tcggcccaca gcatcagttc gtccagggct tgagctacgc tggcggacac ggtgtcgtcc
     4801 atcacggtct gccagtggta gacatgggga tcagcaatgg cgcaaatgaa atcccgccag
     4861 gttgtgtact ggccgatgcc ttggggcccg aatgggccga agccgctggt ttgactgagg
     4921 tcggcggcgg caatagcatc catagcttcg gcgacaggct gtaacacagc aggcagctct
     4981 gtttcgggga ggtcttggag agtaacgcct tgggcgcgtc tactgatgca gtatgtcagg
     5041 ctctcgctga actcgccgat gtccaacact tcggggatgg gtagtgcagc gctggcgaag
     5101 tggcggtaca cgtagcggtc tttgtagaag ccatcagcgc agctattcac ccgcagtaca
     5161 tagccgcgtc cgccgacatc gaagctgaag gctcggctct cttcgccctc cgacaactgc
     5221 atcaggtcgc tcacactgtc gaacttctcg atgagaaatt tttcaacgct ggtagcggtg
     5281 agttcgggct tcttcatggt ggcgctagtg tcagaagaat cgagcttttt gcaaaagcct
     5341 aggcctccaa aaaagcctcc tcactacttc tggaatagct cagaggccga ggcggcctcg
     5401 gcctctgcat aaataaaaaa aattagtcag ccatggggcg gagaatgggc ggaactgggc
     5461 ggagttaggg gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga
     5521 tgcatgcttt gcatacttct gcctgctggg gagcctgggg actttccaca cctggttgct
     5581 gactaattga gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca
     5641 caccctaact gacacacatt ccacagctgg ttctttccgc ctcagaaggt acctaaccaa
     5701 gttcctcttt cagaggttat ttcaggccat ggtgctgcgc
//
