LOCUS       sequence-471873-        5714 bp    DNA     circular SYN 14-JUL-2025
DEFINITION  Express hVTA1-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 5714)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 5714)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: hVTA1-NanoLuc Vector
FEATURES             Location/Qualifiers
     source          1..5714
                     /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(1768..1786)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1769..1786)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     enhancer        1906..2209
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2364..2384
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        2417..2464
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2496..3290
                     /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(2550..2569)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3160..3179
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     rep_origin      complement(3448..4036)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3528..3547)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4162..4210
                     /note="synthetic polyadenylation signal"
     CDS             complement(4234..5271)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             4247..4255
                     /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(5302..5659)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5316..5451
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     enhancer        5468..5659
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     5616..5636
                     /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 agagaaactt ctctgcctgt cgtcagtaac ttgagagcct
      841 tttgtagatt ctggacagca gtgcttacat cttcatactg caaagcactg ccagcatatt
      901 tgcagtactt ctgagctcta gcaaagtctt ctggggttag acgaacatcc ccctgggaaa
      961 ttgtattgaa aagtgcggga tcaatggcag gtatagtctg tggagtaggt tggatagtat
     1021 tacttgctac acctgtgctg tgaggcactt ctgctggtgt attagctgga gcgtgtgcac
     1081 ccggaggaat ctgtattcca gtatagttgc ctgatggcat gttgcttggg tcataagttg
     1141 aagatgatga tggctgagtt ggctgagtgg gcagagaggc tgctccagca tcttcatttt
     1201 cttcaatatc attatcttct tcaattccaa cagggcctgc ttgaggagtc tccccattct
     1261 ttaaacaatt atggatgtat gttgccttcc atctggcata cttcctgtgt ttcacatttt
     1321 catcagtgag ttctccaaat actgttatga catctatcaa aagacttgca gtatagaagg
     1381 acttgatcat gtttttgtga aatcgtccag cacgatcttc attgtctgca tacaaaaaca
     1441 ttttcaaagc ataattctcc aaatgggcac agcccactat ttcttgagta atagcttcat
     1501 tatcacccaa ctgcttcttt agagcttcta actgatccat taactttgat aaaaatttgc
     1561 gacattcagg agttttacta tcgatcttca ttccagtctg cattgcgtat aaacgacagt
     1621 aataagccac cacagggtct cgcttgtcat gctcctgagc cgtcctcaga tgatgctgta
     1681 tgctcttgaa ctgtgcgggg agcgggggca gcggtgcaag cgcggccatg gcgatcgcta
     1741 gctgcaaaaa gaacaagtag cttgtattct atagtgtcac ctaaatgggc gatctgacgg
     1801 ttcactaaac cagctctgct tatataggcc tccctggcgc gccactagtt attaatagta
     1861 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
     1921 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
     1981 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
     2041 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagtc cgccccctat
     2101 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttacggga
     2161 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt
     2221 ttggcagtac accaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca
     2281 ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg
     2341 tcgtaataac cccgccccgt tgacgcaaat gggcggtagg cgtgtacggt gggaggtcaa
     2401 atcgaattct aatactgttg acaattaatc atcggcatag tatatcggca tagtataata
     2461 cgacaaggtg aggaactaaa cccaggaggc agatcatgat tgaacaagat ggattgcacg
     2521 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     2581 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     2641 tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg cggctatcgt
     2701 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     2761 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     2821 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     2881 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     2941 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     3001 aactgttcgc caggctcaag gcgcgcatgc ccgacggcga ggatctcgtc gtgacccatg
     3061 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     3121 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     3181 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     3241 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga ttcgaaatga
     3301 ccgaccaagc gacgcccaac cggtatcagc tcactcaaag gcggtaatac ggttatccac
     3361 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     3421 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     3481 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     3541 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     3601 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     3661 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     3721 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     3781 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     3841 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagga cagtatttgg
     3901 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     3961 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     4021 aaaaaaagga tttcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     4081 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     4141 ccttttatag tccggaatcg tcacacaaaa aaccaacaca cagatgtaat gaaaataaag
     4201 atattttatt gcggccatcg tgatggctag agtctaaact cgacctacct ccttagcgcg
     4261 cggtcgtgta ctgggccggc ggttgccgct gtcggccagc acctcgacgc agccgtcggt
     4321 ccatacggct gcgctccggc gagcgatttg tgtgcgaccg acggtgccgg ccccgctgcg
     4381 gacgatggca tcgcagcggc cttgagccca ggcagcatcg tcgaagttgc cgtccacgag
     4441 gctctggtag agctgatcca ggccgatgcg cagcatgtag gctcgcagac gagggctgcc
     4501 ggccagctcg ggatgccggc gctcgaagta gcgagtctgc tgctccatgc aagccagcca
     4561 gggccgccag aagaagatgt tggccacctc gtactgactg tccccgaaca tagcttcgga
     4621 ccagtcgatt acggcggtga tgcggccgtt gtcggtcagg acgttgttgc tgccgaagtc
     4681 ggcatggacg aggtggcgca cctcgggaca gtcttcggcc cacagcatca gttcgtccag
     4741 ggcttgagct acgctggcgg acacggtgtc gtccatcacg gtctgccagt ggtagacatg
     4801 gggatcagca atggcgcaaa tgaaatcccg ccaggttgtg tactggccga tgccttgggg
     4861 cccgaatggg ccgaagccgc tggtttgact gaggtcggcg gcggcaatag catccatagc
     4921 ttcggcgaca ggctgtaaca cagcaggcag ctctgtttcg gggaggtctt ggagagtaac
     4981 gccttgggcg cgtctactga tgcagtatgt caggctctcg ctgaactcgc cgatgtccaa
     5041 cacttcgggg atgggtagtg cagcgctggc gaagtggcgg tacacgtagc ggtctttgta
     5101 gaagccatca gcgcagctat tcacccgcag tacatagccg cgtccgccga catcgaagct
     5161 gaaggctcgg ctctcttcgc cctccgacaa ctgcatcagg tcgctcacac tgtcgaactt
     5221 ctcgatgaga aatttttcaa cgctggtagc ggtgagttcg ggcttcttca tggtggcgct
     5281 agtgtcagaa gaatcgagct ttttgcaaaa gcctaggcct ccaaaaaagc ctcctcacta
     5341 cttctggaat agctcagagg ccgaggcggc ctcggcctct gcataaataa aaaaaattag
     5401 tcagccatgg ggcggagaat gggcggaact gggcggagtt aggggcggga tgggcggagt
     5461 taggggcggg actatggttg ctgactaatt gagatgcatg ctttgcatac ttctgcctgc
     5521 tggggagcct ggggactttc cacacctggt tgctgactaa ttgagatgca tgctttgcat
     5581 acttctgcct gctggggagc ctggggactt tccacaccct aactgacaca cattccacag
     5641 ctggttcttt ccgcctcaga aggtacctaa ccaagttcct ctttcagagg ttatttcagg
     5701 ccatggtgct gcgc
//
