LOCUS       sequence-473407-        6130 bp    DNA     circular SYN 14-JUL-2025
DEFINITION  Express NanoLuc(R)-hVPS4A 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 6130)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 6130)
  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: NanoLuc-hVPS4A Vector
FEATURES             Location/Qualifiers
     source          1..6130
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        56..413
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        56..247
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(79..99)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      264..399
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             444..1481
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             1460..1468
                     /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)"
     polyA_signal    1505..1553
                     /note="synthetic polyadenylation signal"
     rep_origin      1679..2267
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2168..2187
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2425..3219)
                     /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(2536..2555)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     3146..3165
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(3251..3298)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     primer_bind     complement(3331..3351)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3506..3809
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3929..3947
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3929..3946
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             3986..4498
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     polyA_signal    6033..6114
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6070..6089)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
ORIGIN
        1 gcgcagcacc atggcctgaa ataacctctg aaagaggaac ttggttaggt accttctgag
       61 gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
      121 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
      181 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
      241 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
      301 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tcggcctctg
      361 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcg
      421 attcttctga cactagcgcc accatgaaga agcccgaact caccgctacc agcgttgaaa
      481 aatttctcat cgagaagttc gacagtgtga gcgacctgat gcagttgtcg gagggcgaag
      541 agagccgagc cttcagcttc gatgtcggcg gacgcggcta tgtactgcgg gtgaatagct
      601 gcgctgatgg cttctacaaa gaccgctacg tgtaccgcca cttcgccagc gctgcactac
      661 ccatccccga agtgttggac atcggcgagt tcagcgagag cctgacatac tgcatcagta
      721 gacgcgccca aggcgttact ctccaagacc tccccgaaac agagctgcct gctgtgttac
      781 agcctgtcgc cgaagctatg gatgctattg ccgccgccga cctcagtcaa accagcggct
      841 tcggcccatt cgggccccaa ggcatcggcc agtacacaac ctggcgggat ttcatttgcg
      901 ccattgctga tccccatgtc taccactggc agaccgtgat ggacgacacc gtgtccgcca
      961 gcgtagctca agccctggac gaactgatgc tgtgggccga agactgtccc gaggtgcgcc
     1021 acctcgtcca tgccgacttc ggcagcaaca acgtcctgac cgacaacggc cgcatcaccg
     1081 ccgtaatcga ctggtccgaa gctatgttcg gggacagtca gtacgaggtg gccaacatct
     1141 tcttctggcg gccctggctg gcttgcatgg agcagcagac tcgctacttc gagcgccggc
     1201 atcccgagct ggccggcagc cctcgtctgc gagcctacat gctgcgcatc ggcctggatc
     1261 agctctacca gagcctcgtg gacggcaact tcgacgatgc tgcctgggct caaggccgct
     1321 gcgatgccat cgtccgcagc ggggccggca ccgtcggtcg cacacaaatc gctcgccgga
     1381 gcgcagccgt atggaccgac ggctgcgtcg aggtgctggc cgacagcggc aaccgccggc
     1441 ccagtacacg accgcgcgct aaggaggtag gtcgagttta gactctagcc atcacgatgg
     1501 ccgcaataaa atatctttat tttcattaca tctgtgtgtt ggttttttgt gtgacgattc
     1561 cggactataa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     1621 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     1681 gaaatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     1741 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     1801 gcagagcgca gataccaaat actgtccttc tagtgtagcc gtagttaggc caccacttca
     1861 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     1921 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     1981 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     2041 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     2101 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     2161 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     2221 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     2281 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
     2341 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accggttggg
     2401 cgtcgcttgg tcggtcattt cgaatcagaa gaactcgtca agaaggcgat agaaggcgat
     2461 gcgctgcgaa tcgggagcgg cgataccgta aagcacgagg aagcggtcag cccattcgcc
     2521 gccaagctct tcagcaatat cacgggtagc caacgctatg tcctgatagc ggtccgccac
     2581 acccagccgg ccacagtcga tgaatccaga aaagcggcca ttttccacca tgatattcgg
     2641 caagcaggca tcgccatggg tcacgacgag atcctcgccg tcgggcatgc gcgccttgag
     2701 cctggcgaac agttcggctg gcgcgagccc ctgatgctct tcgtccagat catcctgatc
     2761 gacaagaccg gcttccatcc gagtacgtgc tcgctcgatg cgatgtttcg cttggtggtc
     2821 gaatgggcag gtagccggat caagcgtatg cagccgccgc attgcatcag ccatgatgga
     2881 tactttctcg gcaggagcaa ggtgagatga caggagatcc tgccccggca cttcgcccaa
     2941 tagcagccag tcccttcccg cttcagtgac aacgtcgagc acagctgcgc aaggaacgcc
     3001 cgtcgtggcc agccacgata gccgcgctgc ctcgtcctgc agttcattca gggcaccgga
     3061 caggtcggtc ttgacaaaaa gaaccgggcg cccctgcgct gacagccgga acacggcggc
     3121 atcagagcag ccgattgtct gttgtgccca gtcatagccg aatagcctct ccacccaagc
     3181 ggccggagaa cctgcgtgca atccatcttg ttcaatcatg atctgcctcc tgggtttagt
     3241 tcctcacctt gtcgtattat actatgccga tatactatgc cgatgattaa ttgtcaacag
     3301 tattagaatt cgatttgacc tcccaccgta cacgcctacc gcccatttgc gtcaacgggg
     3361 cggggttatt acgacatttt ggaaagtccc gttgattttg gtgccaaaac aaactcccat
     3421 tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat ccacgcccat
     3481 tggtgtactg ccaaaaccgc atcaccatgg taatagcgat gactaatacg tagatgtact
     3541 gccaagtagg aaagtcccgt aaggtcatgt actgggcata atgccaggcg ggccatttac
     3601 cgtcattgac gtcaataggg ggcggacttg gcatatgata cacttgatgt actgccaagt
     3661 gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct attggcgtta
     3721 ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc ggtcagccag
     3781 gcgggccatt taccgtaagt tatgtaacgc ggaactccat atatgggcta tgaactaatg
     3841 accccgtaat tgattactat taataactag tggcgcgcca gggaggccta tataagcaga
     3901 gctggtttag tgaaccgtca gatcgcccat ttaggtgaca ctatagaata caagctactt
     3961 gttctttttg cagctagcgc tcaccatggt cttcacactc gaagatttcg ttggggactg
     4021 gcgacagaca gccggctaca acctggacca agtccttgaa cagggaggtg tgtccagttt
     4081 gtttcagaat ctcggggtgt ccgtaactcc gatccaaagg attgtcctga gcggtgaaaa
     4141 tgggctgaag atcgacatcc atgtcatcat cccgtatgaa ggtctgagcg gcgaccaaat
     4201 gggccagatc gaaaaaattt ttaaggtggt gtaccctgtg gatgatcatc actttaaggt
     4261 gatcctgcac tatggcacac tggtaatcga cggggttacg ccgaacatga tcgactattt
     4321 cggacggccg tatgaaggca tcgccgtgtt cgacggcaaa aagatcactg taacagggac
     4381 cctgtggaac ggcaacaaaa ttatcgacga gcgcctgatc aaccccgacg gctccctgct
     4441 gttccgagta accatcaacg gagtgaccgg ctggcggctg tgcgaacgca ttctggcggg
     4501 ctcgagcggc gcgatcgcca tgacaacgtc aaccctccag aaagccattg atctggtgac
     4561 gaaagccaca gaggaggaca aagccaagaa ctacgaggag gcgctgcggc tgtaccagca
     4621 tgcggtggag tacttcctcc acgctatcaa gtatgaggcc cacagcgaca aggccaagga
     4681 gagcattcga gccaagtgcg tgcagtacct agaccgggcc gagaagctga aggattattt
     4741 acgaagcaaa gagaaacacg gcaagaagcc agtcaaagag aaccagagtg agggcaaggg
     4801 cagtgacagt gacagtgaag gggataatcc ggagaaaaag aaactgcaag aacagctgat
     4861 gggtgccgtc gtgatggaga agcccaacat acggtggaac gacgtggccg ggctggaggg
     4921 ggccaaggag gccctcaaag aagctgtcat tttgccaatc aaattcccac acttgttcac
     4981 aggcaagcgc accccctggc gggggattct gctgttcgga ccccctggca cagggaaatc
     5041 ctacctggcc aaagccgtgg caacagaggc caacaactcc accttcttct ctgtgtcctc
     5101 ctcagatctg atgtccaagt ggctggggga gagtgaaaag ctggtcaaga acctgtttga
     5161 gctggccagg cagcacaagc cctccatcat cttcatcgat gaggtggatt ccctctgcgg
     5221 gtcccgaaat gaaaatgaga gtgaggccgc ccggaggatc aaaacggagt tcttggtcca
     5281 gatgcagggg gtggggaata acaatgatgg gactctggtt cttggagcca caaacatccc
     5341 atgggtgttg gattcggcca tcaggaggag gtttgaaaaa cgaatttata tccccttgcc
     5401 ggaggaagct gcccgcgccc agatgttccg gttgcatctc gggagcactc cccacaacct
     5461 cacggatgca aacatccacg agctggcccg gaagacggaa ggctactcgg gcgcggacat
     5521 cagcatcatc gtgcgggact ctctcatgca gcccgtgagg aaggtgcagt cggccacaca
     5581 cttcaaaaag gtctgtggcc cctctcgcac caaccccagc atgatgattg atgacctcct
     5641 gactccatgc tcaccagggg acccaggagc catggagatg acttggatgg atgtccctgg
     5701 ggacaaactc ttagagcctg tggtttgcat gtcggacatg ctgcggtctc tggccaccac
     5761 ccggcccacg gtgaatgcag acgacctcct gaaagtgaag aaattctcag aggactttgg
     5821 gcaagagagt gtttaaacga attcgggctc ggtacccggg gatcctctag aaacacctag
     5881 gctctcgtac gaagtcagct gataacgtcg acgtattgcg gccgcggatc ataatcagcc
     5941 ataccacatt tgtagaggtt ttacttgctt taaaaaacct cccacacctc cccctgaacc
     6001 tgaaacataa aatgaatgca attgttgttg ttaacttgtt tattgcagct tataatggtt
     6061 acaaataaag caatagcatc acaaatttca caaataaagc atttttttca ctgccgacgt
     6121 cacgcgtctt
//
