LOCUS       sequence-479073-        6334 bp    DNA     circular SYN 21-JUL-2025
DEFINITION  Express NanoLuc-DGKe 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 6334)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6334)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 21, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: NanoLuc-DGKe Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6334
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        138..517
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        518..721
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     672..692
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          857..989
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     primer_bind     972..991
                     /label=pMT2-F
                     /note="Synthetic intron, forward primer"
     primer_bind     1033..1052
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1033..1051
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1065..1577
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     primer_bind     complement(3413..3431)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3468..3589
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3479..3498)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3533..3552
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3834..4191
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3834..4025
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3857..3877)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4042..4177
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4206..4253
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4285..5079
                     /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(4339..4358)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4949..4968
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5143..5191
                     /note="synthetic polyadenylation signal"
     rep_origin      5348..5893
                     /direction=RIGHT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     5794..5813
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6010..6027
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tcaatattgg ccattagcca tattattcat tggttatata gcataaatca atattggcta
       61 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      121 aatatgaccg ccatgttggc attgattatt gactagttat taatagtaat caattacggg
      181 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      241 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      301 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      361 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      421 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      481 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      541 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      601 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      661 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      721 tggtttagtg aaccgtcaga tcactagaag ctttattgcg gtagtttatc acagttaaat
      781 tgctaacgca gtcagtgctt ctgacacaac agtctcgaac ttaagctgca gaagttggtc
      841 gtgaggcact gggcaggtaa gtatcaaggt tacaagacag gtttaaggag accaatagaa
      901 actgggcttg tcgagacaga gaagactctt gcgtttctga taggcaccta ttggtcttac
      961 tgacatccac tttgcctttc tctccacagg tgtccactcc cagttcaatt acagctctta
     1021 aggctagagt attaatacga ctcactatag ggctagcgct caccatggtc ttcacactcg
     1081 aagatttcgt tggggactgg cgacagacag ccggctacaa cctggaccaa gtccttgaac
     1141 agggaggtgt gtccagtttg tttcagaatc tcggggtgtc cgtaactccg atccaaagga
     1201 ttgtcctgag cggtgaaaat gggctgaaga tcgacatcca tgtcatcatc ccgtatgaag
     1261 gtctgagcgg cgaccaaatg ggccagatcg aaaaaatttt taaggtggtg taccctgtgg
     1321 atgatcatca ctttaaggtg atcctgcact atggcacact ggtaatcgac ggggttacgc
     1381 cgaacatgat cgactatttc ggacggccgt atgaaggcat cgccgtgttc gacggcaaaa
     1441 agatcactgt aacagggacc ctgtggaacg gcaacaaaat tatcgacgag cgcctgatca
     1501 accccgacgg ctccctgctg ttccgagtaa ccatcaacgg agtgaccggc tggcggctgt
     1561 gcgaacgcat tctggcgggc tcgagcggcg cgatcgccat ggaagcggag aggcggccgg
     1621 cgccgggctc gccctccgag ggcctgtttg cggacgggca cctgatcttg tggacgctgt
     1681 gctcggtcct gctgccggtg ttcatcacct tctggtgtag cctccagcgg tcgcgccggc
     1741 agctgcaccg cagggacatc ttccgcaaga gcaagcacgg gtggcgcgac acggacctgt
     1801 tcagccagcc cacctactgc tgcgtgtgcg cgcagcacat tctgcagggc gccttctgcg
     1861 actgctgcgg gctccgcgtg gacgagggct gcctcaggaa ggccgacaag cgcttccagt
     1921 gcaaggagat tatgctcaag aatgacacca aggtcctgga cgccatgccc caccactgga
     1981 tccggggcaa cgtgcccctg tgcagttact gtatggtttg caagcagcag tgtggctgtc
     2041 aacccaagct ttgcgattac aggtgcattt ggtgccagaa aacagtacat gatgagtgca
     2101 tgaaaaatag tttaaagaat gaaaaatgtg attttggaga attcaaaaac ctaatcattc
     2161 caccaagtta tttaacctcc attaatcaga tgcgtaaaga caaaaaaaca gattatgaag
     2221 tgctagcctc taagcttgga aagcagtgga ccccattaat aatcctggcc aactctcgta
     2281 gtggaactaa tatgggagaa ggactgttgg gagaatttag gatcttgttg aatccagtcc
     2341 aggtttttga tgtaactaaa actcctccta tcaaagccct acaactctgt actcttctcc
     2401 catattattc agctcgagta cttgtttgtg gaggggatgg gactgtaggg tgggtcctgg
     2461 atgcagttga tgacatgaag attaagggac aagaaaagta cattccacaa gttgcagttt
     2521 tgcctctggg aacaggcaac gatctatcca atacattggg ttggggtaca ggttatgctg
     2581 gagaaattcc agttgcgcag gttttgcgaa atgtaatgga agcagatgga attaaactag
     2641 atcgatggaa agttcaagta acaaataaag gatactacaa cttaagaaaa cccaaggaat
     2701 tcacaatgaa caactatttt tctgttggac ctgatgctct catggctctc aattttcatg
     2761 ctcatcgtga gaaggcacca tctctgtttt ctagcagaat tcttaataag gcggtttact
     2821 tattctatgg aaccaaagat tgtttagtgc aagaatgtaa agatttgaat aaaaaagttg
     2881 agctagaact ggatggtgag cgagtagcac tgcccagctt ggaaggtatt atagttctga
     2941 acatcggata ctggggcggt ggctgcagac tatgggaagg gatgggggac gagacttacc
     3001 ctctagccag gcatgacgat ggtctgctgg aagtcgttgg agtatatggg tctttccact
     3061 gtgctcagat tcaagtaaaa ctggctaatc cttttcgaat aggacaggca catacagtga
     3121 ggctgatttt gaagtgctcc atgatgccaa tgcaggtgga tggggagcct tgggcccaag
     3181 ggccctgcac tgtcaccata actcacaaga cacatgcaat gatgttatat ttctctggag
     3241 aacaaacaga tgatgacatc tctagtactt cggatcaaga agatataaag gcgactgaag
     3301 tttaaacgaa ttcgggctcg gtacccgggg atcctctaga gtcgacctgc aggcatgcaa
     3361 gctgatccgg ctgctaacaa agcccgaaag gaagctgagt tggctgctgc caccgctgag
     3421 caataactag cataacccct tggggcggcc gcttcgagca gacatgataa gatacattga
     3481 tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg
     3541 tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa
     3601 ttgcattcat tttatgtttc aggttcaggg ggagatgtgg gaggtttttt taagcaagta
     3661 aaacctctac aaatgtggta aaatcgaatt ttaacaaaat attaacgctt acaatttcct
     3721 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gcggatctgc
     3781 gcagcaccat ggcctgaaat aacctctgaa agaggaactt ggttaggtac cttctgaggc
     3841 ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca
     3901 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg tggaaagtcc
     3961 ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccata
     4021 gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc ccattctccg
     4081 ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc ggcctctgag
     4141 ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa aaagcttaat
     4201 taactgttga caattaatca tcggcatagt atatcggcat agtataatac gacaaggtga
     4261 ggaactaaac ccaggaggca gatcatgatt gaacaagatg gattgcacgc aggttctccg
     4321 gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat cggctgctct
     4381 gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac
     4441 ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg gctggccacg
     4501 acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg
     4561 ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa
     4621 gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca
     4681 ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt
     4741 gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc
     4801 aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc
     4861 ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg
     4921 ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt
     4981 ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag
     5041 cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg gggttcgaaa
     5101 tgaccgacca agcgacgccc aacctgccat cacgatggcc gcaataaaat atctttattt
     5161 tcattacatc tgtgtgttgg ttttttgtgt gaatcgatag cgataaggat ccgctcatgc
     5221 gatgataagc tgtcaaacat gagaattaca acttatatcg tatggggctg acttcaggtg
     5281 ctacatttga agagataaat tgcactgaaa tctagaaata ttttatctga ttaataagat
     5341 gatcttcttg agatcgtttt ggtctgcgcg taatctcttg ctctgaaaac gaaaaaaccg
     5401 ccttgcaggg cggtttttcg aaggttctct gagctaccaa ctctttgaac cgaggtaact
     5461 ggcttggagg agcgcagtca ccaaaacttg tcctttcagt ttagccttaa ccggcgcatg
     5521 acttcaagac taactcctct aaatcaatta ccagtggctg ctgccagtgg tgcttttgca
     5581 tgtctttccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcggactga
     5641 acggggggtt cgtgcataca gtccagcttg gagcgaactg cctacccgga actgagtgtc
     5701 aggcgtggaa tgagacaaac gcggccataa cagcggaatg acaccggtaa accgaaaggc
     5761 aggaacagga gagcgcacga gggagccgcc agggggaaac gcctggtatc tttatagtcc
     5821 tgtcgggttt cgccaccact gatttgagcg tcagatttcg tgatgcttgt caggggggcg
     5881 gagcctatgg aaaaacggct ttgccgcggc cctctcactt ccctgttaag tatcttcctg
     5941 gcatcttcca ggaaatctcc gccccgttcg taagccattt ccgctcgccg cagtcgaacg
     6001 accgagcgta gcgagtcagt gagcgaggaa gcggaatata tcctgtatca catattctgc
     6061 tgacgcaccg gtgcagcctt ttttctcctg ccacatgaag cacttcactg acaccctcat
     6121 cagtgccaac atagtaagcc agtatacact ccgctagcgc tgatgtccgg cggtgctttt
     6181 gccgttacgc accaccccgt catgagatta cgcgttttag cggtgcgtac aattaaggga
     6241 ttatggtaaa tccacttact gtctgccctc gtagccatcg agataaaccg cagtactccg
     6301 gccacgatgc gtccggcgta gaggatcgag atct
//
