LOCUS       sequence-479794-        6308 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express DGKe-NanoLuc 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 6308)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6308)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 7, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: DGKe-NanoLuc Fusion Vector
FEATURES             Location/Qualifiers
     source          1..6308
                     /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             2787..3299
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(3387..3405)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    3442..3563
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3453..3472)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3507..3526
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3808..4165
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3808..3999
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3831..3851)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      4016..4151
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        4180..4227
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4259..5053
                     /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(4313..4332)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4923..4942
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5117..5165
                     /note="synthetic polyadenylation signal"
     rep_origin      5322..5867
                     /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     5768..5787
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5984..6001
                     /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 ggctagcgat cgccatggaa gcggagaggc
     1081 ggccggcgcc gggctcgccc tccgagggcc tgtttgcgga cgggcacctg atcttgtgga
     1141 cgctgtgctc ggtcctgctg ccggtgttca tcaccttctg gtgtagcctc cagcggtcgc
     1201 gccggcagct gcaccgcagg gacatcttcc gcaagagcaa gcacgggtgg cgcgacacgg
     1261 acctgttcag ccagcccacc tactgctgcg tgtgcgcgca gcacattctg cagggcgcct
     1321 tctgcgactg ctgcgggctc cgcgtggacg agggctgcct caggaaggcc gacaagcgct
     1381 tccagtgcaa ggagattatg ctcaagaatg acaccaaggt cctggacgcc atgccccacc
     1441 actggatccg gggcaacgtg cccctgtgca gttactgtat ggtttgcaag cagcagtgtg
     1501 gctgtcaacc caagctttgc gattacaggt gcatttggtg ccagaaaaca gtacatgatg
     1561 agtgcatgaa aaatagttta aagaatgaaa aatgtgattt tggagaattc aaaaacctaa
     1621 tcattccacc aagttattta acctccatta atcagatgcg taaagacaaa aaaacagatt
     1681 atgaagtgct agcctctaag cttggaaagc agtggacccc attaataatc ctggccaact
     1741 ctcgtagtgg aactaatatg ggagaaggac tgttgggaga atttaggatc ttgttgaatc
     1801 cagtccaggt ttttgatgta actaaaactc ctcctatcaa agccctacaa ctctgtactc
     1861 ttctcccata ttattcagct cgagtacttg tttgtggagg ggatgggact gtagggtggg
     1921 tcctggatgc agttgatgac atgaagatta agggacaaga aaagtacatt ccacaagttg
     1981 cagttttgcc tctgggaaca ggcaacgatc tatccaatac attgggttgg ggtacaggtt
     2041 atgctggaga aattccagtt gcgcaggttt tgcgaaatgt aatggaagca gatggaatta
     2101 aactagatcg atggaaagtt caagtaacaa ataaaggata ctacaactta agaaaaccca
     2161 aggaattcac aatgaacaac tatttttctg ttggacctga tgctctcatg gctctcaatt
     2221 ttcatgctca tcgtgagaag gcaccatctc tgttttctag cagaattctt aataaggcgg
     2281 tttacttatt ctatggaacc aaagattgtt tagtgcaaga atgtaaagat ttgaataaaa
     2341 aagttgagct agaactggat ggtgagcgag tagcactgcc cagcttggaa ggtattatag
     2401 ttctgaacat cggatactgg ggcggtggct gcagactatg ggaagggatg ggggacgaga
     2461 cttaccctct agccaggcat gacgatggtc tgctggaagt cgttggagta tatgggtctt
     2521 tccactgtgc tcagattcaa gtaaaactgg ctaatccttt tcgaatagga caggcacata
     2581 cagtgaggct gattttgaag tgctccatga tgccaatgca ggtggatggg gagccttggg
     2641 cccaagggcc ctgcactgtc accataactc acaagacaca tgcaatgatg ttatatttct
     2701 ctggagaaca aacagatgat gacatctcta gtacttcgga tcaagaagat ataaaggcga
     2761 ctgaagtttc tctcggctcg agcggcgtct tcacactcga agatttcgtt ggggactggc
     2821 gacagacagc cggctacaac ctggaccaag tccttgaaca gggaggtgtg tccagtttgt
     2881 ttcagaatct cggggtgtcc gtaactccga tccaaaggat tgtcctgagc ggtgaaaatg
     2941 ggctgaagat cgacatccat gtcatcatcc cgtatgaagg tctgagcggc gaccaaatgg
     3001 gccagatcga aaaaattttt aaggtggtgt accctgtgga tgatcatcac tttaaggtga
     3061 tcctgcacta tggcacactg gtaatcgacg gggttacgcc gaacatgatc gactatttcg
     3121 gacggccgta tgaaggcatc gccgtgttcg acggcaaaaa gatcactgta acagggaccc
     3181 tgtggaacgg caacaaaatt atcgacgagc gcctgatcaa ccccgacggc tccctgctgt
     3241 tccgagtaac catcaacgga gtgaccggct ggcggctgtg cgaacgcatt ctggcgtaag
     3301 gccgcgactc tagagtcgac ctgcaggcat gcaagctgat ccggctgcta acaaagcccg
     3361 aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac cccttggggc
     3421 ggccgcttcg agcagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     3481 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     3541 ttataagctg caataaacaa gttaacaaca acaattgcat tcattttatg tttcaggttc
     3601 agggggagat gtgggaggtt tttttaagca agtaaaacct ctacaaatgt ggtaaaatcg
     3661 aattttaaca aaatattaac gcttacaatt tcctgatgcg gtattttctc cttacgcatc
     3721 tgtgcggtat ttcacaccgc atacgcggat ctgcgcagca ccatggcctg aaataacctc
     3781 tgaaagagga acttggttag gtaccttctg aggcggaaag aaccagctgt ggaatgtgtg
     3841 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3901 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
     3961 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
     4021 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
     4081 ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt
     4141 ttttggaggc ctaggctttt gcaaaaagct taattaactg ttgacaatta atcatcggca
     4201 tagtatatcg gcatagtata atacgacaag gtgaggaact aaacccagga ggcagatcat
     4261 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     4321 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     4381 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     4441 ggacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     4501 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     4561 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     4621 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     4681 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     4741 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgcgca tgcccgacgg
     4801 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     4861 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     4921 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     4981 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     5041 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     5101 ccatcacgat ggccgcaata aaatatcttt attttcatta catctgtgtg ttggtttttt
     5161 gtgtgaatcg atagcgataa ggatccgctc atgcgatgat aagctgtcaa acatgagaat
     5221 tacaacttat atcgtatggg gctgacttca ggtgctacat ttgaagagat aaattgcact
     5281 gaaatctaga aatattttat ctgattaata agatgatctt cttgagatcg ttttggtctg
     5341 cgcgtaatct cttgctctga aaacgaaaaa accgccttgc agggcggttt ttcgaaggtt
     5401 ctctgagcta ccaactcttt gaaccgaggt aactggcttg gaggagcgca gtcaccaaaa
     5461 cttgtccttt cagtttagcc ttaaccggcg catgacttca agactaactc ctctaaatca
     5521 attaccagtg gctgctgcca gtggtgcttt tgcatgtctt tccgggttgg actcaagacg
     5581 atagttaccg gataaggcgc agcggtcgga ctgaacgggg ggttcgtgca tacagtccag
     5641 cttggagcga actgcctacc cggaactgag tgtcaggcgt ggaatgagac aaacgcggcc
     5701 ataacagcgg aatgacaccg gtaaaccgaa aggcaggaac aggagagcgc acgagggagc
     5761 cgccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac cactgatttg
     5821 agcgtcagat ttcgtgatgc ttgtcagggg ggcggagcct atggaaaaac ggctttgccg
     5881 cggccctctc acttccctgt taagtatctt cctggcatct tccaggaaat ctccgccccg
     5941 ttcgtaagcc atttccgctc gccgcagtcg aacgaccgag cgtagcgagt cagtgagcga
     6001 ggaagcggaa tatatcctgt atcacatatt ctgctgacgc accggtgcag ccttttttct
     6061 cctgccacat gaagcacttc actgacaccc tcatcagtgc caacatagta agccagtata
     6121 cactccgcta gcgctgatgt ccggcggtgc ttttgccgtt acgcaccacc ccgtcatgag
     6181 attacgcgtt ttagcggtgc gtacaattaa gggattatgg taaatccact tactgtctgc
     6241 cctcgtagcc atcgagataa accgcagtac tccggccacg atgcgtccgg cgtagaggat
     6301 cgagatct
//
