LOCUS       sequence-479793-        5849 bp    DNA     circular SYN 07-AUG-2025
DEFINITION  Express NanoLuc-MAPK14 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 5849)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5849)
  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            "Unpublished"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: NanoLuc-MAPK14 Fusion Vector
FEATURES             Location/Qualifiers
     source          1..5849
                     /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"
     RBS             2590..2598
                     /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)"
     primer_bind     complement(2792..2810)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2847..2968
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2858..2877)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2912..2931
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3213..3570
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3213..3404
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3236..3256)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3421..3556
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3585..3632
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3664..4458
                     /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(3718..3737)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4328..4347
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4522..4570
                     /note="synthetic polyadenylation signal"
     rep_origin      complement(4809..5397)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4889..4908)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     misc_feature    5513..5796
                     /label=cer region
                     /note="ColE1-derived recombination site that helps to 
                     maintain plasmids as monomers"
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 gtctcaggag aggcccacgt
     1621 tctaccggca ggagctgaac aagacaatct gggaggtgcc cgagcgttac cagaacctgt
     1681 ctccagtggg ctctggcgcc tatggctctg tgtgtgctgc ttttgacaca aaaacggggt
     1741 tacgtgtggc agtgaagaag ctctccagac catttcagtc catcattcat gcgaaaagaa
     1801 cctacagaga actgcggtta cttaaacata tgaaacatga aaatgtgatt ggtctgttgg
     1861 acgtttttac acctgcaagg tctctggagg aattcaatga tgtgtatctg gtgacccatc
     1921 tcatgggggc agatctgaac aacattgtga aatgtcagaa gcttacagat gaccatgttc
     1981 agttccttat ctaccaaatt ctccgaggtc taaagtatat acattcagct gacataattc
     2041 acagggacct aaaacctagt aatctagctg tgaatgaaga ctgtgagctg aagattctgg
     2101 attttggact ggctcggcac acagatgatg aaatgacagg ctacgtggcc actaggtggt
     2161 acagggctcc tgagatcatg ctgaactgga tgcattacaa ccagacagtt gatatttggt
     2221 cagtgggatg cataatggcc gagctgttga ctggaagaac attgtttcct ggtacagacc
     2281 atattaacca gcttcagcag attatgcgtc tgacaggaac accccccgct tatctcatta
     2341 acaggatgcc aagccatgag gcaagaaact atattcagtc tttgactcag atgccgaaga
     2401 tgaactttgc gaatgtattt attggtgcca atcccctggc tgtcgacttg ctggagaaga
     2461 tgcttgtatt ggactcagat aagagaatta cagcggccca agcccttgca catgcctact
     2521 ttgctcagta ccacgatcct gatgatgaac cagtggccga tccttatgat cagtcctttg
     2581 aaagcaggga cctccttata gatgagtgga aaagcctgac ctatgatgaa gtcatcagct
     2641 ttgtgccacc accccttgac caagaagaga tggagtccgt ttaaacgaat tcgggctcgg
     2701 tacccgggga tcctctagag tcgacctgca ggcatgcaag ctgatccggc tgctaacaaa
     2761 gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt
     2821 ggggcggccg cttcgagcag acatgataag atacattgat gagtttggac aaaccacaac
     2881 tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg ctttatttgt
     2941 aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt ttatgtttca
     3001 ggttcagggg gagatgtggg aggttttttt aagcaagtaa aacctctaca aatgtggtaa
     3061 aatcgaattt taacaaaata ttaacgctta caatttcctg atgcggtatt ttctccttac
     3121 gcatctgtgc ggtatttcac accgcatacg cggatctgcg cagcaccatg gcctgaaata
     3181 acctctgaaa gaggaacttg gttaggtacc ttctgaggcg gaaagaacca gctgtggaat
     3241 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     3301 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     3361 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     3421 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     3481 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     3541 ggcttttttg gaggcctagg cttttgcaaa aagcttaatt aactgttgac aattaatcat
     3601 cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc caggaggcag
     3661 atcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     3721 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     3781 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     3841 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     3901 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     3961 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     4021 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     4081 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     4141 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     4201 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     4261 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     4321 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     4381 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     4441 cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca
     4501 acctgccatc acgatggccg caataaaata tctttatttt cattacatct gtgtgttggt
     4561 tttttgtgtg aatcgatagc gataaggatc ctctttgcgc ttgcgttttc ccttgtccag
     4621 atagcccagt agctgacatt catccggggt cagcaccgtt tctgcggact ggctttctac
     4681 ccggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
     4741 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
     4801 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
     4861 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
     4921 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
     4981 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
     5041 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
     5101 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
     5161 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
     5221 tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc tctgctgaag
     5281 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
     5341 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atttcaagaa
     5401 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
     5461 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttata gtccggaaat
     5521 acaggaacgc acgctggatg gcccttcgct gggatggtga aaccatgaaa aatggcagct
     5581 tcagtggatt aagtgggggt aatgtggcct gtaccctctg gttgcatagg tattcatacg
     5641 gttaaaattt atcaggcgcg attgcggcag tttttcgggt ggtttgttgc catttttacc
     5701 tgtctgctgc cgtgatcgcg ctgaacgcgt tttagcggtg cgtacaatta agggattatg
     5761 gtaaatccac ttactgtctg ccctcgtagc catcgagata aaccgcagta ctccggccac
     5821 gatgcgtccg gcgtagagga tcgagatct
//
