LOCUS       sequence-470888-        5636 bp    DNA     circular SYN 02-MAY-2025
DEFINITION  Express PNCK-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 5636)
  TITLE     Promega Early Access Collection
REFERENCE   2  (bases 1 to 5636)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 2, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: PNCK-NanoLuc Fusion Vector
FEATURES             Location/Qualifiers
     source          1..5636
                     /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             2115..2627
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     primer_bind     complement(2715..2733)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     polyA_signal    2770..2891
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2781..2800)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2835..2854
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        3136..3493
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3136..3327
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3159..3179)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3344..3479
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        3508..3555
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3587..4381
                     /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(3641..3660)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4251..4270
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4445..4493
                     /note="synthetic polyadenylation signal"
     rep_origin      4650..5195
                     /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     5096..5115
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5312..5329
                     /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 cgccatgctg ctgctgaaga
     1081 agcacaccga ggacatcagc tccgtgtatg agatcaggga gcgcctggga tccggagcct
     1141 tctctgaggt ggtgctggca caggagaggg gatccgccca cctggtggcc ctgaagtgca
     1201 tcccaaagaa ggccctgagg ggcaaggagg ccctggtgga gaacgagatc gccgtgctgc
     1261 ggagaatctc tcacccaaat atcgtggccc tggaggacgt gcacgagtct cccagccacc
     1321 tgtacctggc aatggagctg gtgaccggag gagagctgtt tgacaggatc atggagcgcg
     1381 gcagctacac agagaaggat gcctcccacc tggtgggaca ggtgctggga gccgtgagct
     1441 atctgcactc cctgggcatc gtgcaccggg atctgaagcc cgagaacctg ctgtacgcca
     1501 cacctttcga ggactctaag atcatggtga gcgattttgg cctgtccaag atccaggcag
     1561 gaaatatgct gggaaccgca tgcggaacac ctggatacgt ggcaccagag ctgctggagc
     1621 agaagcctta tggcaaggca gtggacgtgt gggcactggg cgtgatctcc tatatcctgc
     1681 tgtgcggcta cccccctttc tatgacgagt ccgatcctga gctgttttct cagatcctgc
     1741 gggcctctta cgagttcgat agcccatttt gggacgatat ctccgagtct gccaaggact
     1801 tcatccggca cctgctggag agagatcccc agaagaggtt tacctgtcag caggccctga
     1861 ggcacctgtg gatcagcggc gatacagcct tcgacagaga tatcctgggc agcgtgtccg
     1921 agcagatccg caagaacttt gcccggaccc actggaagag agccttcaat gccacatctt
     1981 ttctgcggca catcagaaag ctgggccaga tccctgaggg agagggagca agcgagcagg
     2041 gcatggccag gcactctcac agcggcctgc gcgccggcca gccacccaag tgggtttctc
     2101 tcggctcgag cggcgtcttc acactcgaag atttcgttgg ggactggcga cagacagccg
     2161 gctacaacct ggaccaagtc cttgaacagg gaggtgtgtc cagtttgttt cagaatctcg
     2221 gggtgtccgt aactccgatc caaaggattg tcctgagcgg tgaaaatggg ctgaagatcg
     2281 acatccatgt catcatcccg tatgaaggtc tgagcggcga ccaaatgggc cagatcgaaa
     2341 aaatttttaa ggtggtgtac cctgtggatg atcatcactt taaggtgatc ctgcactatg
     2401 gcacactggt aatcgacggg gttacgccga acatgatcga ctatttcgga cggccgtatg
     2461 aaggcatcgc cgtgttcgac ggcaaaaaga tcactgtaac agggaccctg tggaacggca
     2521 acaaaattat cgacgagcgc ctgatcaacc ccgacggctc cctgctgttc cgagtaacca
     2581 tcaacggagt gaccggctgg cggctgtgcg aacgcattct ggcgtaaggc cgcgactcta
     2641 gagtcgacct gcaggcatgc aagctgatcc ggctgctaac aaagcccgaa aggaagctga
     2701 gttggctgct gccaccgctg agcaataact agcataaccc cttggggcgg ccgcttcgag
     2761 cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg cagtgaaaaa
     2821 aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt ataagctgca
     2881 ataaacaagt taacaacaac aattgcattc attttatgtt tcaggttcag ggggagatgt
     2941 gggaggtttt tttaagcaag taaaacctct acaaatgtgg taaaatcgaa ttttaacaaa
     3001 atattaacgc ttacaatttc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     3061 cacaccgcat acgcggatct gcgcagcacc atggcctgaa ataacctctg aaagaggaac
     3121 ttggttaggt accttctgag gcggaaagaa ccagctgtgg aatgtgtgtc agttagggtg
     3181 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     3241 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     3301 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     3361 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3421 cgaggccgcc tcggcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3481 aggcttttgc aaaaagctta attaactgtt gacaattaat catcggcata gtatatcggc
     3541 atagtataat acgacaaggt gaggaactaa acccaggagg cagatcatga ttgaacaaga
     3601 tggattgcac gcaggttctc cggccgcttg ggtggagagg ctattcggct atgactgggc
     3661 acaacagaca atcggctgct ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc
     3721 ggttcttttt gtcaagaccg acctgtccgg tgccctgaat gaactgcagg acgaggcagc
     3781 gcggctatcg tggctggcca cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac
     3841 tgaagcggga agggactggc tgctattggg cgaagtgccg gggcaggatc tcctgtcatc
     3901 tcaccttgct cctgccgaga aagtatccat catggctgat gcaatgcggc ggctgcatac
     3961 gcttgatccg gctacctgcc cattcgacca ccaagcgaaa catcgcatcg agcgagcacg
     4021 tactcggatg gaagccggtc ttgtcgatca ggatgatctg gacgaagagc atcaggggct
     4081 cgcgccagcc gaactgttcg ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt
     4141 cgtgacccat ggcgatgcct gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg
     4201 attcatcgac tgtggccggc tgggtgtggc ggaccgctat caggacatag cgttggctac
     4261 ccgtgatatt gctgaagagc ttggcggcga atgggctgac cgcttcctcg tgctttacgg
     4321 tatcgccgct cccgattcgc agcgcatcgc cttctatcgc cttcttgacg agttcttctg
     4381 agcgggactc tggggttcga aatgaccgac caagcgacgc ccaacctgcc atcacgatgg
     4441 ccgcaataaa atatctttat tttcattaca tctgtgtgtt ggttttttgt gtgaatcgat
     4501 agcgataagg atccgctcat gcgatgataa gctgtcaaac atgagaatta caacttatat
     4561 cgtatggggc tgacttcagg tgctacattt gaagagataa attgcactga aatctagaaa
     4621 tattttatct gattaataag atgatcttct tgagatcgtt ttggtctgcg cgtaatctct
     4681 tgctctgaaa acgaaaaaac cgccttgcag ggcggttttt cgaaggttct ctgagctacc
     4741 aactctttga accgaggtaa ctggcttgga ggagcgcagt caccaaaact tgtcctttca
     4801 gtttagcctt aaccggcgca tgacttcaag actaactcct ctaaatcaat taccagtggc
     4861 tgctgccagt ggtgcttttg catgtctttc cgggttggac tcaagacgat agttaccgga
     4921 taaggcgcag cggtcggact gaacgggggg ttcgtgcata cagtccagct tggagcgaac
     4981 tgcctacccg gaactgagtg tcaggcgtgg aatgagacaa acgcggccat aacagcggaa
     5041 tgacaccggt aaaccgaaag gcaggaacag gagagcgcac gagggagccg ccagggggaa
     5101 acgcctggta tctttatagt cctgtcgggt ttcgccacca ctgatttgag cgtcagattt
     5161 cgtgatgctt gtcagggggg cggagcctat ggaaaaacgg ctttgccgcg gccctctcac
     5221 ttccctgtta agtatcttcc tggcatcttc caggaaatct ccgccccgtt cgtaagccat
     5281 ttccgctcgc cgcagtcgaa cgaccgagcg tagcgagtca gtgagcgagg aagcggaata
     5341 tatcctgtat cacatattct gctgacgcac cggtgcagcc ttttttctcc tgccacatga
     5401 agcacttcac tgacaccctc atcagtgcca acatagtaag ccagtataca ctccgctagc
     5461 gctgatgtcc ggcggtgctt ttgccgttac gcaccacccc gtcatgagat tacgcgtttt
     5521 agcggtgcgt acaattaagg gattatggta aatccactta ctgtctgccc tcgtagccat
     5581 cgagataaac cgcagtactc cggccacgat gcgtccggcg tagaggatcg agatct
//
