LOCUS       sequence-474672-        5395 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express IFNb in luciferase reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5395)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5395)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: pNL [NlucP/IFNb/Hygro] Vector
FEATURES             Location/Qualifiers
     source          1..5395
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      433..442
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             439..951
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             955..1074
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     polyA_signal    1126..1247
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1137..1156)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1191..1210
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        1441..1798
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1441..1632
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1464..1484)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1649..1784
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1711..1730
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1829..2866
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             2845..2853
                     /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    2890..2938
                     /note="synthetic polyadenylation signal"
     rep_origin      3375..3920
                     /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     3821..3840
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4277..5068,5069..5137))
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     polyA_signal    5242..5290
                     /note="synthetic polyadenylation signal"
     misc_feature    5304..5395
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     5344..5363
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggcctaactg gccggtacca actgtatctt ttagtgtttt gaggttcttg aattctcagg
       61 tcgtttgctt tcctttgctt tctcccaagt cttgttttac aatttgcttt agtcattcac
      121 tgaaacttta aaaaacatta gaaaacctca cagtttgtaa atctttttcc ctattatata
      181 tatcataaga taggagctta aataaagagt tttagaaact actaaaatgt aaatgacata
      241 ggaaaactga aagggagaag tgaaagtggg aaattcctct gaatagagag aggaccatct
      301 catataaata ggccataccc atggagaaag gacattctaa ctgcaacctt tcgaagcctt
      361 tgctctggca caacaggtag taggcgacac tgttcgtgtt gtcaagcttg gcaatccggt
      421 actgttggta aagccaccat ggtcttcaca ctcgaagatt tcgttgggga ctggcgacag
      481 acagccggct acaacctgga ccaagtcctt gaacagggag gtgtgtccag tttgtttcag
      541 aatctcgggg tgtccgtaac tccgatccaa aggattgtcc tgagcggtga aaatgggctg
      601 aagatcgaca tccatgtcat catcccgtat gaaggtctga gcggcgacca aatgggccag
      661 atcgaaaaaa tttttaaggt ggtgtaccct gtggatgatc atcactttaa ggtgatcctg
      721 cactatggca cactggtaat cgacggggtt acgccgaaca tgatcgacta tttcggacgg
      781 ccgtatgaag gcatcgccgt gttcgacggc aaaaagatca ctgtaacagg gaccctgtgg
      841 aacggcaaca aaattatcga cgagcgcctg atcaaccccg acggctccct gctgttccga
      901 gtaaccatca acggagtgac cggctggcgg ctgtgcgaac gcattctggc gaattctcac
      961 ggctttccgc ctgaggttga agagcaagcc gccggtacat tgcctatgtc ctgcgcacaa
     1021 gaaagcggta tggaccggca cccagccgct tgtgcttcag ctcgcatcaa cgtctaaggc
     1081 cgcgactcta gagtcggggc ggccggccgc ttcgagcaga catgataaga tacattgatg
     1141 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     1201 atgctattgc tttatttgta accattataa gctgcaataa acaagttaac aacaacaatt
     1261 gcattcattt tatgtttcag gttcaggggg aggtgtggga ggttttttaa agcaagtaaa
     1321 acctctacaa atgtggtaaa atcgataagg atccgtttgc gtattgggcg ctcttccgct
     1381 gatctgcgca gcaccatggc ctgaaataac ctctgaaaga ggaacttggt tagctacctt
     1441 ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg
     1501 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg
     1561 aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc
     1621 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     1681 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     1741 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     1801 gctcgattct tctgacacta gcgccaccat gaagaagccc gaactcaccg ctaccagcgt
     1861 tgaaaaattt ctcatcgaga agttcgacag tgtgagcgac ctgatgcagt tgtcggaggg
     1921 cgaagagagc cgagccttca gcttcgatgt cggcggacgc ggctatgtac tgcgggtgaa
     1981 tagctgcgct gatggcttct acaaagaccg ctacgtgtac cgccacttcg ccagcgctgc
     2041 actacccatc cccgaagtgt tggacatcgg cgagttcagc gagagcctga catactgcat
     2101 cagtagacgc gcccaaggcg ttactctcca agacctcccc gaaacagagc tgcctgctgt
     2161 gttacagcct gtcgccgaag ctatggatgc tattgccgcc gccgacctca gtcaaaccag
     2221 cggcttcggc ccattcgggc cccaaggcat cggccagtac acaacctggc gggatttcat
     2281 ttgcgccatt gctgatcccc atgtctacca ctggcagacc gtgatggacg acaccgtgtc
     2341 cgccagcgta gctcaagccc tggacgaact gatgctgtgg gccgaagact gtcccgaggt
     2401 gcgccacctc gtccatgccg acttcggcag caacaacgtc ctgaccgaca acggccgcat
     2461 caccgccgta atcgactggt ccgaagctat gttcggggac agtcagtacg aggtggccaa
     2521 catcttcttc tggcggccct ggctggcttg catggagcag cagactcgct acttcgagcg
     2581 ccggcatccc gagctggccg gcagccctcg tctgcgagcc tacatgctgc gcatcggcct
     2641 ggatcagctc taccagagcc tcgtggacgg caacttcgac gatgctgcct gggctcaagg
     2701 ccgctgcgat gccatcgtcc gcagcggggc cggcaccgtc ggtcgcacac aaatcgctcg
     2761 ccggagcgca gccgtatgga ccgacggctg cgtcgaggtg ctggccgaca gcggcaaccg
     2821 ccggcccagt acacgaccgc gcgctaagga ggtaggtcga gtttaaactc tagaaccggt
     2881 catggccgca ataaaatatc tttattttca ttacatctgt gtgttggttt tttgtgtgtt
     2941 cgaactagat gctgtcgacc gatgcccttg agagccttca acccagtcag ctccttccgg
     3001 tgggcgcggg gcatgactat cgtcgccgca cttatgactg tcttctttat catgcaactc
     3061 gtaggacagg tgccggcagc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     3121 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     3181 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     3241 taaaaaggcc gcgttgctgg caaacatgag aattacaact tatatcgtat ggggctgact
     3301 tcaggtgcta catttgaaga gataaattgc actgaaatct agaaatattt tatctgatta
     3361 ataagatgat cttcttgaga tcgttttggt ctgcgcgtaa tctcttgctc tgaaaacgaa
     3421 aaaaccgcct tgcagggcgg tttttcgaag gttctctgag ctaccaactc tttgaaccga
     3481 ggtaactggc ttggaggagc gcagtcacca aaacttgtcc tttcagttta gccttaaccg
     3541 gcgcatgact tcaagactaa ctcctctaaa tcaattacca gtggctgctg ccagtggtgc
     3601 ttttgcatgt ctttccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc
     3661 ggactgaacg gggggttcgt gcatacagtc cagcttggag cgaactgcct acccggaact
     3721 gagtgtcagg cgtggaatga gacaaacgcg gccataacag cggaatgaca ccggtaaacc
     3781 gaaaggcagg aacaggagag cgcacgaggg agccgccagg gggaaacgcc tggtatcttt
     3841 atagtcctgt cgggtttcgc caccactgat ttgagcgtca gatttcgtga tgcttgtcag
     3901 gggggcggag cctatggaaa aacggctttg ccgcggccct ctcacttccc tgttaagtat
     3961 cttcctggca tcttccagga aatctccgcc ccgttcgtaa gccatttccg ctcgccgcag
     4021 tcgaacgacc gagcgtagcg agtcagtgag cgaggaagcg gaatatatcc tgtatcacat
     4081 attctgctga cgcaccggtg cagccttttt tctcctgcca catgaagcac ttcactgaca
     4141 ccctcatcag tgccaacata gtaagccagt atacactccg ctattttaaa ttaaaaatga
     4201 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagcgg ccgcaaatgc
     4261 taaaccactg cagtggttac cagtgcttga tcagtgaggc accgatctca gcgatctgcc
     4321 tatttcgttc gtccatagtg gcctgactcc ccgtcgtgta gatcactacg attcgtgagg
     4381 gcttaccatc aggccccagc gcagcaatga tgccgcgaga gccgcgttca ccggcccccg
     4441 atttgtcagc aatgaaccag ccagcaggga gggccgagcg aagaagtggt cctgctactt
     4501 tgtccgcctc catccagtct atgagctgct gtcgtgatgc tagagtaaga agttcgccag
     4561 tgagtagttt ccgaagagtt gtggccattg ctactggcat cgtggtatca cgctcgtcgt
     4621 tcggtatggc ttcgttcaac tctggttccc agcggtcaag ccgggtcaca tgatcaccca
     4681 tattatgaag aaatgcagtc agctccttag ggcctccgat cgttgtcaga agtaagttgg
     4741 ccgcggtgtt gtcgctcatg gtaatggcag cactacacaa ttctcttacc gtcatgccat
     4801 ccgtaagatg cttttccgtg accggcgagt actcaaccaa gtcgttttgt gagtagtgta
     4861 tacggcgacc aagctgctct tgcccggcgt ctatacggga caacaccgcg ccacatagca
     4921 gtactttgaa agtgctcatc atcgggaatc gttcttcggg gcggaaagac tcaaggatct
     4981 tgccgctatt gagatccagt tcgatatagc ccactcttgc acccagttga tcttcagcat
     5041 cttttacttt caccagcgtt tcggggtgtg caaaaacagg caagcaaaat gccgcaaaga
     5101 agggaatgag tgcgacacga aaatgttgga tgctcatact cgtccttttt caatattatt
     5161 gaagcattta tcagggttac tagtacgtct ctcaaggata agtaagtaat attaaggtac
     5221 gggaggtatt ggacaggccg caataaaata tctttatttt cattacatct gtgtgttggt
     5281 tttttgtgtg aatcgatagt actaacatac gctctccatc aaaacaaaac gaaacaaaac
     5341 aaactagcaa aataggctgt ccccagtgca agtgcaggtg ccagaacatt tctct
//
