LOCUS       sequence-471834-        4992 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express SIE response element in luciferase reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4992)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4992)
  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            "Unpublished"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/SIE/Hygro] Vector
FEATURES             Location/Qualifiers
     source          1..4992
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        196..227
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      254..263
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             260..772
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             776..895
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     polyA_signal    947..1068
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(958..977)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1012..1031
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        1262..1619
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1262..1453
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1285..1305)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1470..1605
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1532..1551
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1650..2687
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             2666..2674
                     /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    2711..2759
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(2915..2932)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3086..3674)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3166..3185)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3874..4665,4666..4734))
                     /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    4839..4887
                     /note="synthetic polyadenylation signal"
     misc_feature    4901..4992
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4941..4960
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggcctaactg gccggtacct gagctcagct tcatttcccg taaatcgtcg aagcttcatt
       61 tcccgtaaat cgtcgaagct tcatttcccg taaatcgtcg aagcttcatt tcccgtaaat
      121 cgtcgaagct tcatttcccg taaatcgtcg actcgaggat atcaagatct ggcctcggcg
      181 gccaagctta gacactagag ggtatataat ggaagctcga cttccagctt ggcaatccgg
      241 tactgttggt aaagccacca tggtcttcac actcgaagat ttcgttgggg actggcgaca
      301 gacagccggc tacaacctgg accaagtcct tgaacaggga ggtgtgtcca gtttgtttca
      361 gaatctcggg gtgtccgtaa ctccgatcca aaggattgtc ctgagcggtg aaaatgggct
      421 gaagatcgac atccatgtca tcatcccgta tgaaggtctg agcggcgacc aaatgggcca
      481 gatcgaaaaa atttttaagg tggtgtaccc tgtggatgat catcacttta aggtgatcct
      541 gcactatggc acactggtaa tcgacggggt tacgccgaac atgatcgact atttcggacg
      601 gccgtatgaa ggcatcgccg tgttcgacgg caaaaagatc actgtaacag ggaccctgtg
      661 gaacggcaac aaaattatcg acgagcgcct gatcaacccc gacggctccc tgctgttccg
      721 agtaaccatc aacggagtga ccggctggcg gctgtgcgaa cgcattctgg cgaattctca
      781 cggctttccg cctgaggttg aagagcaagc cgccggtaca ttgcctatgt cctgcgcaca
      841 agaaagcggt atggaccggc acccagccgc ttgtgcttca gctcgcatca acgtctaagg
      901 ccgcgactct agagtcgggg cggccggccg cttcgagcag acatgataag atacattgat
      961 gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt
     1021 gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat
     1081 tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa
     1141 aacctctaca aatgtggtaa aatcgataag gatccgtttg cgtattgggc gctcttccgc
     1201 tgatctgcgc agcaccatgg cctgaaataa cctctgaaag aggaacttgg ttagctacct
     1261 tctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     1321 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     1381 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     1441 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     1501 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     1561 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     1621 agctcgattc ttctgacact agcgccacca tgaagaagcc cgaactcacc gctaccagcg
     1681 ttgaaaaatt tctcatcgag aagttcgaca gtgtgagcga cctgatgcag ttgtcggagg
     1741 gcgaagagag ccgagccttc agcttcgatg tcggcggacg cggctatgta ctgcgggtga
     1801 atagctgcgc tgatggcttc tacaaagacc gctacgtgta ccgccacttc gccagcgctg
     1861 cactacccat ccccgaagtg ttggacatcg gcgagttcag cgagagcctg acatactgca
     1921 tcagtagacg cgcccaaggc gttactctcc aagacctccc cgaaacagag ctgcctgctg
     1981 tgttacagcc tgtcgccgaa gctatggatg ctattgccgc cgccgacctc agtcaaacca
     2041 gcggcttcgg cccattcggg ccccaaggca tcggccagta cacaacctgg cgggatttca
     2101 tttgcgccat tgctgatccc catgtctacc actggcagac cgtgatggac gacaccgtgt
     2161 ccgccagcgt agctcaagcc ctggacgaac tgatgctgtg ggccgaagac tgtcccgagg
     2221 tgcgccacct cgtccatgcc gacttcggca gcaacaacgt cctgaccgac aacggccgca
     2281 tcaccgccgt aatcgactgg tccgaagcta tgttcgggga cagtcagtac gaggtggcca
     2341 acatcttctt ctggcggccc tggctggctt gcatggagca gcagactcgc tacttcgagc
     2401 gccggcatcc cgagctggcc ggcagccctc gtctgcgagc ctacatgctg cgcatcggcc
     2461 tggatcagct ctaccagagc ctcgtggacg gcaacttcga cgatgctgcc tgggctcaag
     2521 gccgctgcga tgccatcgtc cgcagcgggg ccggcaccgt cggtcgcaca caaatcgctc
     2581 gccggagcgc agccgtatgg accgacggct gcgtcgaggt gctggccgac agcggcaacc
     2641 gccggcccag tacacgaccg cgcgctaagg aggtaggtcg agtttaaact ctagaaccgg
     2701 tcatggccgc aataaaatat ctttattttc attacatctg tgtgttggtt ttttgtgtgt
     2761 tcgaactaga tgctgtcgac cgatgccctt gagagccttc aacccagtca gctccttccg
     2821 gtgggcgcgg ggcatgacta tcgtcgccgc acttatgact gtcttcttta tcatgcaact
     2881 cgtaggacag gtgccggcag cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     2941 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3001 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3061 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3121 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3181 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3241 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3301 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3361 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3421 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3481 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     3541 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3601 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3661 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3721 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     3781 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     3841 acagcggccg caaatgctaa accactgcag tggttaccag tgcttgatca gtgaggcacc
     3901 gatctcagcg atctgcctat ttcgttcgtc catagtggcc tgactccccg tcgtgtagat
     3961 cactacgatt cgtgagggct taccatcagg ccccagcgca gcaatgatgc cgcgagagcc
     4021 gcgttcaccg gcccccgatt tgtcagcaat gaaccagcca gcagggaggg ccgagcgaag
     4081 aagtggtcct gctactttgt ccgcctccat ccagtctatg agctgctgtc gtgatgctag
     4141 agtaagaagt tcgccagtga gtagtttccg aagagttgtg gccattgcta ctggcatcgt
     4201 ggtatcacgc tcgtcgttcg gtatggcttc gttcaactct ggttcccagc ggtcaagccg
     4261 ggtcacatga tcacccatat tatgaagaaa tgcagtcagc tccttagggc ctccgatcgt
     4321 tgtcagaagt aagttggccg cggtgttgtc gctcatggta atggcagcac tacacaattc
     4381 tcttaccgtc atgccatccg taagatgctt ttccgtgacc ggcgagtact caaccaagtc
     4441 gttttgtgag tagtgtatac ggcgaccaag ctgctcttgc ccggcgtcta tacgggacaa
     4501 caccgcgcca catagcagta ctttgaaagt gctcatcatc gggaatcgtt cttcggggcg
     4561 gaaagactca aggatcttgc cgctattgag atccagttcg atatagccca ctcttgcacc
     4621 cagttgatct tcagcatctt ttactttcac cagcgtttcg gggtgtgcaa aaacaggcaa
     4681 gcaaaatgcc gcaaagaagg gaatgagtgc gacacgaaaa tgttggatgc tcatactcgt
     4741 cctttttcaa tattattgaa gcatttatca gggttactag tacgtctctc aaggataagt
     4801 aagtaatatt aaggtacggg aggtattgga caggccgcaa taaaatatct ttattttcat
     4861 tacatctgtg tgttggtttt ttgtgtgaat cgatagtact aacatacgct ctccatcaaa
     4921 acaaaacgaa acaaaacaaa ctagcaaaat aggctgtccc cagtgcaagt gcaggtgcca
     4981 gaacatttct ct
//
