LOCUS       sequence-474673-        4950 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express NFAT response element in luciferase reporter assay.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4950)
  AUTHORS   Promega
  TITLE     Promega Plasmid Collection
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 4950)
  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            "Unpublished"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/NFAT-RE/Hygro] Vector
FEATURES             Location/Qualifiers
     source          1..4950
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    33..62
                     /label=NFAT binding site
                     /bound_moiety="NFAT transcription factor"
                     /note="NFAT binding site from the human interleukin 2 
                     (IL-2) gene (Mattila et al., 1990)"
     protein_bind    63..92
                     /label=NFAT binding site
                     /bound_moiety="NFAT transcription factor"
                     /note="NFAT binding site from the human interleukin 2 
                     (IL-2) gene (Mattila et al., 1990)"
     protein_bind    93..122
                     /label=NFAT binding site
                     /bound_moiety="NFAT transcription factor"
                     /note="NFAT binding site from the human interleukin 2 
                     (IL-2) gene (Mattila et al., 1990)"
     promoter        154..185
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     regulatory      212..221
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             218..730
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="MVFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQR
                     IVLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGV
                     TPNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGW
                     RLCERILA"
     CDS             734..853
                     /codon_start=1
                     /product="PEST degradation sequence from mouse ornithine 
                     decarboxylase"
                     /label=hPEST
                     /note="human codon-optimized"
                     /translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
     polyA_signal    905..1026
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(916..935)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     970..989
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        1220..1577
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        1220..1411
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(1243..1263)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1428..1563
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1490..1509
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1608..2645
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     EVGRV"
     RBS             2624..2632
                     /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    2669..2717
                     /note="synthetic polyadenylation signal"
     primer_bind     complement(2873..2890)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3044..3632)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3124..3143)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3832..4623,4624..4692))
                     /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    4797..4845
                     /note="synthetic polyadenylation signal"
     misc_feature    4859..4950
                     /label=pause site
                     /note="RNA polymerase II transcriptional pause signal from 
                     the human alpha-2 globin gene"
     primer_bind     4899..4918
                     /label=RVprimer3
                     /note="pGL3 vector, forward primer"
ORIGIN
        1 ggcctaactg gccggtacct gagctcgcta gcggaggaaa aactgtttca tacagaaggc
       61 gtggaggaaa aactgtttca tacagaaggc gtggaggaaa aactgtttca tacagaaggc
      121 gtagatctgg cctcggcggc caagcttaga cactagaggg tatataatgg aagctcgact
      181 tccagcttgg caatccggta ctgttggtaa agccaccatg gtcttcacac tcgaagattt
      241 cgttggggac tggcgacaga cagccggcta caacctggac caagtccttg aacagggagg
      301 tgtgtccagt ttgtttcaga atctcggggt gtccgtaact ccgatccaaa ggattgtcct
      361 gagcggtgaa aatgggctga agatcgacat ccatgtcatc atcccgtatg aaggtctgag
      421 cggcgaccaa atgggccaga tcgaaaaaat ttttaaggtg gtgtaccctg tggatgatca
      481 tcactttaag gtgatcctgc actatggcac actggtaatc gacggggtta cgccgaacat
      541 gatcgactat ttcggacggc cgtatgaagg catcgccgtg ttcgacggca aaaagatcac
      601 tgtaacaggg accctgtgga acggcaacaa aattatcgac gagcgcctga tcaaccccga
      661 cggctccctg ctgttccgag taaccatcaa cggagtgacc ggctggcggc tgtgcgaacg
      721 cattctggcg aattctcacg gctttccgcc tgaggttgaa gagcaagccg ccggtacatt
      781 gcctatgtcc tgcgcacaag aaagcggtat ggaccggcac ccagccgctt gtgcttcagc
      841 tcgcatcaac gtctaaggcc gcgactctag agtcggggcg gccggccgct tcgagcagac
      901 atgataagat acattgatga gtttggacaa accacaacta gaatgcagtg aaaaaaatgc
      961 tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag ctgcaataaa
     1021 caagttaaca acaacaattg cattcatttt atgtttcagg ttcaggggga ggtgtgggag
     1081 gttttttaaa gcaagtaaaa cctctacaaa tgtggtaaaa tcgataagga tccgtttgcg
     1141 tattgggcgc tcttccgctg atctgcgcag caccatggcc tgaaataacc tctgaaagag
     1201 gaacttggtt agctaccttc tgaggcggaa agaaccagct gtggaatgtg tgtcagttag
     1261 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     1321 agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca
     1381 tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     1441 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     1501 aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     1561 gcctaggctt ttgcaaaaag ctcgattctt ctgacactag cgccaccatg aagaagcccg
     1621 aactcaccgc taccagcgtt gaaaaatttc tcatcgagaa gttcgacagt gtgagcgacc
     1681 tgatgcagtt gtcggagggc gaagagagcc gagccttcag cttcgatgtc ggcggacgcg
     1741 gctatgtact gcgggtgaat agctgcgctg atggcttcta caaagaccgc tacgtgtacc
     1801 gccacttcgc cagcgctgca ctacccatcc ccgaagtgtt ggacatcggc gagttcagcg
     1861 agagcctgac atactgcatc agtagacgcg cccaaggcgt tactctccaa gacctccccg
     1921 aaacagagct gcctgctgtg ttacagcctg tcgccgaagc tatggatgct attgccgccg
     1981 ccgacctcag tcaaaccagc ggcttcggcc cattcgggcc ccaaggcatc ggccagtaca
     2041 caacctggcg ggatttcatt tgcgccattg ctgatcccca tgtctaccac tggcagaccg
     2101 tgatggacga caccgtgtcc gccagcgtag ctcaagccct ggacgaactg atgctgtggg
     2161 ccgaagactg tcccgaggtg cgccacctcg tccatgccga cttcggcagc aacaacgtcc
     2221 tgaccgacaa cggccgcatc accgccgtaa tcgactggtc cgaagctatg ttcggggaca
     2281 gtcagtacga ggtggccaac atcttcttct ggcggccctg gctggcttgc atggagcagc
     2341 agactcgcta cttcgagcgc cggcatcccg agctggccgg cagccctcgt ctgcgagcct
     2401 acatgctgcg catcggcctg gatcagctct accagagcct cgtggacggc aacttcgacg
     2461 atgctgcctg ggctcaaggc cgctgcgatg ccatcgtccg cagcggggcc ggcaccgtcg
     2521 gtcgcacaca aatcgctcgc cggagcgcag ccgtatggac cgacggctgc gtcgaggtgc
     2581 tggccgacag cggcaaccgc cggcccagta cacgaccgcg cgctaaggag gtaggtcgag
     2641 tttaaactct agaaccggtc atggccgcaa taaaatatct ttattttcat tacatctgtg
     2701 tgttggtttt ttgtgtgttc gaactagatg ctgtcgaccg atgcccttga gagccttcaa
     2761 cccagtcagc tccttccggt gggcgcgggg catgactatc gtcgccgcac ttatgactgt
     2821 cttctttatc atgcaactcg taggacaggt gccggcagcg ctcttccgct tcctcgctca
     2881 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     2941 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     3001 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3061 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     3121 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     3181 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     3241 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     3301 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3361 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     3421 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     3481 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     3541 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     3601 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     3661 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     3721 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     3781 atgagtaaac ttggtctgac agcggccgca aatgctaaac cactgcagtg gttaccagtg
     3841 cttgatcagt gaggcaccga tctcagcgat ctgcctattt cgttcgtcca tagtggcctg
     3901 actccccgtc gtgtagatca ctacgattcg tgagggctta ccatcaggcc ccagcgcagc
     3961 aatgatgccg cgagagccgc gttcaccggc ccccgatttg tcagcaatga accagccagc
     4021 agggagggcc gagcgaagaa gtggtcctgc tactttgtcc gcctccatcc agtctatgag
     4081 ctgctgtcgt gatgctagag taagaagttc gccagtgagt agtttccgaa gagttgtggc
     4141 cattgctact ggcatcgtgg tatcacgctc gtcgttcggt atggcttcgt tcaactctgg
     4201 ttcccagcgg tcaagccggg tcacatgatc acccatatta tgaagaaatg cagtcagctc
     4261 cttagggcct ccgatcgttg tcagaagtaa gttggccgcg gtgttgtcgc tcatggtaat
     4321 ggcagcacta cacaattctc ttaccgtcat gccatccgta agatgctttt ccgtgaccgg
     4381 cgagtactca accaagtcgt tttgtgagta gtgtatacgg cgaccaagct gctcttgccc
     4441 ggcgtctata cgggacaaca ccgcgccaca tagcagtact ttgaaagtgc tcatcatcgg
     4501 gaatcgttct tcggggcgga aagactcaag gatcttgccg ctattgagat ccagttcgat
     4561 atagcccact cttgcaccca gttgatcttc agcatctttt actttcacca gcgtttcggg
     4621 gtgtgcaaaa acaggcaagc aaaatgccgc aaagaaggga atgagtgcga cacgaaaatg
     4681 ttggatgctc atactcgtcc tttttcaata ttattgaagc atttatcagg gttactagta
     4741 cgtctctcaa ggataagtaa gtaatattaa ggtacgggag gtattggaca ggccgcaata
     4801 aaatatcttt attttcatta catctgtgtg ttggtttttt gtgtgaatcg atagtactaa
     4861 catacgctct ccatcaaaac aaaacgaaac aaaacaaact agcaaaatag gctgtcccca
     4921 gtgcaagtgc aggtgccaga acatttctct
//
