LOCUS       sequence-227706-        5654 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for NFAT activation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5654)
  AUTHORS   Jutz S, Leitner J, Schmetterer K, Doel-Perez I, Majdic O, 
            Grabmeier-Pfistershammer K, Paster W, Huppa JB, Steinberger P
  TITLE     Assessment of costimulation and coinhibition in a triple parameter T
            cell reporter line: Simultaneous measurement of NF-kappaB, NFAT and 
            AP-1.
  JOURNAL   J Immunol Methods. 2016 Jan 15. pii: S0022-1759(16)30007-2. doi: 
            10.1016/j.jim.2016.01.007.
  PUBMED    26780292
REFERENCE   2  (bases 1 to 5654)
  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: "J Immunol 
            Methods. 2016 Jan 15. pii: S0022-1759(16)30007-2. doi: 
            10.1016/j.jim.2016.01.007."
COMMENT     Sequence Label: pSIRV-NFAT-eGFP
FEATURES             Location/Qualifiers
     source          1..5654
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    55..412
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             477..893
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     823..845
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     861..877
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    903..1277
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     1215..1231
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     protein_bind    1291..1320
                     /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    1321..1350
                     /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    1351..1380
                     /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    1381..1410
                     /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        1430..1461
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     CDS             join(1502..1504,1505..1507,1508..2221)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1502..1504,1505..1507,1508..2218)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1547..1568)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1808..1827)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2155..2176
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    2711..2727
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2735..2741,2742..2759,2760..2765))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2780..2801
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2918..2935)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3089..3677)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3169..3188)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3841..4632,4633..4701))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4464..4483
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4702..4806)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        4963..5266
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5267..5470
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5421..5441
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
ORIGIN
        1 cgtccgggat cgggagaccc ctgcccaggg accaccgacc caccaccggg aggtaagctg
       61 gccagcaact tatctgtgtc tgtccgattg tctagtgtct atgactgatt ttatgcgcct
      121 gcgtcggtac tagttagcta actagctctg tatctggcgg acccgtggtg gaactgacga
      181 gttcggaaca cccggccgca accctgggag acgtcccagg gacttcgggg gccgtttttg
      241 tggcccgacc tgagtccaaa aatcccgatc gttttggact ctttggtgca ccccccttag
      301 aggagggata tgtggttctg gtaggagacg agaacctaaa acagttcccg cctccgtctg
      361 aatttttgct ttcggtttgg gaccgaagcc gcgccgcgcg tcttgtctgc tgcagcatcg
      421 ttctgtgttg tctctgtctg actgtgtttc tgtatttgtc tgaaaatatg ggcccgggcc
      481 agactgttac cactccctta agtttgacct taggtcactg gaaagatgtc gagcggatcg
      541 ctcacaacca gtcggtagat gtcaagaaga gacgttgggt taccttctgc tctgcagaat
      601 ggccaacctt taacgtcgga tggccgcgag acggcacctt taaccgagac ctcatcaccc
      661 aggttaagat caaggtcttt tcacctggcc cgcatggaca cccagaccag gtcccctaca
      721 tcgtgacctg ggaagccttg gcttttgacc cccctccctg ggtcaagccc tttgtacacc
      781 ctaagcctcc gcctcctctt cctccatccg ccccgtctct cccccttgaa cctcctcgtt
      841 cgaccccgcc tcgatcctcc ctttatccag ccctcactcc ttctctaggc gcccccatat
      901 ggccatatga gatcttatat ggggcacccc cgccccttgt aaacttccct gaccctgaca
      961 tgacaagagt tactaacagc ccctctctcc aagctcactt acaggctctc tacttagtcc
     1021 agcacgaagt ctggagacct ctggcggcag cctaccaaga acaactggac cgaccggtgg
     1081 tacctcaccc ttaccgagtc ggcgacacag tgtgggtccg ccgacaccag actaagaacc
     1141 tagaacctcg ctggaaagga ccttacacag tcctgctgac cacccccacc gccctcaaag
     1201 tagacggcat cgcagcttgg atacacgccg cccacgtgaa ggctgccgac cccgggggtg
     1261 gaccatcctc tagactgccg gatccaagct ggaggaaaaa ctgtttcata cagaaggcgt
     1321 ggaggaaaaa ctgtttcata cagaaggcgt ggaggaaaaa ctgtttcata cagaaggcgt
     1381 ggaggaaaaa ctgtttcata cagaaggcgt cgcgaattcg cggagactct agagggtata
     1441 taatggaagc tcgatttcca gcttggcatt ccggtactgt tggtaaacac caagcttcac
     1501 catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga
     1561 cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta
     1621 cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac
     1681 cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa
     1741 gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt
     1801 cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct
     1861 ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca
     1921 caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa
     1981 cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc
     2041 cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     2101 ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt
     2161 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta
     2221 aagcggccgc cagcacagtg gtcgatcgac gataaaataa aagattttat ttagtctcca
     2281 gaaaaagggg ggaatgaaag accccacctg taggtttggc aagctagctt aagtaacgcc
     2341 attttgcaag gcatggaaaa atacataact gagaatagaa aagttcagat caaggtcagg
     2401 aacagatgga acagggtcgc gtcccgcaat aaaagagccc acaacccctc actcggggcg
     2461 ccagtcctcc gattgactga gtcgcccggg tacccgtgta tccaataaac cctcttgcag
     2521 ttgcatccga cttgtggtct cgctgttcct tgggagggtc tcctctgagt gattgactac
     2581 ccgtcagcgg gggtctttca catgcagcat gtatcaaaat taatttggtt ttttttctta
     2641 agtatttaca ttaaatggcc atagtagttc attatggaca gcgcagaaag agctggggag
     2701 aattgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     2761 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     2821 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     2881 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     2941 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     3001 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     3061 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     3121 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     3181 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3241 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3301 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3361 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3421 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3481 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3541 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3601 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3661 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     3721 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     3781 tccttttgcg gccggccgca aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3841 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3901 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     3961 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4021 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     4081 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     4141 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     4201 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     4261 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     4321 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     4381 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     4441 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     4501 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     4561 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     4621 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4681 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4741 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4801 tccgcgcaca tttccccgaa aagtgccacc agctttgctc ttaggagttt cctaatacat
     4861 cccaaactca aatatataaa gcatttgact tgttctatgc cctagttatt aatagtaatc
     4921 aattacgggg tcattagttc atagcccata tatggagttc cgcgttacat aacttacggt
     4981 aaatggcccg cctggctgac cgcccaacga cccccgccca ttgacgtcaa taatgacgta
     5041 tgttcccata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
     5101 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
     5161 cgtcaatgac ggtaaatggc ccgcctggca ttatgcccag tacatgacct tatgggactt
     5221 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtga tgcggttttg
     5281 gcagtacatc aatgggcgtg gatagcggtt tgactcacgg ggatttccaa gtctccaccc
     5341 cattgacgtc aatgggagtt tgttttggca ccaaaatcaa cgggactttc caaaatgtcg
     5401 taacaactcc gccccattga cgcaaatggg cggtaggcgt gtacggtggg aggtctatat
     5461 aagcagagct caataaaaga gcccacaacc cctcactcgg cgcgccagtc ctccgattga
     5521 ctgagtcgcc cgggtacccg tgtatccaat aaaccctctt gcagttgcat ccgacttgtg
     5581 gtctcgctgt tccttgggag ggtctcctct gagtgattga ctacccgtca gcgggggtct
     5641 ttcatttggg ggct
//
