LOCUS       sequence-223071-        5655 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 5655)
  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 5655)
  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..5655
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     323..342
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(561..665)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        822..1127
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1128..1331
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1282..1302
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    1570..1927
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1992..2408
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     primer_bind     2338..2360
                     /label=pLXSN 5'
                     /note="Murine stem cell virus, forward primer. Also called 
                     MSCV"
     primer_bind     2376..2392
                     /label=pBABE 5'
                     /note="Psi packaging signal, forward primer for pBABE 
                     vectors"
     misc_feature    2418..2792
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     primer_bind     2730..2746
                     /label=pBMN 5'
                     /note="MMLV sequence, for inserts in pBMN retroviral 
                     vector"
     protein_bind    2806..2835
                     /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    2836..2865
                     /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    2866..2895
                     /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    2896..2925
                     /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        2945..2976
                     /label=minP
                     /note="minimal TATA-box promoter with low basal activity"
     CDS             join(3017..3019,3020..3022,3023..3736)
                     /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(3017..3019,3020..3022,3023..3733)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3062..3083)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3323..3342)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3670..3691
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    4225..4241
                     /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(4249..4255,4256..4273,4274..4279))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4294..4315
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4432..4449)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4603..5191)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4683..4702)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5355..5655,1..491,492..560))
                     /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"
ORIGIN
        1 gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat ggcttcattc
       61 agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg caaaaaagcg
      121 gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt gttatcactc
      181 atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag atgcttttct
      241 gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg accgagttgc
      301 tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt aaaagtgctc
      361 atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct gttgagatcc
      421 agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac tttcaccagc
      481 gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat aagggcgaca
      541 cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat ttatcagggt
      601 tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca aataggggtt
      661 ccgcgcacat ttccccgaaa agtgccacca gctttgctct taggagtttc ctaatacatc
      721 ccaaactcaa atatataaag catttgactt gttctatgcc ctagttatta atagtaatca
      781 attacggggt cattagttca tagcccatat atggagttcc gcgttacata acttacggta
      841 aatggcccgc ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat
      901 gttcccatag taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg
      961 taaactgccc acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac
     1021 gtcaatgacg gtaaatggcc cgcctggcat tatgccccag tacatgacct ttatgggact
     1081 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     1141 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     1201 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     1261 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     1321 taagcagagc tcaataaaag agcccacaac ccctcactcg gcgcgccagt cctccgattg
     1381 actgagtcgc ccgggtaccc gtgtatccaa taaaccctct tgcagttgca tccgacttgt
     1441 ggtctcgctg ttccttggga gggtctcctc tgagtgattg actacccgtc agcgggggtc
     1501 tttcatttgg gggctcgtcc gggatcggga gacccctgcc cagggaccac cgacccacca
     1561 ccgggaggta agctggccag caacttatct gtgtctgtcc gattgtctag tgtctatgac
     1621 tgattttatg cgcctgcgtc ggtactagtt agctaactag ctctgtatct ggcggacccg
     1681 tggtggaact gacgagttcg gaacacccgg ccgcaaccct gggagacgtc ccagggactt
     1741 cgggggccgt ttttgtggcc cgacctgagt ccaaaaatcc cgatcgtttt ggactctttg
     1801 gtgcaccccc cttagaggag ggatatgtgg ttctggtagg agacgagaac ctaaaacagt
     1861 tcccgcctcc gtctgaattt ttgctttcgg tttgggaccg aagccgcgcc gcgcgtcttg
     1921 tctgctgcag catcgttctg tgttgtctct gtctgactgt gtttctgtat ttgtctgaaa
     1981 atatgggccc gggccagact gttaccactc ccttaagttt gaccttaggt cactggaaag
     2041 atgtcgagcg gatcgctcac aaccagtcgg tagatgtcaa gaagagacgt tgggttacct
     2101 tctgctctgc agaatggcca acctttaacg tcggatggcc gcgagacggc acctttaacc
     2161 gagacctcat cacccaggtt aagatcaagg tcttttcacc tggcccgcat ggacacccag
     2221 accaggtccc ctacatcgtg acctgggaag ccttggcttt tgacccccct ccctgggtca
     2281 agccctttgt acaccctaag cctccgcctc ctcttcctcc atccgccccg tctctccccc
     2341 ttgaacctcc tcgttcgacc ccgcctcgat cctcccttta tccagccctc actccttctc
     2401 taggcgcccc catatggcca tatgagatct tatatggggc acccccgccc cttgtaaact
     2461 tccctgaccc tgacatgaca agagttacta acagcccctc tctccaagct cacttacagg
     2521 ctctctactt agtccagcac gaagtctgga gacctctggc ggcagcctac caagaacaac
     2581 tggaccgacc ggtggtacct cacccttacc gagtcggcga cacagtgtgg gtccgccgac
     2641 accagactaa gaacctagaa cctcgctgga aaggacctta cacagtcctg ctgaccaccc
     2701 ccaccgccct caaagtagac ggcatcgcag cttggataca cgccgcccac gtgaaggctg
     2761 ccgaccccgg gggtggacca tcctctagac tgccggatcc aagctggagg aaaaactgtt
     2821 tcatacagaa ggcgtggagg aaaaactgtt tcatacagaa ggcgtggagg aaaaactgtt
     2881 tcatacagaa ggcgtggagg aaaaactgtt tcatacagaa ggcgtcgcga attcgcggag
     2941 actctagagg gtatataatg gaagctcgat ttccagcttg gcattccggt actgttggta
     3001 aacaccaagc ttcaccatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     3061 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
     3121 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     3181 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     3241 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
     3301 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     3361 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     3421 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
     3481 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
     3541 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
     3601 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
     3661 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
     3721 cgagctgtac aagtaaagcg gccgccagca cagtggtcga tcgacgataa ataaaagatt
     3781 ttatttagtc tccagaaaaa ggggggaatg aaagacccca cctgtaggtt tggcaagcta
     3841 gcttaagtaa cgccattttg caaggcatgg aaaaatacat aactgagaat agaaaagttc
     3901 agatcaaggt caggaacaga tggaacaggg tcgcgtcccg caataaaaga gcccacaacc
     3961 cctcactcgg ggcgccagtc ctccgattga ctgagtcgcc cgggtacccg tgtatccaat
     4021 aaaccctctt gcagttgcat ccgacttgtg gtctcgctgt tccttgggag ggtctcctct
     4081 gagtgattga ctacccgtca gcgggggtct ttcacatgca gcatgtatca aaattaattt
     4141 ggtttttttt cttaagtatt tacattaaat ggccatagta gttcattatg gacagcgcag
     4201 aaagagctgg ggagaattgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4261 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4321 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     4381 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     4441 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     4501 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     4561 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     4621 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     4681 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     4741 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     4801 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     4861 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     4921 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     4981 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     5041 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     5101 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     5161 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     5221 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     5281 gatcttcacc tagatccttt tgcggccggc cgcaaatcaa tctaaagtat atatgagtaa
     5341 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5401 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5461 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5521 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5581 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5641 aatagtttgc gcaac
//
