LOCUS       sequence-325857-        7044 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Used to provide a steady-state readout of cPLA2 activation in live 
            cells. Amino-terminally fused to mKate2 for red fluorescence..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7044)
  AUTHORS   Enyedi B, Kala S, Nikolich-Zugich T, Niethammer P
  TITLE     Tissue damage detection by osmotic surveillance.
  JOURNAL   Nat Cell Biol. 2013 Sep;15(9):1123-30. doi: 10.1038/ncb2818. Epub 
            2013 Aug 11.
  PUBMED    23934216
REFERENCE   2  (bases 1 to 7044)
  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"; doi: "10.1038/ncb2818"; 
            journalName: "Nat Cell Biol"; date: "2013-09"; volume: "15"; issue: 
            "9"; pages: "1123-30"
COMMENT     Sequence Label: pCS2+cPLA2-mKate2
FEATURES             Location/Qualifiers
     source          1..7044
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        10..987
                     /label=CMV IE94 promoter
                     /note="enhancer/promoter region of simian cytomegalovirus 
                     major immediate early transcription unit IE94"
     promoter        1022..1040
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     1022..1039
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             3335..4045
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKAVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRAD
                     MALKLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKL"
     CDS             3335..4045
                     /codon_start=1
                     /product="monomeric photoactivatable red fluorescent 
                     protein (Gunewardene et al., 2011)"
                     /label=PAmKate
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKAVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRAD
                     MALKLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKL"
     polyA_signal    4083..4217
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4107..4126)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4161..4180
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     complement(4330..4350)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4330..4348)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4369..4385)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4369..4385)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4382..4404)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4393..4409
                     /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(4417..4423,4424..4441,4442..4447))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4462..4483
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4600..4617)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4771..5359)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4851..4870)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5530..6321,6322..6390))
                     /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     6153..6172
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6391..6495)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6521..6976)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6658..6679)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6870..6889
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cagtcgaccg ccaattcaat atggcgtata tggactcatg ccaattcaat atggtggatc
       61 tggacctgtg ccaattcaat atggcgtata tggactcgtg ccaattcaat atggtggatc
      121 tggaccccag ccaattcaat atggcggact tggcaccatg ccaattcaat atggcggacc
      181 tggcactgtg ccaactgggg aggggtctac ttggcacggt gccaagtttg aggaggggtc
      241 ttggccctgt gccaagtccg ccatattgaa ttggcatggt gccaataatg gcggccatat
      301 tggctatatg ccaggatcaa tatataggca atatccaata tggccctatg ccaatatggc
      361 tattggccag gttcaatact atgtattggc cctatgccat atagtattcc atatatgggt
      421 tttcctattg acgtagatag cccctcccaa tgggcggtcc catataccat atatggggct
      481 tcctaatacc gcccatagcc actcccccat tgacgtcaat ggtctctata tatggtcttt
      541 cctattgacg tcatatgggc ggtcctattg acgtatatgg cgcctccccc attgacgtca
      601 attacggtaa atggcccgcc tggctcaatg cccattgacg tcaataggac cacccaccat
      661 tgacgtcaat gggatggctc attgcccatt catatccgtt ctcacgcccc ctattgacgt
      721 caatgacggt aaatggccca cttggcagta catcaatatc tattaatagt aacttggcaa
      781 gtacattact attggaagta cgccagggta cattggcagt actcccattg acgtcaatgg
      841 cggtaaatgg cccgcgatgg ctgccaagta catccccatt gacgtcaatg gggaggggca
      901 atgacgcaaa tgggcgttcc attgacgtaa atgggcggta ggcgtgccta atgggaggtc
      961 tatataagca atgctcgttt agggaaccgc cattctgcct ggggacgtcg gagcaagctt
     1021 gatttaggtg acactataga atacaagcta cttgttcttt ttgcaggatc ccatcgatgc
     1081 cacaatgtcc aacattatag ttgagcatca gtattctcat agactgaaat tgactgtggt
     1141 gagagctgaa aatgtaacta aaggagcatt tggggattta ttggacaccc cggaccctta
     1201 tgttgagctg tcagtgccaa ccacaccaga gtcaagaaaa cgaacccgcc acattaacaa
     1261 tgacattaac cccaagtgga acgagacctt tgagttcata ttggatccca atcaatccaa
     1321 tgttttggag gtgacactga tggatgccaa ttatgtcatg gatgaaactc ttggcacagc
     1381 aaaatattcc ctctcaaaac tcaaagtggc acaaatggag catgtgaccc tctctatcgg
     1441 caagacaacc aaagtgttcc tcgacttgct cttggaagtt tgcgcatcca cagacctgcg
     1501 cttcagcatg acactgtgtg atcaagagaa gctgttcatg cagacacgca gagacagggt
     1561 catgctcagc ataaagaagc ttcttaaaat ggaaaaccct cgtttccttc catcctcacc
     1621 aagagaggtt cccaccatcg ctattctggg ctctggagga ggttttcgtg caatggtggg
     1681 cttctctggt gtgatgaagg ctttgtatga atctggcgtg tttgactgtg caacatatgt
     1741 ggcaggactt tcaggatcaa catggtatat gtctatgctg tactctcatc cagagtttcc
     1801 tgctaaaggc ccaggcgaca ttaataaaga gctcatgaac agagtcagta ataacccact
     1861 caaactgctc ctgccccaaa acatcaaccg ctacgtcaaa gcactgtgga agaagaaatc
     1921 tgctggacag ccggtcacct tcactgacat cttcgggatg ttgataggag agacgcttat
     1981 tccagggaga atgaacatta agctgagctc attgaaaggg aaaattaatg aaggtcagag
     2041 ccctcttcct ctcttcacct gtctgcacgt caaacctgac gtctctgagc tcatgtttgc
     2101 agattgggtg gagttcagcc cttatgaaat tggcatggcc aaatatggca ccttcatgtc
     2161 tcctgggctg tttggcagca aattcttcat gggaagtgtt gtgaaacaat atgaggaaaa
     2221 ccccttacat ttcctgatgg gtgtgtgggg aagtgctttt tccatcctct ttaaccgggt
     2281 tttgggtgtg aaggagacga ccagcagtag cactatggag gaggagctgg agcaaattaa
     2341 accagagcac attgtgggtg acgattctgc tgataacgag gaggaaactc aaagaggagg
     2401 cacagagtct gcagatgcag aggacgagcg ccagcgtcat gcccaggcca gctgggtaca
     2461 gcgcatgttg acctccatca tgggcgacac cactctgttc accactcgtg agggccgcgc
     2521 aggcaaagtg cacaacttca tgctgggact gaacctgaac tcaaccctgc ccttctctcc
     2581 cttcagcggc atcacacacc agacctcgct ggaggaggaa gtggacgctg ttactgatcc
     2641 ggatgagttt gagcgaatct atgagcctct ggatgtgaag agcaagaaaa tccatgtggt
     2701 ggacagtggc ttgaccttta acctgcccta ccctctcatt ctgcggtgtc agagaggagt
     2761 cgacctgatt atttcattcg acttctcagc ccgacccagt gactccagcc caccgttcaa
     2821 agagttactg ctggctgaaa aatgggcccg tatgaataag ctgcccttcc cgaagattga
     2881 ttcgaaagta ttcgacagag agggtctaaa agagtgctat gtgttcaagc ctgctaaagg
     2941 agacaagaac tgccccacca tcattcattt tgtcttggcc aacatcaact tccggaattt
     3001 taaagcccct ggagttcctc gagattcaga caaggacata gagtttggtg actttgacat
     3061 atttgacgag ccagcctccc cttactccac tttcaacttc aaatacaaca accaggcctt
     3121 caagagactt catgacctca tggagttcaa tacgcttaac aacatcgagg ttatcaaaga
     3181 agccattaaa gacagcattc tgctgagacg agagaaccca gcgcgatgtt cggtttccct
     3241 ttcgctcagt gaaatcgaaa acaagaagtt cctgaaacga gacaacagca ttgcaaaacg
     3301 acccaccgga gctggaggaa ttcgtcctgg atctatggtg tctaagggcg aagagctgat
     3361 taaggagaac atgcacatga agctgtacat ggagggcacc gtgaacaacc accacttcaa
     3421 gtgcacatcc gagggcgaag gcaagcccta cgagggcacc cagaccatga gaatcaaggc
     3481 cgtcgagggc ggccctctcc ccttcgcctt cgacatcctg gctaccagct tcatgtacgg
     3541 cagcaaaacc ttcatcaacc acacccaggg catccccgac ttctttaagc agtccttccc
     3601 tgagggcttc acatgggaga gagtcaccac atacgaagac gggggcgtgc tgaccgctac
     3661 ccaggacacc agcctccagg acggctgcct catctacaac gtcaagatca gaggggtgaa
     3721 cttcccatcc aacggccctg tgatgcagaa gaaaacactc ggctgggagg cctccaccga
     3781 gaccctgtac cccgctgacg gcggcctgga aggcagagcc gacatggccc tgaagctcgt
     3841 gggcgggggc cacctgatct gcaacttgaa gaccacatac agatccaaga aacccgctaa
     3901 gaacctcaag atgcccggcg tctactatgt ggacagaaga ctggaaagaa tcaaggaggc
     3961 cgacaaagag acctacgtcg agcagcacga ggtggctgtg gccagatact gcgacctccc
     4021 tagcaaactg gggcacaaac tgtaactcga gcctctagaa ctatagtgag tcgtattacg
     4081 tagatccaga catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt
     4141 gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa
     4201 gctgcaataa acaagttaac aacaacaatt gcattcattt tatgtttcag gttcaggggg
     4261 aggtgtggga ggttttttaa ttcgcggccg cggcgccaat gcattgggcc cggtacccag
     4321 cttttgttcc ctttagtgag ggttaattgc gcgcttggcg taatcatggt catagctgtt
     4381 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg ggagcataaa
     4441 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4501 gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     4561 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     4621 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     4681 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     4741 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     4801 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     4861 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     4921 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     4981 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     5041 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     5101 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     5161 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     5221 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     5281 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     5341 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     5401 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     5461 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     5521 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     5581 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     5641 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     5701 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     5761 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     5821 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     5881 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     5941 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6001 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6061 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6121 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6181 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     6241 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     6301 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     6361 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     6421 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     6481 aataggggtt ccgcgcacat ttccccgaaa agtgccacct aaattgtaag cgttaatatt
     6541 ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat tttttaacca ataggccgaa
     6601 atcggcaaaa tcccttataa atcaaaagaa tagaccgaga tagggttgag tgttgttcca
     6661 gtttggaaca agagtccact attaaagaac gtggactcca acgtcaaagg gcgaaaaacc
     6721 gtctatcagg gcgatggccc actacgtgaa ccatcaccct aatcaagttt tttggggtcg
     6781 aggtgccgta aagcactaaa tcggaaccct aaagggagcc cccgatttag agcttgacgg
     6841 ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag cgaaaggagc gggcgctagg
     6901 gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca cacccgccgc gcttaatgcg
     6961 ccgctacagg gcgcgtccca ttcgccattc aggctgcgca actgttggga agggcgatcg
     7021 gtgcgggcct cttcgctatt acgc
//
