LOCUS       sequence-259088-        6706 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression of Lyn-SponGee, a lipid raft-targeted variant 
            of the cGMP buffer SponGee that prevents downstream signaling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6706)
  AUTHORS   Ros O, Zagar Y, Ribes S, Baudet S, Loulier K, Couvet S, Ladarre D, 
            Aghaie A, Louail A, Petit C, Mechulam Y, Lenkei Z, Nicol X
  TITLE     SponGee: A Genetic Tool for Subcellular and Cell-Specific cGMP 
            Manipulation.
  JOURNAL   Cell Rep. 2019 Jun 25;27(13):4003-4012.e6. doi: 
            10.1016/j.celrep.2019.05.102.
  PUBMED    31242429
REFERENCE   2  (bases 1 to 6706)
  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: 
            "10.1016/j.celrep.2019.05"; journalName: "Cell Rep"; date: 
            "2019-06-25- 25"; volume: "27"; issue: "13"; pages: "4003-4012.e6"
COMMENT     Sequence Label: pCX-Lyn-SponGee
FEATURES             Location/Qualifiers
     source          1..6706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1674
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1598..1620
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1682..1701
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1765..1774
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      1849..1858
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1858..1881
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      2932..2941
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2938..3612
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /note="mammalian codon-optimized"
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRPS
                     TGA"
     primer_bind     complement(3068..3086)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3336..3355
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     3530..3553
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(3730..3749)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3795..3850
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3849..3868)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4209..4225)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4209..4225)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4222..4244)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4233..4249
                     /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(4257..4263,4264..4281,4282..4287))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4302..4323
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4382..4577
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4428..4563
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4583..4717
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4620..4639)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4674..4693
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4785..4802)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4956..5544)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5036..5055)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5715..6506,6507..6575))
                     /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     6338..6357
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6576..6680)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtac ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gcccccggag cgccggcggc
     1321 tgtcgaggcg cggcgagccg cagccattgc cttttatggt aatcgtgcga gagggcgcag
     1381 ggacttcctt tgtcccaaat ctgtgcggag ccgaaatctg ggaggcgccg ccgcaccccc
     1441 tctagcgggc gcggggcgaa gcggtgcggc gccggcagga aggaaatggg cggggagggc
     1501 cttcgtgcgt cgccgcgccg ccgtcccctt ctccctctcc agcctcgggg ctgtccgcgg
     1561 ggggacggct gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc
     1621 ggcggctcta gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg
     1681 ggcaacgtgc tggttattgt gctgtctcat cattttggca aagaattctg atatctttaa
     1741 ttaaaccccg ggaccggtgc tagcgccacc atgggctgca tcaagagcaa gcgcaaggac
     1801 aagatgggct gcatcaagag caagcgcaag gacaaggcta gctccggagc caccatggac
     1861 tacaaagacg atgacgacaa gtccggaatc gatatgagcg agctggagga agactttgcc
     1921 aagattctca tgctcaagga ggagaggatc aaagagctgg agaagcggct gtcagagaag
     1981 gaggaagaaa tccaggagct gaagaggaaa ctccataaat gccagtcagt gctgcccgtg
     2041 ccctcgaccc acatcggccc ccggaccacc cgggcacagg gcatctcggc cgagccgcag
     2101 acctacaggt ccttccacga cctccgagtg accctgccct tctaccccaa gagtccacag
     2161 tccaagatcg atctcataaa ggaggccatc cttgacaatg actttatgaa gaacttggag
     2221 ctgtcacaga tccaagagat tgtggattgt atgtacccag tggagtacgg caaagacagc
     2281 tgcatcatca aagaaggaga tgtggggtca ctggtgtatg tcatggaaga tggtaaggtt
     2341 gaagttacaa aagaaggcgt gaagctgtgc acaatgggtc ctggtaaagt gtttggagag
     2401 ttggctatcc tttacaactg tacccggacg gcgaccgtca aaactcttgt aaatgtgaaa
     2461 ctctgggcca ttgatcgaca atgttttcag acgataatga tgaggacagg acttatcaag
     2521 cataccgagt atatggaatt tttaaaaagc gttccaacat tccagagcct tcctgaagag
     2581 atcctcagta aacttgctga cgtccttgaa gagacccact atgaaaatgg ggaatatatc
     2641 atcaggcaag gtgcaagagg ggacaccttc tttatcatca gtaaaggaaa ggttaatgtc
     2701 actcgtgaag actcgcccaa tgaagaccca gtctttctta gaaccttagg aaaaggagat
     2761 tggtttggag agaaagcctt gcagggggaa gatgtgagaa cagcgaatgt aattgcggca
     2821 gaagctgtaa cctgccttgt gatcgacaga gactctttca aacatttgat tggaggatta
     2881 gatgatgttt ctaataaagc atatgaagat gcagaagcta aggcggatcc cgccaccatg
     2941 gcctcctccg aggacgtcat caaggagttc atgcgcttca aggtgcgcat ggagggctcc
     3001 gtgaacggcc acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc
     3061 cagaccgcca agctgaaggt gaccaagggc ggccccctgc ccttcgcctg ggacatcctg
     3121 tcccctcagt tccagtacgg ctccaaggcc tacgtgaagc accccgccga catccccgac
     3181 tacttgaagc tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggat
     3241 ggcggcgtgg tgaccgtgac ccaggactcc tccctgcagg acggcgagtt catctacaag
     3301 gtgaagctgc gcggcaccaa cttcccctcc gacggccccg taatgcagaa gaagaccatg
     3361 ggctgggagg cctccaccga gcggatgtac cccgaggacg gcgccctgaa gggcgagatc
     3421 aagatgaggc tgaagctgaa ggacggcggc cactacgacg ccgaggtcaa gaccacctac
     3481 atggccaaga agcccgtgca gctgcccggc gcctacaaga ccgacatcaa gctggacatc
     3541 acctcccaca acgaggacta caccatcgtg gaacagtacg agcgcgccga gggccgcccc
     3601 tccaccggcg cctgtacaag taagaattct gcagataaac gctagcgaat tcactcctca
     3661 ggtgcaggct gcctatcaga aggtggtggc tggtgtggcc aatgccctgg ctcacaaata
     3721 ccactgagat ctttttccct ctgccaaaaa ttatggggac atcatgaagc cccttgagca
     3781 tctgacttct ggctaataaa ggaaatttat tttcattgca atagtgtgtt ggaatttttt
     3841 gtgtctctca ctcggaagga catatgggag ggcaaatcat ttaaaacatc agaatgagta
     3901 tttggtttag agtttggcaa catatgccat atgctggctg ccatgaacaa aggtggctat
     3961 aaagaggtca tcagtatatg aaacagcccc ctgctgtcca ttccttattc catagaaaag
     4021 ccttgacttg aggttagatt ttttttatat tttgttttgt gttatttttt tctttaacat
     4081 ccctaaaatt ttccttacat gttttactag ccagattttt cctcctctcc tgactactcc
     4141 cagtcatagc tgtccctctt ctcttatgaa gatccctcga cctgcagccc aagcttggcg
     4201 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4261 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4321 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagcggatc
     4381 cgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa
     4441 ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag
     4501 aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag
     4561 gcctaggctt ttgcaaaaag ctaacttgtt tattgcagct tataatggtt acaaataaag
     4621 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
     4681 gtccaaactc atcaatgtat cttatcatgt ctggatccgc tgcattaatg aatcggccaa
     4741 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4801 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4861 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4921 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4981 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5041 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5101 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca atgctcacgc
     5161 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     5221 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     5281 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     5341 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     5401 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     5461 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     5521 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     5581 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5641 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5701 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5761 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5821 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5881 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5941 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     6001 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     6061 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     6121 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     6181 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     6241 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     6301 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     6361 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     6421 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     6481 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     6541 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6601 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6661 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctg
//
