LOCUS       sequence-351723-        7369 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of G-alpha-q(L254A) fused to 
            mNeonGreen-eDHFR(69K6).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7369)
  AUTHORS   Suzuki S, Nakamura A, Hatano Y, Yoshikawa M, Yoshii T, Sawada S, 
            Atsuta-Tsunoda K, Aoki K, Tsukiji S
  TITLE     A chemogenetic platform for controlling plasma membrane signaling 
            and synthetic signal oscillation.
  JOURNAL   Cell Chem Biol. 2022 Sep 15;29(9):1446-1464.e10. doi: 
            10.1016/j.chembiol.2022.06.005. Epub 2022 Jul 13.
  PUBMED    35835118
REFERENCE   2  (bases 1 to 7369)
  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.chembiol.2022.06.005"; journalName: "Cell Chem Biol"; 
            date: "2022-09-15- 15"; volume: "29"; issue: "9"; pages: 
            "1446-1464.e10"
COMMENT     Sequence Label: pCAGGS-mNG-eDHFR(69K6)-G-alpha-q(L254A)
FEATURES             Location/Qualifiers
     source          1..7369
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          121..1137
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1061..1083
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1145..1164
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1192..1201
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1198..1905
                     /codon_start=1
                     /product="bright monomeric yellow-green fluorescent protein
                     derived from LanYFP (Shaner et al., 2013)"
                     /label=mNeonGreen
                     /translation="MVSKGEEDNMASLPATHELHIFGSINGVDFDMVGQGTGNPNDGYE
                     ELNLKSTKGDLQFSPWILVPHIGYGFHQYLPYPDGMSPFQAAMVDGSGYQVHRTMQFED
                     GASLTVNYRYTYEGSHIKGEAQVKGTGFPADGPVMTNSLTAADWCRSKKTYPNDKTIIS
                     TFKWSYTTGNGKRYRSTARTTYTFAKPMAANYLKNQPMYVFRKTELKHSKTELNFKEWQ
                     KAFTDVMGMDELYK"
     regulatory      1453..1462
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(3731..3751)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3731..3749)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3770..3786)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3888..3907)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3953..4008
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4007..4026)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(4369..4385)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     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."
     promoter        4542..4737
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4588..4723
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4743..4877
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4780..4799)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4834..4853
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4945..4962)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5116..5704)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5196..5215)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5875..6666,6667..6735))
                     /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     6498..6517
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6736..6840)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        6871..7250
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt
       61 ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg
      121 ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg
      181 ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg cccttctcct
      241 ccgggctgta attagcgctt ggtttaatga cggcttgttt cttttctgtg gctgcgtgaa
      301 agccttgagg ggctccggga gggccctttg tgcgggggga gcggctcggg gggtgcgtgc
      361 gtgtgtgtgt gcgtggggag cgccgcgtgc ggctccgcgc tgcccggcgg ctgtgagcgc
      421 tgcgggcgcg gcgcggggct ttgtgcgctc cgcagtgtgc gcgaggggag cgcggccggg
      481 ggcggtgccc cgcggtgcgg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg
      541 tgcgtggggg ggtgagcagg gggtgtgggc gcgtcggtcg ggctgcaacc ccccctgcac
      601 ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg tacggggcgt
      661 ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg
      721 gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc
      781 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg
      841 cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc
      901 ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag
      961 ggccttcgtg cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg
     1021 cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg
     1081 accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc
     1141 ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatt ggccaccatg
     1201 gtgagcaagg gcgaggagga taacatggcc tctctcccag cgacacatga gttacacatc
     1261 tttggctcca tcaacggtgt ggactttgac atggtgggtc agggcaccgg caatccaaat
     1321 gatggttatg aggagttaaa cctgaagtcc accaagggtg acctccagtt ctccccctgg
     1381 attctggtcc ctcatatcgg gtatggcttc catcagtacc tgccctaccc tgacgggatg
     1441 tcgcctttcc aggccgccat ggtagatggc tccggctacc aagtccatcg cacaatgcag
     1501 tttgaagatg gtgcctccct tactgttaac taccgctaca cctacgaggg aagccacatc
     1561 aaaggagagg cccaggtgaa ggggactggt ttccctgctg acggtcctgt gatgaccaac
     1621 tcgctgaccg ctgcggactg gtgcaggtcg aagaagactt accccaacga caaaaccatc
     1681 atcagtacct ttaagtggag ttacaccact ggaaatggca agcgctaccg gagcactgcg
     1741 cggaccacct acacctttgc caagccaatg gcggctaact atctgaagaa ccagccgatg
     1801 tacgtgttcc gtaagacgga gctcaagcac tccaagaccg agctcaactt caaggagtgg
     1861 caaaaggcct ttaccgatgt gatgggcatg gacgagctct acaagtctgg gtccgcagcc
     1921 agtagcgccg gaagtatgat cagtctgatt gcggcgttag cggtagatcg cgttatcggc
     1981 atggaaaacg ccatgccgtg gaacctgcct gccgatctcg cctggtttaa acgcaacacc
     2041 ttaaataaac ccgtgattat gggccgccat acctgggaat caatcggtcg tccgttgcca
     2101 ggacgcaaaa atattatcct cagcagtcaa ccgggtacgg acaaaaaaaa gaaaaagaaa
     2161 gatcgcgtaa cgtgggtgaa gtcggtggat gaagccatcg cggcgtgtgg tgacgtacca
     2221 gaaatcatgg tgattggcgg cggtcgcgtt tatgaacagt tcttgccaaa agcgcaaaaa
     2281 ctgtatctga cgcatatcga cgcagaagtg gaaggcgaca cccatttccc ggattacgag
     2341 ccggatgact gggaatcggt attcagcgaa ttccacgatg ctgatgcgca gaactctcac
     2401 agctattgct ttgagattct ggagcggcgg ctgtactctg gtggcggagg ctcgggcgga
     2461 ggtgggtcgg gtggcggcgg atcttcagct ggagcttcgt tgtacaaggc aagaggagcc
     2521 gctgcgggtg ccggaggcgc tggtaggagc ggcaaaaagg ggaagaaagg gaaaaagggc
     2581 accacaatga ctctggagtc catcatggca tcctccctga gcgaggaggc caaggaagcc
     2641 cggaggatca acgacgagat cgagcggcag ctgcgcaggg acaagcgcga cgcccgccgg
     2701 gagctcaagc tgctgctgct ggggacaggg gagagtggca agtcgacctt catcaagcag
     2761 atgaggatca tccacgggtc gggctactct gacgaagaca agcgcggctt caccaagctg
     2821 gtgtatcaga acatcttcac ggccatgcag gccatgatca gagcgatgga cacactcaag
     2881 atcccataca agtatgaaca caataaggct catgcacaat tggttcgaga ggttgatgtg
     2941 gagaaggtgt ctgcttttga gaatccatat gtagatgcaa taaagagctt gtggaatgat
     3001 cctggaatcc aggagtgcta cgacagacga cgggaatatc agttatctga ctctaccaaa
     3061 tactatctga atgacttgga ccgtgtagcc gacccttcct atctgcctac acaacaagac
     3121 gtgcttagag ttcgagtccc cactacaggg atcatcgaat acccctttga cttacaaagt
     3181 gtcattttca gaatggtcga tgtagggggc ctgaggtcag agagaagaaa atggatacac
     3241 tgctttgaaa atgtcacctc catcatgttt ctagtagcgc ttagcgaata tgatcaagtt
     3301 cttgtggagt cagacaatga gaaccgcatg gaggagagca aagcagcctt tagaacaatt
     3361 atcacctacc cctggttcca gaactcctct gtgattctgt tcttaaacaa gaaagatctt
     3421 ctagaggaga aaatcatgta ttcccaccta gtcgactact tcccagaata tgatggaccc
     3481 cagagagatg cccaggcagc tcgagaattc atcctgaaaa tgttcgtgga cctgaacccc
     3541 gacagtgaca aaatcatcta ctcccacttc acgtgcgcca cagataccga gaacatccgc
     3601 ttcgtctttg cagccgtcaa ggacaccatc ctgcagctga acctgaagga gtacaatctg
     3661 gtctaacagt tgctcgagag tacccgggat actcgcggcc gccaccgcgg tggagctcca
     3721 gcttttgttc cctttagtga gggttaattg cgcgcttggc gtaatcatgg tcatagctgt
     3781 ttcctgtgtg aaattgttat ccgctaattc actcctcagg tgcaggctgc ctatcagaag
     3841 gtggtggctg gtgtggccaa tgccctggct cacaaatacc actgagatct ttttccctct
     3901 gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg ctaataaagg
     3961 aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact cggaaggaca
     4021 tatgggaggg caaatcattt aaaacatcag aatgagtatt tggtttagag tttggcaaca
     4081 tatgcccata tgctggctgc catgaacaaa ggttggctat aaagaggtca tcagtatatg
     4141 aaacagcccc ctgctgtcca ttccttattc catagaaaag ccttgacttg aggttagatt
     4201 ttttttatat tttgttttgt gttatttttt tctttaacat ccctaaaatt ttccttacat
     4261 gttttactag ccagattttt cctcctctcc tgactactcc cagtcatagc tgtccctctt
     4321 ctcttatgga gatccctcga cctgcagccc aagcttggcg taatcatggt catagctgtt
     4381 tcctgtgtga aattgttatc cgctcacaat tccacacaac atacgagccg gaagcataaa
     4441 gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca ttaattgcgt tgcgctcact
     4501 gcccgctttc cagtcgggaa acctgtcgtg ccagcggatc cgcatctcaa ttagtcagca
     4561 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     4621 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctcggcc
     4681 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     4741 ctaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca
     4801 caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat
     4861 cttatcatgt ctggatccgc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     4921 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     4981 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     5041 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     5101 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     5161 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     5221 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     5281 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     5341 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     5401 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     5461 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     5521 cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct
     5581 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     5641 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     5701 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     5761 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     5821 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     5881 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     5941 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     6001 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     6061 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     6121 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     6181 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     6241 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     6301 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     6361 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     6421 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     6481 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     6541 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     6601 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     6661 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     6721 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     6781 tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg
     6841 cacatttccc cgaaaagtgc cacctgggtc gacattgatt attgactagt tattaatagt
     6901 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
     6961 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
     7021 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
     7081 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt acgcccccta
     7141 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttatggg
     7201 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtcgaggtga
     7261 gccccacgtt ctgcttcact ctccccatct cccccccctc cccaccccca attttgtatt
     7321 tatttatttt ttaattattt tgtgcagcga tgggggcggg ggggggggg
//
