LOCUS       sequence-210153-        7161 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GsX chimera with G-alpha-i C-terminus (Pertussis toxin insensitive 
            mutant).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7161)
  AUTHORS   Ballister ER, Rodgers J, Martial F, Lucas RJ
  TITLE     A live cell assay of GPCR coupling allows identification of 
            optogenetic tools for controlling Go and Gi signaling.
  JOURNAL   BMC Biol. 2018 Jan 16;16(1):10. doi: 10.1186/s12915-017-0475-2.
  PUBMED    29338718
REFERENCE   2  (bases 1 to 7161)
  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: "BMC Biol.";
            date: "2018-01-16"; volume: "16(1):10. doi"; pages: " 
            10.1186/s12915-017-0475-2"
COMMENT     Sequence Label: pEM705 Gsi (Ser)
FEATURES             Location/Qualifiers
     source          1..7161
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     gap             424..439
                     /estimated_length=16
     intron          560..1576
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1500..1522
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1584..1603
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1646..1655
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(2959..2978)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3024..3079
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3078..3097)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     protein_bind    3443..3476
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    3489..3610
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3526..3545)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3580..3599
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3629..3649)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3629..3647)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3668..3684)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3668..3684)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3681..3703)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3692..3708
                     /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(3716..3722,3723..3740,3741..3746))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3761..3782
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3899..3916)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4070..4658)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4150..4169)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4829..5620,5621..5689))
                     /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     5452..5471
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5690..5794)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    5867..5915
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     protein_bind    6127..6160
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     CDS             6165..6764
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(6165..6184)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6661..6681
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(6900..6917)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6906..7017
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc
      361 cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg
      421 gggnnnnnnn nnnnnnnnng gcggggcgag gcggagaggt gcggcggcag ccaatcagag
      481 cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc cctataaaaa
      541 gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc cccgctccgc
      601 cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactcccac aggtgagcgg
      661 gcgggacggc ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac ggcttgtttc
      721 ttttctgtgg ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt gcggggggag
      781 cggctcgggg ggtgcgtgcg tgtgtgtgtg cgtggggagc gccgcgtgcg gctccgcgct
      841 gcccggcggc tgtgagcgct gcgggcgcgg cgcggggctt tgtgcgctcc gcagtgtgcg
      901 cgaggggagc gcggccgggg gcggtgcccc gcggtgcggg gggggctgcg aggggaacaa
      961 aggctgcgtg cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg cgtcggtcgg
     1021 gctgcaaccc cccctgcacc cccctccccg agttgctgag cacggcccgg cttcgggtgc
     1081 ggggctccgt acggggcgtg gcgcggggct cgccgtgccg ggcggggggt ggcggcaggt
     1141 gggggtgccg ggcggggcgg ggccgcctcg ggccggggag ggctcggggg aggggcgcgg
     1201 cggcccccgg agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt gccttttatg
     1261 gtaatcgtgc gagagggcgc agggacttcc tttgtcccaa atctgtgcgg agccgaaatc
     1321 tgggaggcgc cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg gcgccggcag
     1381 gaaggaaatg ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc ttctccctct
     1441 ccagcctcgg ggctgtccgc ggggggacgg ctgccttcgg gggggacggg gcagggcggg
     1501 gttcggcttc tggcgtgtga ccggcggctc tagagcctct gctaaccatg ttcatgcctt
     1561 cttctttttc ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg
     1621 caaagaattg ctcgaccacc ggcgcgccac catgggctgc ctcgggaaca gtaagaccga
     1681 ggaccagcgc aacgaggaga aggcgcagcg tgaggccaac aaaaagatcg agaagcagct
     1741 gcagaaggac aagcaggtct accgggccac gcaccgcctg ctgctgctgg gtgctggaga
     1801 atctggtaaa agcaccattg tgaagcagat gaggatcctg catgttaatg ggtttaatgg
     1861 agagggcggc gaagaggacc cgcaggctgc aaggagcaac agcgatggtg agaaggcaac
     1921 caaagtgcag gacatcaaaa acaacctgaa agaggcgatt gaaaccattg tggccgccat
     1981 gagcaacctg gtgccccccg tggagctggc caaccccgag aaccagttca gagtggacta
     2041 cattctgagt gtgatgaacg tgcctgactt tgacttccct cccgaattct atgagcatgc
     2101 caaggctctg tgggaggatg aaggagtgcg tgcctgctac gaacgctcca acgagtacca
     2161 gctgattgac tgtgcccagt acttcctgga caagatcgac gtgatcaagc aggctgacta
     2221 tgtgccgagc gatcaggacc tgcttcgctg ccgtgtcctg acttctggaa tctttgagac
     2281 caagttccag gtggacaaag tcaacttcca catgtttgac gtgggtggcc agcgcgatga
     2341 acgccgcaag tggatccagt gcttcaacga tgtgactgcc atcatcttcg tggtggccag
     2401 cagcagctac aacatggtca tccgggagga caaccagacc aaccgcctgc aggaggctct
     2461 gaacctcttc aagagcatct ggaacaacag atggctgcgc accatctctg tgatcctgtt
     2521 cctcaacaag caagatctgc tcgctgagaa agtccttgct gggaaatcga agattgagga
     2581 ctactttcca gaatttgctc gctacactac tcctgaggat gctactcccg agcccggaga
     2641 ggacccacgc gtgacccggg ccaagtactt cattcgagat gagtttctga ggatcagcac
     2701 tgccagtgga gatgggcgtc actactgcta ccctcatttc acctgcgctg tggacactga
     2761 gaacatccgc cgtgtgttca acgactgccg tgatgtcatc ataaaaaata atctaaaaga
     2821 ttctggtctc ttttgaaccg gtctgcagcc taggcttaag cgctcgaggc ggccgcaatt
     2881 cactcctcag gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc
     2941 tcacaaatac cactgagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc
     3001 ccttgagcat ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg
     3061 gaattttttg tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca
     3121 gaatgagtat ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa
     3181 aggttggcta taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt
     3241 ccatagaaaa gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt
     3301 ttctttaaca tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc
     3361 ctgactactc ccagtcatag ctgtccctct tctcttatgg agatccctcg acctgcaccg
     3421 tcgagctagc atatggatcc atataacttc gtataatgta tgctatacga agttattacg
     3481 ataccgtcaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     3541 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     3601 atgtatctta ggtacccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt
     3661 aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     3721 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     3781 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     3841 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     3901 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     3961 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     4021 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     4081 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     4141 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     4201 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     4261 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     4321 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     4381 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     4441 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     4501 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     4561 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     4621 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     4681 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     4741 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     4801 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     4861 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     4921 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     4981 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     5041 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     5101 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     5161 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     5221 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     5281 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     5341 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     5401 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     5461 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     5521 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     5581 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     5641 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     5701 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     5761 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccaagct
     5821 taataaacac ggaaggagac aataccggaa ggaacccgcg ctatgacggc aataaaaaga
     5881 cagaataaaa cgcacgggtg ttgggtcgtt tgttcataaa cgcggggttc ggtcccaggg
     5941 ctggcactct gtcgataccc caccgagacc ccattggggc caatacgccc gcgtttcttc
     6001 cttttcccca ccccaccccc caagttcggg tgaaggccca gggctcgcag ccaacgtcgg
     6061 ggcggcaggc cctgccatag ccactggccc cgtgggttag ggacggggtc ccccattcga
     6121 gaataaataa cttcgtataa agtatcctat acgaagttat caccatgacc gagtacaagc
     6181 ccacggtgcg cctcgccacc cgcgacgacg tcccccgggc cgtacgcacc ctcgccgccg
     6241 cgttcgccga ctaccccgcc acgcgccaca ccgtcgaccc ggaccgccac atcgagcggg
     6301 tcaccgagct gcaagaactc ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg
     6361 tcgcggacga cggcgccgcg gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg
     6421 cggtgttcgc cgagatcggc ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc
     6481 agcaacagat ggaaggcctc ctggcgccgc accggcccaa ggagcccgcg tggttcctgg
     6541 ccaccgtcgg cgtctcgccc gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc
     6601 ccggagtgga ggcggccgag cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc
     6661 gcaacctccc cttctacgag cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg
     6721 aaggaccgcg cacctggtgc atgacccgca agcccggtgc ctgacgcccg ccccacgacc
     6781 cgcagcgccc gaccgaaagg agcgcacgac cccatggctc cgaccgaagc cacccggggc
     6841 ggccccgccg accccgcacc cgcccccgag gcccaccgac tctagagctc gctgatcagc
     6901 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
     6961 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
     7021 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
     7081 ggattgggaa gacaatagca ctagttatta atagtaatca attacggggt cattagttca
     7141 tagcccatat atggagttcc g
//
