LOCUS       sequence-343903-        6867 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For the mammalian expression of rat G-alpha-i3 internally tagged 
            with YFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6867)
  AUTHORS   Marivin A, Leyme A, Parag-Sharma K, DiGiacomo V, Cheung AY, Nguyen 
            LT, Dominguez I, Garcia-Marcos M
  TITLE     Dominant-negative Galpha subunits are a mechanism of dysregulated 
            heterotrimeric G protein signaling in human disease.
  JOURNAL   Sci Signal. 2016 Apr 12;9(423):ra37. doi: 10.1126/scisignal.aad2429.
  PUBMED    27072656
REFERENCE   2  (bases 1 to 6867)
  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: "Sci 
            Signal."; date: "2016-04-12"; volume: "9(423):ra37. doi"; pages: " 
            10.1126/scisignal.aad2429"
COMMENT     Sequence Label: pcDNA3.1(-)-G-alpha-i3-YFP (Citrine variant, b/c 
            loo...
FEATURES             Location/Qualifiers
     source          1..6867
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(86..415)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      100..235
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(154..173)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     400..420
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(429..857)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(539..560)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     751..770
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    903..1127
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     1116..1133
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(join(2002..2712,2713..2715,2716..2718))
                     /codon_start=1
                     /product="enhanced variant of YFP (Heikal et al., 2001)"
                     /label=Citrine
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLMCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2044..2065)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2393..2412
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2652..2673
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3126..3145)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3127..3145)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(3190..3393)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(3219..3239)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3394..3773
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     3945..3964
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        4039..4143
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4144..4212,4213..5004)
                     /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     complement(4362..4381)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5175..5763
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5664..5683
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5917..5934
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    6051..6072
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6087..6092,6093..6110,6111..6117)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6125..6141
                     /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)."
     primer_bind     6130..6152
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6149..6165
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6149..6165
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    6214..6335
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6225..6244)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     6279..6298
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6465..6839)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(join(6858..6867,1..38))
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
ORIGIN
        1 actatgccga tatactatgc cgatgattaa ttgtcaacac gtgctgatca gatccgaaaa
       61 tggatataca agctcccggg agctttttgc aaaagcctag gcctccaaaa aagcctcctc
      121 actacttctg gaatagctca gaggcagagg cggcctcggc ctctgcataa ataaaaaaaa
      181 ttagtcagcc atggggcgga gaatgggcgg aactgggcgg agttaggggc gggatgggcg
      241 gagttagggg cgggactatg gttgctgact aattgagatg catgctttgc atacttctgc
      301 ctgctgggga gcctggggac tttccacacc tggttgctga ctaattgaga tgcatgcttt
      361 gcatacttct gcctgctggg gagcctgggg actttccaca ccctaactga cacacattcc
      421 acagaattaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
      481 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
      541 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
      601 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
      661 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
      721 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
      781 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
      841 gccgctacag ggcgcgtggg gataccccct agagccccag ctggttcttt ccgcctcaga
      901 agccatagag cccaccgcat ccccagcatg cctgctattg tcttcccaat cctccccctt
      961 gctgtcctgc cccaccccac cccccagaat agaatgacac ctactcagac aatgcgatgc
     1021 aatttcctca ttttattagg aaaggacagt gggagtggca ccttccaggg tcaaggaagg
     1081 cacgggggag gggcaaacaa cagatggctg gcaactagaa ggcacagtcg aggctgatca
     1141 gcggtttaaa cttaagcttg gtaccgagct cggatccact agtccagtgt ggtggaattc
     1201 tgcagatatc cagcacagtg gcggccgctc gagcggccgc cagtgtgatg gatatctgca
     1261 gaattcggat cctcagtaaa gcccacattc ctttaagttg tttttaatga tgacatctgt
     1321 aacagcatca aaaacaaact gcacgttttt ggtgtcagtg gcacaggtaa agtgagtgta
     1381 gacctccttg gtgtcctttc ttcggttcag atcttcaaac tggcactgga tgtaagcagc
     1441 tgcctcttcg tacgtattgg aacctgtgta ttctggataa cagattgtta atggactcct
     1501 ctttattttt tcctcaaaaa ggtctttctt attaaggaag agaatgattg aagtgtctgt
     1561 aaaccatttg ttgttacaaa tgctgtcaaa caatttcatg ctttcatgca ttcggttcat
     1621 ttcctcgtcc tcagccagaa caaggtcgta atcactgaga gccacacaaa agataattgc
     1681 tgtcactccc tcaaaacagt ggatccactt tttccgttcg gatctttggc cacctacatc
     1741 aaacattttg aagtagagct ccttgaaggt gaagtgggtc tccacaatgc ctgtagtctt
     1801 cactctcgtc cgaagaacat cttgctgagt tggaatgtag ttggtctggg atattctatc
     1861 caaatcattt aggtaatatg aagcagaatc attgagctga tattccctgg acctgctgaa
     1921 gcatgcctgc accccgccat ctcgccataa acgtttaatc acgcctgcta gttctgaagt
     1981 catgactcct tcctcagcgc tcttgtacag ctcgtccatg ccgagagtga tcccggcggc
     2041 ggtcacgaac tccagcagga ccatgtgatc gcgcttctcg ttggggtctt tgctcagggc
     2101 ggactggtag ctcaggtagt ggttgtcggg cagcagcacg gggccgtcgc cgatgggggt
     2161 gttctgctgg tagtggtcgg cgagctgcac gctgccgtcc tcgatgttgt ggcggatctt
     2221 gaagttcacc ttgatgccgt tcttctgctt gtcggccatg atatagacgt tgtggctgtt
     2281 gtagttgtac tccagcttgt gccccaggat gttgccgtcc tccttgaagt cgatgccctt
     2341 cagctcgatg cggttcacca gggtgtcgcc ctcgaacttc acctcggcgc gggtcttgta
     2401 gttgccgtcg tccttgaaga agatggtgcg ctcctggacg tagccttcgg gcatggcgga
     2461 cttgaagaag tcgtgctgct tcatgtggtc ggggtagcgg gcgaagcaca tcaggccgta
     2521 gccgaaggtg gtcacgaggg tgggccaggg cacgggcagc ttgccggtgg tgcagatgaa
     2581 cttcagggtc agcttgccgt aggtggcatc gccctcgccc tcgccggaca cgctgaactt
     2641 gtggccgttt acgtcgccgt ccagctcgac caggatgggc accaccccgg tgaacagctc
     2701 ctcgcccttg ctcaccatag cgctgccagc taaaacaaat aactgtcggg catcatccgc
     2761 tctggcagct tccccaaaat caatcttcaa ccgtcccatg gctcttatga ttgcaatgat
     2821 ggactgaatg gtattgctgt agacgacaac tttatactgc ttacactcgt cctcggaata
     2881 gccatcctca tgaatgattt tcatctgttt cacaatagta cttttaccag attctccagc
     2941 gccgagcagc agcagcttca cttctttggc cgctttctct ccgtcctccc gcaagttgcg
     3001 gtcgatcatc ttactccgct ccaccgccgc cttgtcctcg gcgctcaacg tgcagcccat
     3061 gagaattcca gcacactggc ggccgttact agagggcccg tttaaacgct agccagcttg
     3121 ggtctcccta tagtgagtcg tattaatttc gataagccag taagcagtgg gttctctagt
     3181 tagccagaga gctctgctta tatagacctc ccaccgtaca cgcctaccgc ccatttgcgt
     3241 caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt gccaaaacaa
     3301 actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa accgctatcc
     3361 acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga ctaatacgta
     3421 gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat gccaggcggg
     3481 ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca cttgatgtac
     3541 tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga aagtccctat
     3601 tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt cgttgggcgg
     3661 tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat atgggctatg
     3721 aactaatgac cccgtaattg attactatta ataactagtc aataatcaat gtcaacgcgt
     3781 atatctggcc cgtacatcgc gaagcagcgc aaaacgccta accctaagca gattcttcat
     3841 gcaattgtcg gtcaagcctt gccttgttgt agcttaaatt ttgctcgcgc actactcagc
     3901 gacctccaac acacaagcag ggagcagata ctggcttaac tatgcggcat cagagcagat
     3961 tgtactgaga gtgcaccata ggggatcggg agatctcccg atccgtcgac gtcaggtggc
     4021 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     4081 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     4141 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     4201 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     4261 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     4321 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     4381 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     4441 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     4501 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     4561 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     4621 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     4681 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     4741 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     4801 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     4861 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     4921 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     4981 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     5041 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     5101 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     5161 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     5221 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     5281 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     5341 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     5401 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     5461 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     5521 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     5581 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     5641 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     5701 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     5761 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     5821 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     5881 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     5941 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     6001 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt
     6061 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
     6121 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag
     6181 ctctagctag aggtcgacgg tatacagaca tgataagata cattgatgag tttggacaaa
     6241 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     6301 tatttgtaac cattataagc tgcaataaac aagttggggt gggcgaagaa ctccagcatg
     6361 agatccccgc gctggaggat catccagccg gcgtcccgga aaacgattcc gaagcccaac
     6421 ctttcataga aggcggcggt ggaatcgaaa tctcgtagca cgtgtcagtc ctgctcctcg
     6481 gccacgaagt gcacgcagtt gccggccggg tcgcgcaggg cgaactcccg cccccacggc
     6541 tgctcgccga tctcggtcat ggccggcccg gaggcgtccc ggaagttcgt ggacacgacc
     6601 tccgaccact cggcgtacag ctcgtccagg ccgcgcaccc acacccaggc cagggtgttg
     6661 tccggcacca cctggtcctg gaccgcgctg atgaacaggg tcacgtcgtc ccggaccaca
     6721 ccggcgaagt cgtcctccac gaagtcccgg gagaacccga gccggtcggt ccagaactcg
     6781 accgctccgg cgacgtcgcg cgcggtgagc accggaacgg cactggtcaa cttggccatg
     6841 gtttagttcc tcaccttgtc gtattat
//
