LOCUS       sequence-407833-        7476 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express 3x FLAG tagged-GINIP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7476)
  AUTHORS   Park JC, Luebbers A, Dao M, Semeano A, Nguyen AM, Papakonstantinou 
            MP, Broselid S, Yano H, Martemyanov KA, Garcia-Marcos M
  TITLE     Fine-tuning GPCR-mediated neuromodulation by biasing signaling 
            through different G protein subunits.
  JOURNAL   Mol Cell. 2023 Jul 20;83(14):2540-2558.e12. doi: 
            10.1016/j.molcel.2023.06.006. Epub 2023 Jun 29.
  PUBMED    37390816
REFERENCE   2  (bases 1 to 7476)
  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.molcel.2023.06.006"; journalName: "Mol Cell"; date: 
            "2023-07-20- 20"; volume: "83"; issue: "14"; pages: "2540-2558.e12"
COMMENT     Sequence Label: p3xFLAG-CMV14-GINIP
FEATURES             Location/Qualifiers
     source          1..7476
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     126..148
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     140..157
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     141..157
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     enhancer        318..697
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        698..901
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     852..872
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             2159..2224
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    2235..2857
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2434..2453)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     primer_bind     complement(2881..2901)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2899..3196
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             3250..4044
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3304..3323)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3914..3933
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4696..4830
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4733..4752)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4787..4806
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4835..4854)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4836..4854)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4868..4884)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4868..4884)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4881..4903)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4892..4908
                     /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(4916..4922,4923..4940,4941..4946))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4961..4982
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5099..5116)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5270..5858)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5350..5369)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6029..6820,6821..6889))
                     /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     6652..6671
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6890..6994)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      7021..7449
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7108..7127)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7318..7339
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ccattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
       61 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
      121 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagctgatct atacattgaa
      181 tcaatattgg caattagcca tattagtcat tggttatata gcataaatca atattggcta
      241 ttggccattg catacgttgt atctatatca taatatgtac atttatattg gctcatgtcc
      301 aatatgaccg ccatgttgac attgattatt gactagttat taatagtaat caattacggg
      361 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
      421 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
      481 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
      541 ccacttggca gtacatcaag tgtatcatat gccaagtccg ccccctattg acgtcaatga
      601 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttacgggact ttcctacttg
      661 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacac
      721 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
      781 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaataaccc
      841 cgccccgttg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
      901 tcgtttagtg aaccgtcaga attaagcttg cggccgcgaa ttcatggggg tgttgatgtc
      961 caagcggcag acagtggagc aggtgcagaa ggtgagtctg gctgtgtctg ctttcaagga
     1021 tgggctgcgg gacaggcctt ccatccgacg cacaggtgag ctgccagggt cccgccgtgg
     1081 cactgtagag ggctccgtcc aggaggtaca ggaggagaag gaagcagaag caggaacctc
     1141 ggtggtccag gaagagagta gtgccggccg cgcagcctgg gagcggctcc gagatgggcg
     1201 cggcgtggag cctgaggagt ttgacaggac aagtcgattc acaccccctg cgttcatccg
     1261 ccccacccgg aagctggatg atgacaaacc tccagaaatc tgcctggagc ccagagagcc
     1321 tgttgtcaac gatgagatgt gtgatgtttg tgaggtctgg acagctgaga gcctcttccc
     1381 gtgcagggtc tgcaccaggg ttttccatga tggctgcctg cgccgcatgg gctacatcca
     1441 aggagacagt gcagcggagg tgacggagat ggcccacaca gaaacaggct ggagctgcca
     1501 ctactgtgac aacatcaact tgctgcttac tgaggaggaa atgtatagcc tcacggagac
     1561 ctttcagcgg tgtaaagtca tccctgattg ctccctgaca ctggaggact ttctgcgtta
     1621 ccgccaccaa gcagccaagc ggggggaccg tgacagggcc ctgagtgagg agcaagaaga
     1681 gcaggcggcc cgccagtttg ctgccctgga ccctgaacat cgaggccaca tagagtggcc
     1741 tgacttcttg tcccatgagt ccctcctgct tctgcagcag ttgcgtcccc agaactctct
     1801 gttgaggctt ctgacagtga aggagcggga gcgagcccga gccgccttcc tggctcgggg
     1861 cagtgggagc accgtcagtg aggcagagtg ccgccgggcc cagcactctt ggttttgcaa
     1921 acggttccca gaggctcctt cctgcagtgt cagcatcagc catgtgggtc ccatcgcaga
     1981 tagcagccca gccagcagca gcagcaagag tcaggacaag accctgctgc ccacagagca
     2041 ggagtccaga tttgtggact ggcctacctt cctgcaagag aatgtcctct acatcctggc
     2101 tgctcgcccc aacagcgcag ccattcacct gaagccccca ggaacgggat cccgggctga
     2161 ctacaaagac catgacggtg attataaaga tcatgacatc gactacaagg atgacgatga
     2221 caagtagtga tcccgggtgg catccctgtg acccctcccc agtgcctctc ctggccctgg
     2281 aagttgccac tccagtgccc accagccttg tcctaataaa attaagttgc atcattttgt
     2341 ctgactaggt gtccttctat aatattatgg ggtggagggg ggtggtatgg agcaaggggc
     2401 aagttgggaa gacaacctgt agggcctgcg gggtctattg ggaaccaagc tggagtgcag
     2461 tggcacaatc ttggctcact gcaatctccg cctcctgggt tcaagcgatt ctcctgcctc
     2521 agcctcccga gttgttggga ttccaggcat gcatgaccag gctcagctaa tttttgtttt
     2581 tttggtagag acggggtttc accatattgg ccaggctggt ctccaactcc taatctcagg
     2641 tgatctaccc accttggcct cccaaattgc tgggattaca ggcgtgaacc actgctccct
     2701 tccctgtcct tctgatttta aaataactat accagcagga ggacgtccag acacagcata
     2761 ggctacctgg ccatgcccaa ccggtgggac atttgagttg cttgcttggc actgtcctct
     2821 catgcgttgg gtccactcag tagatgcctg ttgaattggg tacgcggcca gcttggctgt
     2881 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     2941 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     3001 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     3061 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     3121 ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc cagaagtagt
     3181 gaggaggctt ttttggagga attgatcagc ttgggatctg atcaagagac aggatgagga
     3241 tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag
     3301 aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc
     3361 cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg
     3421 aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc
     3481 gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg
     3541 ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct
     3601 gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg
     3661 aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat
     3721 ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgcgc
     3781 atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg
     3841 gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc
     3901 tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct
     3961 gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat
     4021 cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc gaccaagcga
     4081 cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct
     4141 tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg
     4201 agttcttcgc ccaccccggg ctcgatcccc tcgcgagttg gttcagctgc tgcctgaggc
     4261 tggacgacct cgcggagttc taccggcagt gcaaatccgt cggcatccag gaaaccagca
     4321 gcggctatcc gcgcatccat gcccccgaac tgcaggagtg gggaggcacg atggccgctt
     4381 tggtcgaccc ggacgggacg ctcctgcgcc tgatacagaa cgaattgctt gcaggcatct
     4441 catgagtgtg tcttcccgtt ttccgcctga ggtcactgcg tggatggagc gctggcgcct
     4501 gctgcgcgac ggcgagctgc tcaccaccca ctcgccaagc tggaaccgta aaaaggccgc
     4561 gttgctggcg tttttccata ggctccgccg atcataatca gccataccac atttgtagag
     4621 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     4681 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     4741 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     4801 ctcatcaatg tatcttatca tgtctggatc aattccctat agtgagtcgt attaaattcg
     4861 taatcatgtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca
     4921 tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat
     4981 taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt
     5041 aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct
     5101 cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa
     5161 aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa
     5221 aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc
     5281 tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga
     5341 caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc
     5401 cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt
     5461 ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct
     5521 gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg
     5581 agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta
     5641 gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct
     5701 acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa
     5761 gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt
     5821 gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta
     5881 cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat
     5941 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     6001 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     6061 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     6121 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct
     6181 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     6241 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     6301 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     6361 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     6421 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     6481 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     6541 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     6601 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     6661 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     6721 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     6781 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     6841 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     6901 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     6961 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     7021 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     7081 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     7141 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     7201 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     7261 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     7321 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     7381 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     7441 acgcgaattt taacaaaata ttaacgctta caattt
//
