LOCUS       sequence-476308-        7386 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express GPR119 in insect cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7386)
  AUTHORS   Kim D, Liu W, Viner R, Cherezov V
  TITLE     Native mass spectrometry prescreening of G protein-coupled receptor 
            complexes for cryo-EM structure determination.
  JOURNAL   Structure. 2024 Dec 5;32(12):2206-2219.e4. doi: 
            10.1016/j.str.2024.10.004. Epub 2024 Oct 28.
  PUBMED    39471802
REFERENCE   2  (bases 1 to 7386)
  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.str.2024.10.004"; journalName: "Structure"; date: 
            "2024-12-5- 5"; volume: "32"; issue: "12"; pages: "2206-2219.e4"
COMMENT     Sequence Label: IMPT-11371
FEATURES             Location/Qualifiers
     source          1..7386
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      2..456
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(89..108)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     298..319
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        482..586
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(587..655,656..1447)
                     /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(805..824)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1618..2206
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2107..2126
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2360..2377
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  2511..2735
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(2802..3335)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(3524..3529,3530..3546,3547..3552))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     promoter        3904..3995
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     3920..3937
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             4098..4121
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4122..4151
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4152..4175
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             5997..6020
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(6030..6032,6033..6743)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     polyA_signal    6876..7010
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6913..6932)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6967..6986
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  7039..7204
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
       61 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
      121 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
      181 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
      241 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt cttaatagtg
      301 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
      361 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
      421 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
      481 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
      541 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
      601 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      661 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
      721 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
      781 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
      841 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      901 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
      961 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     1021 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     1081 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     1141 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     1201 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1261 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     1321 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     1381 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     1441 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     1501 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     1561 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     1621 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     1681 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     1741 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     1801 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     1861 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     1921 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     1981 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     2041 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     2101 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     2161 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     2221 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     2281 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     2341 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcctgatgcg
     2401 gtattttctc cttacgcatc tgtgcggtat ttcacaccgc atagaccagc cgcgtaacct
     2461 ggcaaaatcg gttacggttg agtaataaat ggatgccctg cgtaagcggg tgtgggcgga
     2521 caataaagtc ttaaactgaa caaaatagat ctaaactatg acaataaagt cttaaactag
     2581 acagaatagt tgtaaactga aatcagtcca gttatgctgt gaaaaagcat actggacttt
     2641 tgttatggct aaagcaaact cttcattttc tgaagtgcaa attgcccgtc gtattaaaga
     2701 ggggcgtggc caagggcatg gtaaagacta tattcgcggc gttgtgacaa tttaccgaac
     2761 aactccgcgg ccgggaagcc gatctcggct tgaacgaatt gttaggtggc ggtacttggg
     2821 tcgatatcaa agtgcatcac ttcttcccgt atgcccaact ttgtatagag agccactgcg
     2881 ggatcgtcac cgtaatctgc ttgcacgtag atcacataag caccaagcgc gttggcctca
     2941 tgcttgagga gattgatgag cgcggtggca atgccctgcc tccggtgctc gccggagact
     3001 gcgagatcat agatatagat ctcactacgc ggctgctcaa acttgggcag aacgtaagcc
     3061 gcgagagcgc caacaaccgc ttcttggtcg aaggcagcaa gcgcgatgaa tgtcttacta
     3121 cggagcaagt tcccgaggta atcggagtcc ggctgatgtt gggagtaggt ggctacgtct
     3181 ccgaactcac gaccgaaaag atcaagagca gcccgcatgg atttgacttg gtcagggccg
     3241 agcctacatg tgcgaatgat gcccatactt gagccaccta actttgtttt agggcgactg
     3301 ccctgctgcg taacatcgtt gctgctgcgt aacatcgttg ctgctccata acatcaaaca
     3361 tcgacccacg gcgtaacgcg cttgctgctt ggatgcccga ggcatagact gtacaaaaaa
     3421 acagtcataa caagccatga aaaccgccac tgcgccgtta ccaccgctgc gttcggtcaa
     3481 ggttctggac cagttgcgtg agcgcatacg ctacttgcat tacagtttac gaaccgaaca
     3541 ggcttatgtc aactgggttc gtgccttcat ccgtttccac ggtgtgcgtc acccggcaac
     3601 cttgggcagc agcgaagtcg aggcatttct gtcctggctg gcgaacgagc gcaaggtttc
     3661 ggtctccacg catcgtcagg cattggcggc cttgctgttc ttctacggca aggtgctgtg
     3721 cacggatctg ccctggcttc aggagatcgg aagacctcgg ccgtcgcggc gcttgccggt
     3781 ggtgctgacc ccggatgaag tggttcgcat cctcggtttt ctggaaggcg agcatcgttt
     3841 gttcgcccag gactctagct atagttctag tggttggcta cgtatactcc ggaatattaa
     3901 tagatcatgg agataattaa aatgataacc atctcgcaaa taaataagta ttttactgtt
     3961 ttcgtaacag ttttgtaata aaaaaaccta taaatattcc ggattattca taccgtccca
     4021 ccatcgggcg cggatccatg aagacgatta tcgcactttc gtacattttt tgtcttgtct
     4081 tcgccggcgc tcctggtgat tacaaggatg atgacgataa acatcatcac catcaccatc
     4141 atcatcatca tctggaagtg ctttttcaag ggccgggcgc cgacctggaa gacaactggg
     4201 aaaccttaaa tgacaatctg aaagtaatag aaaaggcaga caatgcggct caagtaaagg
     4261 atgctctgac caagatgcgg gcagctgcgc ttgatgcgca gaaagccaca cccccaaaat
     4321 tagaagacaa gtcacctgac agcccagaga tgaaggactt caggcacggt ttcgacatat
     4381 tagtgggaca gatcgacgat gccctaaagc tcgccaacga aggaaaggtc aaagaagcac
     4441 aggctgcggc cgaacaattg aaaacaacgc gtaacgctta tatccaaaaa tacctaggct
     4501 cgccaataat ggagagctcg ttttcatttg gggttatcct agcggtattg gcgagcctca
     4561 tcatagccac gaacacactt gttgcggtag cggtactgct cttaatccat aaaaacgacg
     4621 gcgtatccct atgttttacc ctcaacctag cggttgccga caccttaata ggtgtcgcaa
     4681 tttcgggatt actgacagac caactgtcat ccccttctag acctacgcag aagacacttt
     4741 gctcgctcag aatggctttc gtgacgtcgt ccgcagcagc atctgttctg accgtgatgt
     4801 tgataacctt tgaccggtat ctagcaatta agcaaccctt ccgatacttg aagataatgt
     4861 cgggatttgt cgcgggggcg tgcattgctg gactttggct cgtctcttat ttaatagggt
     4921 ttctgccact ggggataccg atgttccaac agacagcgta caaaggtcag tgctcatttt
     4981 tcgccgtttt ccaccctcac tttgtgctaa ctctgtcctg tgtcggattc ttccccgcaa
     5041 tgcttctttt tgtgttcttc tattgcgata tgctaaaaat tgcttctatg cactctcaac
     5101 aaatacgtaa gatggagcat gccggcgcca tggcgggagg ttatagaagt ccgcgaacac
     5161 cctctgattt caaagccctc cggactgtga gcgtgctcat cgggagtttt gcgttatcat
     5221 ggaccccgtt ccttatcact ggaattgtac aagtagcttg tcaagagtgt cacttgtact
     5281 tagtgttgga acgctatctg tggctcctag gcgtgggcaa cagtctgctc aatcctctga
     5341 tctatgcata ttggcaaaag gaggtcaggt tgcaattata ccatatggca ctgggggtaa
     5401 agaaggtcct tacgagtttt ctcctgttcc tcagtgctcg caactgtggg cccgaacgac
     5461 cacgtgaatc cagctgccac attgttacca ttagtagcag tgagtttgac ggggtcttta
     5521 ctttggagga ctttgttgga gattgggaac aaacggctgc ttataacctg gatcaagttc
     5581 tcgagcaagg aggcgtttct tcgctacttc agaatcttgc cgtttccgtt actccgatcc
     5641 agcgaatagt ccggtctggg gaaaatgcgc tgaaaataga tatacacgtg atcataccct
     5701 acgaggggct ctctgcggac cagatggccc agatcgagga ggtgttcaaa gtcgtgtatc
     5761 ctgttgacga ccaccacttt aaggtcattt taccgtacgg aaccctagtc attgacggag
     5821 tgacgcccaa tatgctcaat tatttcggcc ggccatacga agggattgcg gtttttgatg
     5881 gcaagaagat aacggtcact ggcacgcttt ggaacggtaa caaaattatc gacgagcgtt
     5941 tgattacacc tgatgggtcg atgctcttta gagtaactat taactccggc ggatcacttg
     6001 aagtcttgtt tcagggaccg ggaggttccg tatcaaaggg agaggagcta tttacgggtg
     6061 ttgtaccaat cctcgtcgaa ttggatggtg atgtgaatgg tcacaagttc tcggtaagcg
     6121 gcgagggtga gggtgatgca acctatggca aattgacact gaaattcatt tgcaccactg
     6181 ggaaattgcc agtcccgtgg cctactctcg tcacaacctt gacttatggc gttcagtgct
     6241 tcagcagata tcctgaccat atgaaacagc atgacttttt caaatccgct atgcccgagg
     6301 ggtatgttca ggagcgcact atcttcttca aagatgacgg gaactacaag acgagggcag
     6361 aagttaaatt cgaaggtgat acactcgtaa ataggattga actaaagggc atcgatttca
     6421 aagaggatgg caatatacta ggtcataagt tggaatataa ttacaacagt cataacgtat
     6481 acatcatggc agacaagcag aaaaatggca taaaagttaa ttttaaaatt cgacacaata
     6541 tagaagacgg ttcagtacag ttagccgacc attaccaaca gaatactcca attggagatg
     6601 gtccggtgtt attaccagat aaccactacc tttctaccca gtcagcctta agtaaagatc
     6661 ctaacgagaa acgcgatcac atggtattgc tagaatttgt gactgcagct ggaatcacat
     6721 taggtatgga tgagctatac aaatgaaagc ttgtcgagaa gtactagagg atcataatca
     6781 gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga
     6841 acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg
     6901 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
     6961 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctggatc tgatcactgc
     7021 ttgagcctag gagatccgaa ccagataagt gaaatctagt tccaaactat tttgtcattt
     7081 ttaattttcg tattagctta cgacgctaca cccagttccc atctattttg tcactcttcc
     7141 ctaaataatc cttaaaaact ccatttccac ccctcccagt tcccaactat tttgtccgcc
     7201 cacagcgggg catttttctt cctgttatgt ttttaatcaa acatcctgcc aactccatgt
     7261 gacaaaccgt catcttcggc tactttttct ctgtcacaga atgaaaattt ttctgtcatc
     7321 tcttcgttat taatgtttgt aattgactga atatcaacgc ttatttgcag cctgaatggc
     7381 gaatgg
//
