LOCUS       sequence-430857-        6953 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  The DuET construct encodes the named human GPCR engineered with a 
            cleaved hemagglutinin signal peptide, a 5' FLAG and a 3' 1D4 epitope
            tag optimized for expression in mammalian cells and proteomics 
            studies..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6953)
  AUTHORS   Dahl L, Kotliar IB, Bendes A, Dodig-Crnkovic T, Fromm S, Elofsson A,
            Uhlen M, Sakmar TP, Schwenk JM
  TITLE     Multiplexed selectivity screening of anti-GPCR antibodies.
  JOURNAL   Sci Adv. 2023 May 3;9(18):eadf9297. doi: 10.1126/sciadv.adf9297. 
            Epub 2023 May 3.
  PUBMED    37134173
REFERENCE   2  (bases 1 to 6953)
  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 Adv."; 
            date: "2023-05-3"; volume: "9(18):eadf9297. doi"; pages: " 
            10.1126/sciadv.adf9297. Epub 2023 May 3"
COMMENT     Sequence Label: HTR2A-DuET
FEATURES             Location/Qualifiers
     source          1..6953
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     412..431
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(650..754)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(829..848)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1020..1399
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1400..1603
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1554..1574
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1648..1667
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1648..1666
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3329..3346)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3335..3559
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3605..4033
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3692..3711)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3902..3923
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4042..4062)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4047..4376
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      4227..4362
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     4289..4308
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             4443..5237
                     /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(4497..4516)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5107..5126
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    5411..5532
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5448..5467)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5502..5521
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5581..5597)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5581..5597)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5594..5616)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5605..5621
                     /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(5629..5635,5636..5653,5654..5659))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5674..5695
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5812..5829)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5983..6571)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6063..6082)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6742..6953,1..580,581..649))
                     /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"
ORIGIN
        1 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
       61 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
      121 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
      181 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
      241 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
      301 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
      361 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
      421 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
      481 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
      541 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
      601 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
      661 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
      721 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
      781 acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta caatctgctc
      841 tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg tcgctgagta
      901 gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacattt gcatgaagaa
      961 tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta cgggccagat atacgcgttg
     1021 acattgatta ttgactagtt attaatagta atcaattacg gggtcattag ttcatagccc
     1081 atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct gaccgcccaa
     1141 cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac
     1201 tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca
     1261 agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg
     1321 gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt
     1381 agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc gtggatagcg
     1441 gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga gtttgttttg
     1501 gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat
     1561 gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctctctggc taactagaga
     1621 acccactgct tactggctta tcgaaattaa tacgactcac tatagggaga cccaagctgg
     1681 ctagcgttta aacttaagct tagcgccacc atgaagacga tcatcgccct gagctacatc
     1741 ttctgcctgg tattcgccga ctacaaggac gatgatgacg ccagcatcga tatggatatc
     1801 ttgtgcgagg agaatacctc actgtccagc acaactaact ctctcatgca gctcaatgat
     1861 gatacccgac tttactccaa cgacttcaac agtggtgagg ctaatacatc agacgccttc
     1921 aattggacag tagacagtga aaaccggacc aatctgtcct gtgagggatg cctgtcccca
     1981 agttgtttgt ctctcctgca tttgcaggaa aagaactggt ctgcgctgct caccgccgtg
     2041 gttatcatac tcactatagc tggcaatatt ctcgtaatca tggccgtgtc cctggaaaaa
     2101 aagctgcaga acgccacgaa ctacttcctg atgtctctgg cgattgcgga tatgctgctt
     2161 gggtttctgg tgatgcctgt atccatgttg actatactgt acgggtaccg ctggccactc
     2221 ccatccaagt tgtgtgccgt ctggatctat ctggacgttc tctttagtac cgcaagcata
     2281 atgcaccttt gtgctattag cctggaccgc tacgtcgcta ttcaaaaccc catccaccac
     2341 agcagattca acagccgaac taaagccttt ctgaaaatca tcgcggtctg gaccatttca
     2401 gttggcatca gcatgccgat acctgtattt ggccttcagg atgactctaa ggtcttcaaa
     2461 gagggcagtt gccttctggc tgatgacaat ttcgtactta taggaagctt cgtgtccttc
     2521 ttcattcctc ttaccattat ggtgatcact tacttcctga caattaagtc ccttcaaaag
     2581 gaagcgaccc tttgcgttag cgatttgggc acccgggcca agcttgcaag ttttagcttc
     2641 ctgcctcaat cctccctgag cagcgagaaa ctgttccagc ggagcataca ccgagagccc
     2701 gggtcttata ctggccggcg caccatgcaa agcatcagca acgagcagaa ggcttgcaag
     2761 gtcctgggca ttgtgttttt cctgtttgtt gtgatgtggt gtccattctt tataacgaac
     2821 atcatggcgg ttatctgcaa agagtcctgc aacgaggatg ttatcggagc tttgttgaac
     2881 gtgtttgtat ggatcggata tctgtcctca gctgtgaatc cattggtcta cacccttttt
     2941 aacaaaacct atcggtccgc tttttcccgg tacatccagt gtcagtacaa agagaataag
     3001 aagccactcc agcttatact ggtcaatact atacccgcac tggcatataa gagctcccaa
     3061 ctgcagatgg ggcagaaaaa aaactctaaa caagatgcca agaccacaga caacgactgc
     3121 agtatggtgg ccctgggcaa acagcacagt gaagaagcct ccaaagacaa tagtgacggc
     3181 gttaacgaga aagtctcctg cgttatcgat accggtggac gcaccggtga attcgatgag
     3241 gccagcacca ccgtgtctaa gaccgagact tctcaggtgg cccctgccta actcgagtct
     3301 agagggcccg tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct
     3361 gttgtttgcc cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt
     3421 tcctaataaa atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg
     3481 ggtggggtgg ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg
     3541 gatgcggtgg gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat
     3601 ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     3661 accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc
     3721 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     3781 tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt
     3841 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     3901 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     3961 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4021 tttaacgcga attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct
     4081 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa
     4141 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     4201 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     4261 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     4321 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     4381 tcccgggagc ttgtatatcc attttcggat ctgatcaaga gacaggatga ggatcgtttc
     4441 gcatgattga acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat
     4501 tcggctatga ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt
     4561 cagcgcaggg gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac
     4621 tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg
     4681 tgctcgacgt tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc
     4741 aggatctcct gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa
     4801 tgcggcggct gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc
     4861 gcatcgagcg agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg
     4921 aagagcatca ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg
     4981 acggcgagga tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa
     5041 atggccgctt ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg
     5101 acatagcgtt ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct
     5161 tcctcgtgct ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc
     5221 ttgacgagtt cttctgagcg ggactctggg gttcgaaatg accgaccaag cgacgcccaa
     5281 cctgccatca cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
     5341 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
     5401 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
     5461 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
     5521 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
     5581 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     5641 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     5701 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
     5761 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
     5821 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
     5881 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
     5941 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
     6001 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
     6061 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
     6121 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
     6181 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
     6241 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
     6301 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
     6361 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
     6421 aagaacggta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
     6481 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
     6541 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
     6601 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
     6661 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
     6721 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
     6781 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
     6841 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
     6901 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtg
//
