LOCUS       sequence-224188-        7963 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mammalian expression plasmid for FLAG-tagged human T1R3 with signal 
            peptide of HLA class I histocompatibility antigen A-2 alpha chain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7963)
  AUTHORS   Park J, Selvam B, Sanematsu K, Shigemura N, Shukla D, Procko E
  TITLE     Structural architecture of a dimeric class C GPCR based on 
            co-trafficking of sweet taste receptor subunits.
  JOURNAL   J Biol Chem. 2019 Feb 5. pii: RA118.006173. doi: 
            10.1074/jbc.RA118.006173.
  PUBMED    30723160
REFERENCE   2  (bases 1 to 7963)
  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: "J Biol 
            Chem. 2019 Feb 5. pii: RA118.006173. doi: 10.1074/jbc.RA118.006173."
COMMENT     Sequence Label: pcDNA3.1(+)-HLA-Flag-natT1R3
FEATURES             Location/Qualifiers
     source          1..7963
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(38..55)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    44..268
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      314..742
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(401..420)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     611..632
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(751..771)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        756..1085
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      936..1071
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             1152..1946
                     /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(1206..1225)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1816..1835
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2120..2241
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2157..2176)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2211..2230
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2290..2306)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2290..2306)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2303..2325)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2314..2330
                     /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(2338..2344,2345..2362,2363..2368))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2383..2404
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2521..2538)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2692..3280)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2772..2791)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3451..4242,4243..4311))
                     /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     4074..4093
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4312..4416)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4491..4510)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4682..5061
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5062..5265
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5216..5236
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5310..5329
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5310..5328
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      5348..5357
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5432..5455
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
ORIGIN
        1 ctcgagtcta gagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
       61 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
      121 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
      181 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
      241 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
      301 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
      361 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
      421 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
      481 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
      541 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
      601 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
      661 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
      721 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
      781 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
      841 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
      901 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
      961 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     1021 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     1081 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcaagag acaggatgag
     1141 gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg
     1201 agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt
     1261 tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc
     1321 tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt
     1381 gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag
     1441 tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg
     1501 ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag
     1561 cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg
     1621 atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc
     1681 gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca
     1741 tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc
     1801 gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg
     1861 ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct
     1921 atcgccttct tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc
     1981 gacgcccaac ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg
     2041 cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct
     2101 ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca aataaagcaa
     2161 tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt gtggtttgtc
     2221 caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct agagcttggc
     2281 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     2341 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     2401 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca
     2461 ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc
     2521 ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc
     2581 aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc
     2641 aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag
     2701 gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc
     2761 gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt
     2821 tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct
     2881 ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg
     2941 ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct
     3001 tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat
     3061 tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg
     3121 ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa
     3181 aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt
     3241 ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc
     3301 tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt
     3361 atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta
     3421 aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat
     3481 ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac
     3541 tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg
     3601 ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag
     3661 tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt
     3721 aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt
     3781 gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt
     3841 tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt
     3901 cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct
     3961 tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt
     4021 ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac
     4081 cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa
     4141 actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa
     4201 ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca
     4261 aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct
     4321 ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga
     4381 atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc
     4441 tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag tacaatctgc
     4501 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
     4561 tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag
     4621 aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt
     4681 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
     4741 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     4801 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     4861 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     4921 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     4981 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     5041 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
     5101 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     5161 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     5221 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg gctaactaga
     5281 gaacccactg cttactggct tatcgaaatt aatacgactc actataggga gacccaagct
     5341 ggctagcgcc accatggccg tcatggcgcc ccgaaccctc gtcctgctac tctcgggggc
     5401 cctggccctg acccagacct gggcgggctc cgactacaag gacgatgatg acaagggagg
     5461 atccgcccca ttgtgcctgt cacagcaact taggatgaag ggggactacg tgctgggggg
     5521 gctgttcccc ctgggcgagg ccgaggaggc tggcctccgc agccggacac ggcccagcag
     5581 ccctgtgtgc accaggttct cctcaaacgg cctgctctgg gcactggcca tgaaaatggc
     5641 cgtggaggag atcaacaaca agtcggatct gctgcccggg ctgcgcctgg gctacgacct
     5701 ctttgatacg tgctcggagc ctgtggtggc catgaagccc agcctcatgt tcctggccaa
     5761 ggcaggcagc cgcgacatcg ccgcctactg caactacacg cagtaccagc cccgtgtgct
     5821 ggctgtcatc gggccccact cgtcagagct cgccatggtc accggcaagt tcttcagctt
     5881 cttcctcatg ccccaggtca gctacggtgc tagcatggag ctgctgagcg cccgggagac
     5941 cttcccctcc ttcttccgca ccgtgcccag cgaccgtgtg cagctgacgg ccgccgcgga
     6001 gctgctgcag gagttcggct ggaactgggt ggccgccctg ggcagcgacg acgagtacgg
     6061 ccggcagggc ctgagcatct tctcggccct ggccgcggca cgcggcatct gcatcgcgca
     6121 cgagggcctg gtgccgctgc cccgtgccga tgactcgcgg ctggggaagg tgcaggacgt
     6181 cctgcaccag gtgaaccaga gcagcgtgca ggtggtgctg ctgttcgcct ccgtgcacgc
     6241 cgcccacgcc ctcttcaact acagcatcag cagcaggctc tcgcccaagg tgtgggtggc
     6301 cagcgaggcc tggctgacct ctgacctggt catggggctg cccggcatgg cccagatggg
     6361 cacggtgctt ggcttcctcc agaggggtgc ccagctgcac gagttccccc agtacgtgaa
     6421 gacgcacctg gccctggcca ccgacccggc cttctgctct gccctgggcg agagggagca
     6481 gggtctggag gaggacgtgg tgggccagcg ctgcccgcag tgtgactgca tcacgctgca
     6541 gaacgtgagc gcagggctaa atcaccacca gacgttctct gtctacgcag ctgtgtatag
     6601 cgtggcccag gccctgcaca acactcttca gtgcaacgcc tcaggctgcc ccgcgcagga
     6661 ccccgtgaag ccctggcagc tcctggagaa catgtacaac ctgaccttcc acgtgggcgg
     6721 gctgccgctg cggttcgaca gcagcggaaa cgtggacatg gagtacgacc tgaagctgtg
     6781 ggtgtggcag ggctcagtgc ccaggctcca cgacgtgggc aggttcaacg gcagcctcag
     6841 gacagagcgc ctgaagatcc gctggcacac gtctgacaac cagaagcccg tgtcccggtg
     6901 ctcgcggcag tgccaggagg gccaggtgcg ccgggtcaag gggttccact cctgctgcta
     6961 cgactgtgtg gactgcgagg cgggcagcta ccggcaaaac ccagacgaca tcgcctgcac
     7021 cttttgtggc caggatgagt ggtccccgga gcgaagcaca cgctgcttcc gccgcaggtc
     7081 tcggttcctg gcatggggcg agccggctgt gctgctgctg ctcctgctgc tgagcctggc
     7141 gctgggcctt gtgctggctg ctttggggct gttcgttcac catcgggaca gcccactggt
     7201 tcaggcctcg ggggggcccc tggcctgctt tggcctggtg tgcctgggcc tggtctgcct
     7261 cagcgtcctc ctgttccctg gccagcccag ccctgcccga tgcctggccc agcagccctt
     7321 gtcccacctc ccgctcacgg gctgcctgag cacactcttc ctgcaggcgg ccgagatctt
     7381 cgtggagtca gaactgcctc tgagctgggc agaccggctg agtggctgcc tgcgggggcc
     7441 ctgggcctgg ctggtggtgc tgctggccat gctggtggag gtcgcactgt gcacctggta
     7501 cctggtggcc ttcccgccgg aggtggtgac ggactggcac atgctgccca cggaggcgct
     7561 ggtgcactgc cgcacacgct cctgggtcag cttcggccta gcgcacgcca ccaatgccac
     7621 gctggccttt ctctgcttcc tgggcacttt cctggtgcgg agccagccgg gccgctacaa
     7681 ccgtgcccgt ggcctcacct ttgccatgct ggcctacttc atcacctggg tctcctttgt
     7741 gcccctcctg gccaatgtgc aggtggtcct caggcccgcc gtgcagatgg gcgccctcct
     7801 gctctgtgtc ctgggcatcc tggctgcctt ccacctgccc aggtgttacc tgctcatgcg
     7861 gcagccaggg ctcaacaccc ccgagttctt cctgggaggg ggccctgggg atgcccaagg
     7921 ccagaatgac gggaacacag gaaatcaggg gaaacatgag tga
//
