LOCUS       sequence-442063-        6111 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression plasmid for FLAG-tagged AT1R.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6111)
  AUTHORS   Skiba MA, Sterling SM, Rawson S, Zhang S, Xu H, Jiang H, Nemeth GR, 
            Gilman MSA, Hurley JD, Shen P, Staus DP, Kim J, McMahon C, Lehtinen 
            MK, Rockman HA, Barth P, Wingler LM, Kruse AC
  TITLE     Antibodies expand the scope of angiotensin receptor pharmacology.
  JOURNAL   Nat Chem Biol. 2024 May 14. doi: 10.1038/s41589-024-01620-6.
  PUBMED    38744986
REFERENCE   2  (bases 1 to 6111)
  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: "Nat Chem 
            Biol. 2024 May 14. doi: 10.1038/s41589-024-01620-6."
COMMENT     Sequence Label: pCDNA3.1_Zeo_TetO_FLAG-AT1R
FEATURES             Location/Qualifiers
     source          1..6111
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    820..838
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    841..859
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     869..893
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             981..1004
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     complement(2119..2136)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2125..2349
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2395..2823
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2482..2501)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2692..2713
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2832..2852)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2837..3166
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3017..3152
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3079..3098
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3214..3261
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3280..3654
                     /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"
     polyA_signal    3784..3905
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3821..3840)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3875..3894
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3954..3970)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3954..3970)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3967..3989)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3978..3994
                     /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(4002..4008,4009..4026,4027..4032))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4047..4068
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4185..4202)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4356..4944)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4436..4455)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5115..5906,5907..5975))
                     /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     5738..5757
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5976..6080)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggattttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtccgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta cgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacaccaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaaagtcgta ataaccccgc cccgttgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc
      841 tccctatcag tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atctctagaa
      901 gctgggtacc agctgctagc aagcttgcca ccatgaagac gatcatcgcc ctgagctaca
      961 tcttctgcct ggtgttcgcc gactacaagg acgatgatga caaaattctc aactcttcta
     1021 ctgaagatgg tattaaaaga atccaagatg attgtcccaa agctggaagg cataattaca
     1081 tatttgtcat gattcctact ttatacagta tcatctttgt ggtgggaata tttggaaaca
     1141 gcttggtggt gatagtcatt tacttttata tgaagctgaa gactgtggcc agtgtttttc
     1201 ttttgaattt agcactggct gacttatgct ttttactgac tttgccacta tgggctgtct
     1261 acacagctat ggaataccgc tggccctttg gcaattacct atgtaagatt gcttcagcca
     1321 gcgtcagttt caacctgtac gctagtgtgt ttctactcac gtgtctcagc attgatcgat
     1381 acctggctat tgttcaccca atgaagtccc gccttcgacg cacaatgctt gtagccaaag
     1441 tcacctgcat catcatttgg ctgctggcag gcttggccag tttgccagct ataatccatc
     1501 gaaatgtatt tttcattgag aacaccaata ttacagtttg tgctttccat tatgagtccc
     1561 aaaattcaac ccttccgata gggctgggcc tgaccaaaaa tatactgggt ttcctgtttc
     1621 cttttctgat cattcttaca agttatactc ttatttggaa ggccctaaag aaggcttatg
     1681 aaattcagaa gaacaaacca agaaatgatg atatttttaa gataattatg gcaattgtgc
     1741 ttttcttttt cttttcctgg attccccacc aaatattcac ttttctggat gtattgattc
     1801 aactaggcat catacgtgac tgtagaattg cagatattgt ggacacggcc atgcctatca
     1861 ccatttgtat agcttatttt aacaattgcc tgaatcctct tttttatggc tttctgggga
     1921 aaaaatttaa aagatatttt ctccagcttc taaaatatat tcccccaaaa gccaaatccc
     1981 actcaaacct ttcaacaaaa atgagcacgc tttcctaccg cccctcagat aatgtaagct
     2041 catccaccaa gaagcctgca ccatgttttg aggttgagtg actcgagtct agagggcccg
     2101 tttaaacccg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     2161 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     2221 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     2281 ggcaggacag caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg
     2341 gctctatggc ttctgaggcg gaaagaacca gctggggctc tagggggtat ccccacgcgc
     2401 cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac
     2461 ttgccagcgc cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg
     2521 ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt
     2581 tacggcacct cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc
     2641 cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct
     2701 tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga
     2761 ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga
     2821 attaattctg tggaatgtgt gtcagttagg gtgtggaaag tccccaggct ccccagcagg
     2881 cagaagtatg caaagcatgc atctcaatta gtcagcaacc aggtgtggaa agtccccagg
     2941 ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc
     3001 gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca
     3061 tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct ctgagctatt
     3121 ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc tcccgggagc
     3181 ttgtatatcc attttcggat ctgatcagca cgtgttgaca attaatcatc ggcatagtat
     3241 atcggcatag tataatacga caaggtgagg aactaaacca tggccaagtt gaccagtgcc
     3301 gttccggtgc tcaccgcgcg cgacgtcgcc ggagcggtcg agttctggac cgaccggctc
     3361 gggttctccc gggacttcgt ggaggacgac ttcgccggtg tggtccggga cgacgtgacc
     3421 ctgttcatca gcgcggtcca ggaccaggtg gtgccggaca acaccctggc ctgggtgtgg
     3481 gtgcgcggcc tggacgagct gtacgccgag tggtcggagg tcgtgtccac gaacttccgg
     3541 gacgcctccg ggccggccat gaccgagatc ggcgagcagc cgtgggggcg ggagttcgcc
     3601 ctgcgcgacc cggccggcaa ctgcgtgcac ttcgtggccg aggagcagga ctgacacgtg
     3661 ctacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     3721 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     3781 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     3841 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     3901 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     3961 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     4021 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     4081 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     4141 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     4201 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     4261 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     4321 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     4381 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     4441 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     4501 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     4561 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     4621 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     4681 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     4741 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     4801 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     4861 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     4921 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     4981 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     5041 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     5101 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     5161 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     5221 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     5281 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     5341 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     5401 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     5461 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     5521 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     5581 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     5641 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     5701 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     5761 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     5821 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     5881 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     5941 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     6001 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     6061 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
