LOCUS       sequence-388864-        8103 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To express benzoylphenylalanine-specific variant of E. coli TyrRS 
            (pBpaRS) and its cognate amber suppressor tRNA to allow UAG codon to
            be translated into benzoylphenylalanine..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8103)
  AUTHORS   Hino N, Sakamoto K, Yokoyama S
  TITLE     Site-specific incorporation of unnatural amino acids into proteins 
            in mammalian cells.
  JOURNAL   Methods Mol Biol. 2012;794:215-28. doi: 
            10.1007/978-1-61779-331-8_13.
  PUBMED    21956565
REFERENCE   2  (bases 1 to 8103)
  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: "Methods Mol
            Biol."; date: "2012"; volume: "794:215-28. doi"; pages: " 
            10.1007/978-1-61779-331-8_13"
COMMENT     Sequence Label: pcpBpaRS
FEATURES             Location/Qualifiers
     source          1..8103
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(40..59)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(1857..1876)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     enhancer        1973..2352
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2353..2556
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2507..2527
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    2558..2576
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2579..2597
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2607..2631
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4040..4066
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(4107..4124)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4113..4337
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4383..4811
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4470..4489)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4680..4701
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4820..4840)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4825..5154
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5005..5140
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5067..5086
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        5202..5249
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             5268..5642
                     /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    5772..5893
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5809..5828)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5863..5882
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5942..5958)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5942..5958)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5955..5977)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5966..5982
                     /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(5990..5996,5997..6014,6015..6020))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6035..6056
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6173..6190)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6344..6932)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6424..6443)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7103..7894,7895..7963))
                     /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     7726..7745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7964..8068)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc
       61 atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag
      121 caaaatttaa gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag
      181 ggttaggcgt tttgcgctgc ttcggccctt tcgtcttcaa gaattcagcg agtgttaacc
      241 ctgcctagcg ctccggtttt tctgtgctga acctcagggg acgccgacac acgtacacgt
      301 cggaggggta gcgaagtggc taaacgcggc ggactctaaa tccgctccct ttgggttcgg
      361 cggttcgaat ccgtccccct ccagacaagt gcggtttttt tctccagctc ccgccagcag
      421 actggagcgc tccggttttt ctgtgctgaa cctcagggga cgccgacaca cgtacacgtc
      481 ggaggggtag cgaagtggct aaacgcggcg gactctaaat ccgctccctt tgggttcggc
      541 ggttcgaatc cgtccccctc cagacaagtg cggttttttt ctccagctcc cgccagcttc
      601 ctggagtgct ccggtttttc tgtgctgaac ctcaggggac gccgacacac gtacacgtcg
      661 gaggggtagc gaagtggcta aacgcggcgg actctaaatc cgctcccttt gggttcggcg
      721 gttcgaatcc gtccccctcc agacaagtgc ggtttttttc tccagctccc gcgactacga
      781 tattcgcgca gaagcttagc gagtgttaac cctgcctagc gctccggttt ttctgtgctg
      841 aacctcaggg gacgccgaca cacgtacacg tcggaggggt agcgaagtgg ctaaacgcgg
      901 cggactctaa atccgctccc tttgggttcg gcggttcgaa tccgtccccc tccagacaag
      961 tgcggttttt ttctccagct cccgccagca gactggagcg ctccggtttt tctgtgctga
     1021 acctcagggg acgccgacac acgtacacgt cggaggggta gcgaagtggc taaacgcggc
     1081 ggactctaaa tccgctccct ttgggttcgg cggttcgaat ccgtccccct ccagacaagt
     1141 gcggtttttt tctccagctc ccgccagctt cctggagcgc tccggttttt ctgtgctgaa
     1201 cctcagggga cgccgacaca cgtacacgtc ggaggggtag cgaagtggct aaacgcggcg
     1261 gactctaaat ccgctccctt tgggttcggc ggttcgaatc cgtccccctc cagacaagtg
     1321 cggttttttt ctccagctcc cgcgactacg atattcgcgc aggaattcag cgagtgttaa
     1381 ccctgcctag cgctccggtt tttctgtgct gaacctcagg ggacgccgac acacgtacac
     1441 gtcggagggg tagcgaagtg gctaaacgcg gcggactcta aatccgctcc ctttgggttc
     1501 ggcggttcga atccgtcccc ctccagacaa gtgcggtttt tttctccagc tcccgccagc
     1561 agactggagc gctccggttt ttctgtgctg aacctcaggg gacgccgaca cacgtacacg
     1621 tcggaggggt agcgaagtgg ttaaacgcgg cggactctaa atccgctccc tttgggttcg
     1681 gcggttcgaa tccgtccccc tccagacaag tgcggttttt ttctccagct cccgccagct
     1741 tcctggagcg ctccggtttt tctccagctc ccgcgactac gatattcgcg cagggatcct
     1801 ctacgccgga cgcatcgtgg ccggcatcac cggcgccaca ggtgcggttg ctggcgccta
     1861 tatcgccgac atcaccgatg gggaagatcg ggctcgccac ttcgggctca tgagcgcttg
     1921 tttcggcgtg ggtatggtgg caggcccgat gtacgggcca gatatacgcg ttgacattga
     1981 ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg
     2041 gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     2101 cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat
     2161 tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat
     2221 catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat
     2281 gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc
     2341 gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac
     2401 tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa
     2461 aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt
     2521 aggcgtgtac ggtgggaggt ctatataagc agagctctcc ctatcagtga tagagatctc
     2581 cctatcagtg atagagatcg tcgacgagct cgtttagtga accgtcagat cgcctggaga
     2641 cgccatccac gctgttttga cctccataga agacaccggg accgatccag cctccggact
     2701 ctagcgttta aacttaagct tggtaccgag ctcggatcca ctagtccagt gtggtggaat
     2761 tcccaccatg gcaagcagta acttgattaa acaattgcaa gagcgggggc tggtagccca
     2821 ggtgacggac gaggaagcgt tagcagagcg actggcgcaa ggcccgatcg cgctcggttg
     2881 cggcttcgat cctaccgctg acagcttgca tttggggcat cttgttccat tgttatgcct
     2941 gaaacgcttc cagcaggcgg gccacaagcc ggttgcgctg gtaggcggcg cgacgggtct
     3001 gattggcgac ccgagcttca aagctgccga gcgtaagctg aacaccgaag aaactgttca
     3061 ggagtgggtg gacaaaatcc gtaagcaggt tgccccgttc ctcgatttcg actgtggaga
     3121 aaactctgct atcgcggcga acaactatga ctggttcggc aatatgaatg tgctgacctt
     3181 cctgcgcgat attggcaaac acttctccgt taaccagatg atcaacaaag aagcggttaa
     3241 gcagcgtctc aaccgtgaag atcaggggat ttcgttcact gagttttcct acaacctgtt
     3301 gcagggttat ggcttcgcct gtgcgaacaa acagtacggt gtggtgctgc aaattggtgg
     3361 ttctgaccag tggggtaaca tcacttctgg tatcgacctg acccgtcgtc tgcatcagaa
     3421 tcaggtgttt ggcctgaccg ttccgctgat cactaaagca gatggcacca aatttggtaa
     3481 aactgaaggc ggcgcagtct ggttggatcc gaagaaaacc agcccgtaca aattctacca
     3541 gttctggatc aacactgcgg atgccgacgt ttaccgcttc ctgaagttct tcacctttat
     3601 gagcattgaa gagatcaacg ccctggaaga agaagataaa aacagcggta aagcaccgcg
     3661 cgcccagtat gtactggcgg agcaggtgac tcgtctggtt cacggtgaag aaggtttaca
     3721 ggcggcaaaa cgtattaccg aatgcctgtt cagcggttct ttgagtgcgc tgagtgaagc
     3781 ggacttcgaa cagctggcgc aggacggcgt accgatggtt gagatggaaa agggcgcaga
     3841 cctgatgcag gcactggtcg attctgaact gcaaccttcc cgtggtcagg cacgtaaaac
     3901 tatcgcctcc aatgccatca ccattaacgg tgaaaaacag tccgatcctg aatacttctt
     3961 taaagaagaa gatcgtctgt ttggtcgttt taccttactg cgtcgcggta aaaagaatta
     4021 ctgtctgatt tgctggaaat acccatacga tgttccagat tacgcttaac tcgagtctag
     4081 agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     4141 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     4201 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     4261 tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga
     4321 tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc
     4381 ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     4441 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     4501 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     4561 tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg
     4621 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     4681 tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt
     4741 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     4801 taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc
     4861 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag
     4921 tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc
     4981 atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct
     5041 ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct
     5101 gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc
     5161 ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg
     5221 catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg gccaagttga
     5281 ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg
     5341 accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg gtccgggacg
     5401 acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac accctggcct
     5461 gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc gtgtccacga
     5521 acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg tgggggcggg
     5581 agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag gagcaggact
     5641 gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa
     5701 tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct
     5761 tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     5821 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     5881 tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat
     5941 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     6001 ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg
     6061 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     6121 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     6181 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     6241 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     6301 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     6361 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     6421 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     6481 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     6541 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     6601 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     6661 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     6721 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     6781 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     6841 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     6901 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     6961 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     7021 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     7081 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     7141 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     7201 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     7261 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     7321 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     7381 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     7441 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     7501 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     7561 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     7621 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     7681 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     7741 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     7801 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     7861 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     7921 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     7981 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     8041 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     8101 acg
//
