LOCUS       sequence-390740-        8003 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Protein expression and characterization.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8003)
  AUTHORS   Chen Z, Zakrzewska S, Hajare HS, Alvarez-Buylla A, Abderemane-Ali F,
            Bogan M, Ramirez D, O'Connell LA, Du Bois J, Minor DL Jr
  TITLE     Definition of a saxitoxin (STX) binding code enables discovery and 
            characterization of the anuran saxiphilin family.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 Nov;119(44):e2210114119. doi: 
            10.1073/pnas.2210114119. Epub 2022 Oct 24.
  PUBMED    36279441
REFERENCE   2  (bases 1 to 8003)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2022-11"; volume: "119(44):e2210114119. 
            doi"; pages: " 10.1073/pnas.2210114119. Epub 2022 Oct 24"
COMMENT     Sequence Label: RcSxph F784S
FEATURES             Location/Qualifiers
     source          1..8003
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        146..237
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     162..179
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     CDS             2824..2847
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     CDS             join(2863..2865,2866..3576)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2905..2926)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3166..3185)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3513..3534
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3589..3618
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     polyA_signal    3754..3888
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3791..3810)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3845..3864
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  3917..4082
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     rep_origin      4266..4720
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4353..4372)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4562..4583
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4746..4850
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4851..4919,4920..5711)
                     /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(5069..5088)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5882..6470
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6371..6390
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6624..6641
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  6775..6999
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(7066..7599)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLWSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(7769..7774,7775..7791,7792..7797))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
ORIGIN
        1 ggccgtcgcg gcgcttgccg gtggtgctga ccccggatga agtggttcgc atcctcggtt
       61 ttctggaagg cgagcatcgt ttgttcgccc aggactctag ctatagttct agtggttggc
      121 tacgtatact ccggaatatt aatagatcat ggagataatt aaaatgataa ccatctcgca
      181 aataaataag tattttactg ttttcgtaac agttttgtaa taaaaaaacc tataaatatt
      241 ccggattatt cataccgtcc caccatcggg cgcggatcca tggctcctac cttccagacc
      301 gctctgttct tcaccatcat ctccctgtcc ttcgctatgg ccccgaacgc aaagcaagtc
      361 cgctggtgtg ctatcagcga cctggagcag aagaaatgca acgatctggt tggtagctgc
      421 aacgtgccgg acatcaccct ggtgtgcgtt ctgcgtagca gcaccgaaga ttgcatgacc
      481 gcgatcaagg acggtcaggc ggatgctatg ttcctggaca gcggcgaggt gtacgaagcg
      541 agcaaagatc cgtacaacct gaagccgatc atcgctgaac cgtacagcag caaccgtgac
      601 ctgcagaagt gcctgaaaga gcgtcagcag gcgctggcta agaaaatgat cggtcactac
      661 atcccgcagt gcgatgagaa aggcaactac caaccgcagc agtgccacgg tagcaccggt
      721 cactgctggt gcgttaacgc gatgggtgaa aagatcagcg gtaccaacac cccgccgggt
      781 cagacccgtg ctacctgcga gcgtcacgaa ctgccgaagt gcctgaaaga acgtcaggtg
      841 gcgctgggtg gcgacgagaa agttctgggt cgtttcgtgc cgcagtgcga tgaaaagggc
      901 aactacgagc cgcagcagtt ccacggtagc accggctact cctggtgtgt taacgcgatc
      961 ggcgaggaaa tcgctggcac caaaaccccg ccgggcaaga tcccggctac ctgccagaag
     1021 cacgacctgg tgaccacctg ccactacgcg gttgctatgg tgaagaaaag cagcgcgttc
     1081 cagttcaacc agctgaaggg caaacgtagc tgccacagcg gtgttagcaa aaccgacggc
     1141 tggaaggcgc tggtgaccgt tctggtggaa aagaaactgc tgagctggga tggtccggcg
     1201 aaagagagca tccagcgtgc tatgagcaag ttcttcagcg tgagctgcat tccgggtgct
     1261 acccagacca acctgtgcaa gcagtgcaaa ggcgaggaag gcaagaactg caaaaacagc
     1321 cacgacgaac cgtactacgg taactacggc gcgttccgtt gcctgaagga agacatgggt
     1381 gacgttgcgt tcctgcgtag caccgctctg agcgacgagc acagcgaagt gtacgagctg
     1441 ctgtgcccgg ataacacccg taagccgctg aacaagtaca aagagtgcaa cctgggtacc
     1501 gttccggctg gcaccgtggt tacccgtaag atcagcgaca aaaccgaaga catcaacaac
     1561 ttcctgatgg aggcgcagaa gcgtcagtgc aaactgttca gcagcgctca cggcaaggac
     1621 ctgatgttcg acgatagcac cctgcagctg gctctgctga gcagcgaggt tgatgctttc
     1681 ctgtacctgg gtgtgaaact gttccacgcg atgaaggctc tgaccggcga cgctcacctg
     1741 ccgagcaaga acaaagtgcg ttggtgcacc atcaacaagc tggaaaagat gaaatgcgac
     1801 gattggagcg cggttagcgg tggcgcgatc gcttgcaccg aggctagctg cccgaagggc
     1861 tgcgtgaaac agatcctgaa gggcgaagcg gacgctgtta agctggaagt gcagtacatg
     1921 tacgaggcgc tgatgtgcgg tctgctgccg gctgttgagg aataccacaa caaagacgat
     1981 ttcggtccgt gcaagacccc aggtagcccg tacaccgact tcggcaccct gcgtgcggtt
     2041 gctctggtga agaaaagcaa caaagacatc aactggaaca acatcaaggg caagaagagc
     2101 tgccacaccg gtgtgggcga catcgctggc tgggttatcc cggtgagcct gatccgtcgt
     2161 cagaacgata acagcgacat cgatagcttc ttcggcgaga gctgcgcgcc gggcagcgac
     2221 accaaaagca acctgtgcaa gctgtgcatc ggtgacccga agaacagcgc ggctaacacc
     2281 aaatgcagcc tgagcgacaa ggaagcgtac tacggtaacc agggcgcttt ccgttgcctg
     2341 gtggagaaag gtgacgttgc tttcgtgccg cacaccgtgg ttttcgaaaa caccgatggc
     2401 aagaacccgg cggtttgggc taagaacctg aaaagcgaag acttcgagct gctgtgcctg
     2461 gatggtagcc gtgcgccggt gagcaactac aagagctgca aactgagcgg tatcccgccg
     2521 ccggctatcg ttacccgtga ggaaagcatc agcgacgtgg ttcgtatcgt ggctaaccag
     2581 cagagcctgt acggtcgtaa aggcttcgag aaggatatgt tccagctgtt cagcagcaac
     2641 aagggtaaca acctgctgtt caacgacaac acccagtgcc tgatcaccag cgatcgtcag
     2701 ccgaaagaca tcatggaaga ttacttcggc aagccgtact acaccaccgt gtacggcgcg
     2761 tcccgctccg ctatgtcctc cgaactgatt tccgcctgta ccattaaaca ctgctcgaat
     2821 tctttggaag ttttgtttca aggtccaact gctgccgccg ctgtgagcaa gggcgaggag
     2881 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
     2941 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
     3001 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
     3061 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     3121 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     3181 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     3241 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     3301 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     3361 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     3421 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccaag
     3481 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     3541 gccgggatca ctctcggcat ggacgagctg tacaagtccg gaggtggtca ccatcaccat
     3601 caccaccatc accatcacta atagaagctt gtcgagaagt actagaggat cataatcagc
     3661 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     3721 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     3781 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     3841 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctggatctg atcactgctt
     3901 gagcctagga gatccgaacc agataagtga aatctagttc caaactattt tgtcattttt
     3961 aattttcgta ttagcttacg acgctacacc cagttcccat ctattttgtc actcttccct
     4021 aaataatcct taaaaactcc atttccaccc ctcccagttc ccaactattt tgtccgccca
     4081 cagcggggca tttttcttcc tgttatgttt ttaatcaaac atcctgccaa ctccatgtga
     4141 caaaccgtca tcttcggcta ctttttctct gtcacagaat gaaaattttt ctgtcatctc
     4201 ttcgttatta atgtttgtaa ttgactgaat atcaacgctt atttgcagcc tgaatggcga
     4261 atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     4321 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     4381 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     4441 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     4501 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttcttaat
     4561 agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta ttcttttgat
     4621 ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa
     4681 tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact tttcggggaa
     4741 atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     4801 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     4861 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     4921 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt
     4981 acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt
     5041 ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg
     5101 ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact
     5161 caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg
     5221 ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga
     5281 aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg
     5341 aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa
     5401 tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac
     5461 aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc
     5521 cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca
     5581 ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga
     5641 gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta
     5701 agcattggta actgtcagac caagtttact catatatact ttagattgat ttaaaacttc
     5761 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     5821 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     5881 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     5941 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     6001 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     6061 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     6121 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     6181 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     6241 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     6301 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     6361 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     6421 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     6481 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     6541 cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc
     6601 gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga
     6661 tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatagac cagccgcgta
     6721 acctggcaaa atcggttacg gttgagtaat aaatggatgc cctgcgtaag cgggtgtggg
     6781 cggacaataa agtcttaaac tgaacaaaat agatctaaac tatgacaata aagtcttaaa
     6841 ctagacagaa tagttgtaaa ctgaaatcag tccagttatg ctgtgaaaaa gcatactgga
     6901 cttttgttat ggctaaagca aactcttcat tttctgaagt gcaaattgcc cgtcgtatta
     6961 aagaggggcg tggccaaggg catggtaaag actatattcg cggcgttgtg acaatttacc
     7021 gaacaactcc gcggccggga agccgatctc ggcttgaacg aattgttagg tggcggtact
     7081 tgggtcgata tcaaagtgca tcacttcttc ccgtatgccc aactttgtat agagagccac
     7141 tgcgggatcg tcaccgtaat ctgcttgcac gtagatcaca taagcaccaa gcgcgttggc
     7201 ctcatgcttg aggagattga tgagcgcggt ggcaatgccc tgcctccggt gctcgccgga
     7261 gactgcgaga tcatagatat agatctcact acgcggctgc tcaaacttgg gcagaacgta
     7321 agccgcgaga gcgccaacaa ccgcttcttg gtcgaaggca gcaagcgcga tgaatgtctt
     7381 actacggagc aagttcccga ggtaatcgga gtccggctga tgttgggagt aggtggctac
     7441 gtctccgaac tcacgaccga aaagatcaag agcagcccgc atggatttga cttggtcagg
     7501 gccgagccta catgtgcgaa tgatgcccat acttgagcca cctaactttg ttttagggcg
     7561 actgccctgc tgcgtaacat cgttgctgct ccataacatc aaacatcgac ccacggcgta
     7621 acgcgcttgc tgcttggatg cccgaggcat agactgtaca aaaaaacagt cataacaagc
     7681 catgaaaacc gccactgcgc cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt
     7741 gcgtgagcgc atacgctact tgcattacag tttacgaacc gaacaggctt atgtcaactg
     7801 ggttcgtgcc ttcatccgtt tccacggtgt gcgtcacccg gcaaccttgg gcagcagcga
     7861 agtcgaggca tttctgtcct ggctggcgaa cgagcgcaag gtttcggtct ccacgcatcg
     7921 tcaggcattg gcggccttgc tgttcttcta cggcaaggtg ctgtgcacgg atctgccctg
     7981 gcttcaggag atcggaagac ctc
//
