LOCUS       sequence-434369-        7442 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of secretory BT0294 SP-NanoLuc via 
            BfP1E6-RBS8 promoter-RBS pair in Bacteroides species..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7442)
  AUTHORS   Yeh YH, Kelly VW, Rahman Pour R, Sirk SJ
  TITLE     A molecular toolkit for heterologous protein secretion across 
            Bacteroides species.
  JOURNAL   Nat Commun. 2024 Nov 11;15(1):9741. doi: 10.1038/s41467-024-53845-7.
  PUBMED    39528443
REFERENCE   2  (bases 1 to 7442)
  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 
            Commun."; date: "2024-11-11"; volume: "15(1):9741. doi"; pages: " 
            10.1038/s41467-024-53845-7"
COMMENT     Sequence Label: pYHY2
FEATURES             Location/Qualifiers
     source          1..7442
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             2888..3397
                     /codon_start=1
                     /product="NanoLuc(R) luciferase"
                     /label=Nluc
                     /note="human codon-optimized"
                     /translation="VFTLEDFVGDWRQTAGYNLDQVLEQGGVSSLFQNLGVSVTPIQRI
                     VLSGENGLKIDIHVIIPYEGLSGDQMGQIEKIFKVVYPVDDHHFKVILHYGTLVIDGVT
                     PNMIDYFGRPYEGIAVFDGKKITVTGTLWNGNKIIDERLINPDGSLLFRVTINGVTGWR
                     LCERILA"
     CDS             3419..3484
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     terminator      3511..3582
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(3689..4480,4481..4549))
                     /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     4312..4331
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4550..4654)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      4757..5345
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5246..5265
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     oriT            5505..5614
                     /note="incP origin of transfer"
     CDS             5647..6018
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
ORIGIN
        1 tgatagcaca ctagcaccct aaagagtgcg tgtgtaaggg gacaagcccc cttaaccccc
       61 tgtcagccgg ctgtcaccgg ctgttatcaa tcttcaaagt cgggtggttg tcaagatgat
      121 ttttttggtt tggtgtcgtc tttttttaag ctgccgcata acggtttgcg attgtcacaa
      181 aaatgttaac ttatgggagc aacaagtatt catgtacaag cagtgaagcc ggggagcgag
      241 attcacaact ttagggaaaa agagttggac tatgttcgtc ccgaacttag tcatttgaat
      301 gaaagctggg ttggagatag catttcccat cggctggaga gtgcaaaaca aagatatctc
      361 gatacggttg ggcagaagat gcaggctaaa gccgcaccca tacgagaggg agtaatagta
      421 atcaaacaag aaaccactat gcaggaactc cagcagtttg ccacggtctg caaagaacgt
      481 ttcggtatcg aagcgtttca aatccatata cacaaggacg aaggatacat gaacgcaaag
      541 cagtggacac ctaaccttca tgcccatgta gttttcgatt ggacgcagcc gaatgggaaa
      601 agtgtgcgtt tatcgcgtga tgacatggca gaactccaaa ccatagcatc tgaaacactg
      661 ggcatggaac gtggcgtttc ttctgaccgc aaacacttat cggctatgca gtacaagacc
      721 gaatgtgcga aagaacagct tcaggaacta tccaacgata tatccagtgc attagacaaa
      781 tacaaggacg cacaaaacca gcttcttcag cttcagaagg aactacgttc cattgaaaca
      841 aagaagaacg tccaaaaact catttctaag gcttcagaga agttttacgg cttgattgga
      901 aaaacagtca acgataggga aaaggatgcc ttaaaagcca aagtaaagac tttagagggg
      961 gaaaatgaac aactatccga tagactggga aaggctatac ttgaaaaaga gcgaaacagc
     1021 acaaaggcat tcaaagccga gaatgacaag gagtattata gacaacaaat ggacaacgca
     1081 aggacaacca gtaacctatt aaggacggag aaccaaaaac taaaagccga aaccaaggag
     1141 ttgaaaaaag aacttggtaa gatgaaagac ttgttcaatt ccgaacaact ggaggcttta
     1201 aggcatcatt tcccaaacat atcaaaagct atggaagagg ggaaagacct attcaagcaa
     1261 atcaccagaa gcagaggttt cggcatgggt atgtaacccc cccccgcccc caaaggggga
     1321 gcgaccaaac ggcagcctct ctcaatggag tgttacatcc cgtaaagagg ggctttgcag
     1381 ggggtggact cagaaagatg agaatagatg actattgtag ttgaaacaca tagaaagttg
     1441 ctgatataca gaccgatacg catatcggga tgaaccatga gtacgttctt ttctcaaaaa
     1501 acataaatat tcgaaaagag atgcaataaa ttaaggagag gttataatga acaaagtaaa
     1561 tataaaagat agtcaaaatt ttattacttc aaaatatcac atagaaaaaa taatgaattg
     1621 cataagttta gatgaaaaag ataacatctt tgaaataggt gcagggaaag gtcattttac
     1681 tgctggattg gtaaagagat gtaattttgt aacggcgata gaaattgatt ctaaattatg
     1741 tgaggtaact cgtaataagc tcttaaatta tcctaactat caaatagtaa atgatgatat
     1801 actgaaattt acatttccta gccacaatcc atataaaata tttggcagca taccttacaa
     1861 cataagcaca aatataattc gaaaaattgt ttttgaaagt tcagccacaa taagttattt
     1921 aatagtggaa tatggttttg ctaaaatgtt attagataca aacagatcac tagcattgct
     1981 gttaatggca gaggtagata tttctatatt agcaaaaatt cctaggtatt atttccatcc
     2041 aaaacctaaa gtggatagca cattaattgt attaaaaaga aagccagcaa aaatggcatt
     2101 taaagagaga aaaaaatatg aaacttttgt aatgaaatgg gttaacaaag agtacgaaaa
     2161 actgtttaca aaaaatcaat ttaataaagc tttaaaacat gcgagaatat atgatataaa
     2221 caatattagt ttcgaacaat ttgtatcgct atttaatagt tataaaatat ttaacggcta
     2281 aaaacaatag gccacatgca actgtaaatg tttacgcgtc ctcggtacca acaaatactt
     2341 tcaggacgat gtaaagtcct gaaaaatcaa agctggccaa tgagacgacg gggtcatcac
     2401 ggctcatcat gcgccaaaca aatgtgtgca atacacgctc ggatgactgc atgatgaccg
     2461 cactgactgg ggacagcaga tccacctaag cctgtgagag aagcagacac ccgacagatc
     2521 aaggcagtta actagtgcac tgcagtggcg ataaaacgaa aggctcagtc gaaagactgg
     2581 gtctttcgtt ttacaattgg gctacctttt ttttgttttg tttgcaatgg ttaatctatt
     2641 gttaaaattt aaagtttcac ttgaactttc aaataatgtt cttatatttg cagtgtcgaa
     2701 agaaacaaag taggactgat cgtccatcaa tttaaaattt aaaataatga gaaatttaaa
     2761 atggttgtat gcatgctctt tggcgatagc cttcggtgtg ttgtcttttg taacagtttc
     2821 ctgtcatgat gatgacgatg aaccaaagca agaaccaggt gaagtgattg aaactcccgc
     2881 tccggttgtc ttcacactcg aagatttcgt tggggactgg cgacagacag ccggctacaa
     2941 cctggaccaa gtccttgaac agggaggtgt gtccagtttg tttcagaatc tcggggtgtc
     3001 cgtaactccg atccaaagga ttgtcctgag cggtgaaaat gggctgaaga tcgacatcca
     3061 tgtcatcatc ccgtatgaag gtctgagcgg cgaccaaatg ggccagatcg aaaaaatttt
     3121 taaggtggtg taccctgtgg atgatcatca ctttaaggtg atcctgcact atggcacact
     3181 ggtaatcgac ggggttacgc cgaacatgat cgactatttc ggacggccgt atgaaggcat
     3241 cgccgtgttc gacggcaaaa agatcactgt aacagggacc ctgtggaacg gcaacaaaat
     3301 tatcgacgag cgcctgatca accccgacgg ctccctgctg ttccgagtaa ccatcaacgg
     3361 agtgaccggc tggcggctgt gcgaacgcat tctggcggga tcttctggct cgagcggcga
     3421 ctacaaagac catgacggtg attataaaga tcatgacatc gattacaagg atgacgatga
     3481 caagtgaata gatcaccatt aaccaggcat caaataaaac gaaaggctca gtcgaaagac
     3541 tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tctacagcgg ccgcgattat
     3601 caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa
     3661 gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct
     3721 cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta
     3781 cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcgg gacccacgct
     3841 caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg
     3901 gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa
     3961 gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt
     4021 cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta
     4081 catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca
     4141 gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta
     4201 ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct
     4261 gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg
     4321 cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac
     4381 tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact
     4441 gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa
     4501 atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt
     4561 ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat
     4621 gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg
     4681 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     4741 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     4801 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     4861 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt
     4921 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     4981 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     5041 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     5101 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     5161 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     5221 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     5281 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     5341 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     5401 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtgcggcc gcccctccaa
     5461 agggttcgtg tagactttcc ttggtgtatc caacggcgtc agccgggcag gataggtgaa
     5521 gtaggcccac ccgcgagcgg gtgttccttc ttcactgtcc cttattcgca cctggcggtg
     5581 ctcaacggga atcctgctct gcgaggctgg ccggctaccg ccggcgtaac agatgagggc
     5641 aagcggatgg ctgatgaaac caagccaacc aggaagggca gcccacctat caaggtgtac
     5701 tgccttccag acgaacgaag agcgattgag gaaaaggcgg cggcggccgg catgagcctg
     5761 tcggcctacc tgctggccgt cggccagggc tacaaaatca cgggcgtcgt ggactatgag
     5821 cacgtccgcg agctggcccg catcaatggc gacctgggcc gcctgggcgg cctgctgaaa
     5881 ctctggctca ccgacgaccc gcgcacggcg cggttcggtg atgccacgat cctcgccctg
     5941 ctggcgaaga tcgaagagaa gcaggacgag cttggcaagg tcatgatggg cgtggtccgc
     6001 ccgagggcag agccatgact tttttagccg ctaaaacggc cggggggtgc gcgtgattgc
     6061 caagcacgtc cccataacgc atagcgctcc ccgaccgatg atttttaagt gactgatttt
     6121 gtgctgtttt gggggtatat taagaataaa agaaatagaa taagttaagt acttgataca
     6181 caatataggg cattttccat attggaaatt ctcattttcc aatctagaaa ataccgattt
     6241 cctaatatga tactaaatag gaaaataata tttcccttaa tattgttttt atggaaaata
     6301 atagtttact ttgtggagaa taatatttcc caaaaacata tcaaaatgga aaataaaaaa
     6361 gcagttaagt taaccgattt tcaaaagaac gaagaaaatc cttttatgaa acaagctata
     6421 gaaggtattg aaaatcatgt tgttaaaaag tataagagta atagtggtgg cgataagaga
     6481 gctgtagtag ctttagccga cactgaaact ggagaagtgt ttaagacttc gtttatccgt
     6541 caaatagaag tagatgaaga acaattcact aaattgtatc tttctaactt tgctgcattc
     6601 tttgacctat cacaagcagc tattcgggtt tttggttact ttatgacctg catgaaaccc
     6661 aaaaatgatt taatcatttt caatagaaaa aaatgcctag aatataccaa atacaaaaca
     6721 gacaaagccg tttataaagg acttgcagaa cttgtaaaag ctgaaatcat agcccgagga
     6781 ccagccgata atctttggtt tattaatcct ctgatagtat tcaatggtga ccgagtgaca
     6841 tttgctaaaa catacgttcg gaaaaagact ttagctgccc aaaagaaaga agaagcagag
     6901 aaacgacaat tatcacttgg ctttgatgaa cagtaacact ccattgagtg aagctgccgt
     6961 ttggtcgctc cccctttggg ggcggggggg gatagataaa gttcctctat gtaaagttat
     7021 aatgggggat gaaaggcaag gttcgctaac cttaccccga ttgtttatgc ttcccagcat
     7081 tcaatctatc cccgtttatg ggtttctctt tcaccctaca aagataacct catgggggaa
     7141 aaatgtccaa gaatatgggg taaaaactat caagtcggta gaaaataagt atctttgaag
     7201 tacattttta gtagaggtac tggtgtgcct aggcgaagca gataggcaaa aataaagccg
     7261 ctggagatgt cggatgtccg tcaattaact ccagcggctt gtattttggt aactctcccg
     7321 tatcggtcag ctaccgtttg gtcagcaaca aaggtagtac tttattatcg caaatccaaa
     7381 tgtttttccc tttatttgtg catttacaaa acattcacta acttcgcacc tatgcagtat
     7441 gg
//
