LOCUS       sequence-444799-        5398 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To produce caveospheres with periplasmically encapsulated mScarlet.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5398)
  AUTHORS   T. D. Luong, N. Martel, J. Rae, H. P. Lo, Y-W. Lim, Y. Wu, K-A. 
            McMahon, N. Fletcher, K. Thurecht, A.P.R. Johnston, N. Ariotti, T. 
            E. Hall, R.G. Parton
  TITLE     A modular encapsulation system for precision delivery of proteins, 
            nucleic acids and therapeutics
  JOURNAL   bioRxiv
REFERENCE   2  (bases 1 to 5398)
  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: "bioRxiv"
COMMENT     Sequence Label: pNMTMA_MBP_Z_Cav1_H6_Periplasmic_mScarlet
FEATURES             Location/Qualifiers
     source          1..5398
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(14..32)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        100..203
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(205..273,274..1065)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MRIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(423..442)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1236..1824
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1725..1744
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1978..1995
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    2010..2149
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(2075..2094)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2235..2257
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2310..2329
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2310..2328
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             2402..2424
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2432..3532
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRLTVDEALKDAQT"
     primer_bind     3506..3529
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             3551..3571
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3593..3613
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             3629..3802
                     /codon_start=1
                     /product="IgG-binding unit of Staphylococcus aureus protein
                     A"
                     /label=ProtA
                     /translation="VDNKFNKEQQNAFYEILHLPNLNEEQRNAFIQSLKDDPSQSANLL
                     AEAKKLNDAQAPK"
     CDS             4343..4354
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             4355..4372
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     sig_peptide     4424..4486
                     /codon_start=1
                     /product="signal peptide from the E. coli outer membrane 
                     protein OmpA"
                     /label=OmpA signal peptide
                     /translation="MKKTAIAIAVALAGFATVAQA"
     primer_bind     complement(5306..5324)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5320..5367
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 tgaagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
       61 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
      121 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
      181 caataatatt gaaaaaggaa gagcatgcgt attcaacatt tccgtgtcgc ccttattccc
      241 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
      301 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
      361 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
      421 ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc
      481 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
      541 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
      601 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
      661 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
      721 aacgacgagc gtgacaccac gatgcctgca gcaatggcaa caacgttgcg caaactatta
      781 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
      841 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
      901 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
      961 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     1021 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaaatcga tgaccaagtt
     1081 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     1141 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     1201 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     1261 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     1321 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     1381 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     1441 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     1501 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     1561 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     1621 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     1681 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     1741 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     1801 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     1861 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     1921 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     1981 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     2041 tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     2101 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     2161 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     2221 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     2281 cgcgaggcag gatctcgatc ccgcgaaatt aatacgactc actataggga gaccacaacg
     2341 gtttccctct agtgccggct ccggagagct gcaattgcaa ttcgctagct cttgaaataa
     2401 ttttgtttaa ctttaagaag gagatataaa tatgaaaatc gaagaaggta aactggtaat
     2461 ctggattaac ggcgataaag gctataacgg tctcgctgaa gtcggtaaga aattcgagaa
     2521 agataccgga attaaagtca ccgttgagca tccagataaa ctggaagaga aattcccaca
     2581 ggttgcggca actggcgatg gccctgacat tatcttctgg gcacacgacc gctttggtgg
     2641 ctacgctcaa tctggcctgt tggctgaaat caccccggac aaagcgttcc aggacaagct
     2701 gtatccgttt acctgggatg ccgtacgtta caacggcaag ctgattgctt acccgatcgc
     2761 tgttgaagcg ttatcgctga tttataacaa agatctgctg ccgaacccgc caaaaacctg
     2821 ggaagagatc ccggcgctgg ataaagaact gaaagcgaaa ggtaagagcg cgctgatgtt
     2881 caacctgcaa gaaccgtact tcacctggcc gctgattgct gctgacgggg gttatgcgtt
     2941 caagtatgaa aacggcaagt acgacattaa agacgtgggc gtggataacg ctggcgcgaa
     3001 agcgggtctg accttcctgg ttgacctgat taaaaacaaa cacatgaatg cagacaccga
     3061 ttactccatc gcagaagctg cctttaataa aggcgaaaca gcgatgacca tcaacggccc
     3121 gtgggcatgg tccaacatcg acaccagcaa agtgaattat ggtgtaacgg tactgccgac
     3181 cttcaagggt caaccatcca aaccgttcgt tggcgtgctg agcgcaggta ttaacgccgc
     3241 cagtccgaac aaagagctgg caaaagagtt cctcgaaaac tatctgctga ctgatgaagg
     3301 tctggaagcg gttaataaag acaaaccgct gggtgccgta gcgctgaagt cttacgagga
     3361 agagttggcg aaagatccac gtattgccgc cactatggaa aacgcccaga aaggtgaaat
     3421 catgccgaac atcccgcaga tgtccgcttt ctggtatgcc gtgcgtactg cggtgatcaa
     3481 cgccgccagc ggtcgtctga ctgtcgatga agccctgaaa gacgcgcaga ctgcaagctc
     3541 ttctggttcc gaaaacctgt atttccaggg ctcccatatg tcttctggtt ccgaaaacct
     3601 gtatttccag ggctcccatc ccatggctgt agacaacaaa ttcaacaaag aacaacaaaa
     3661 cgcgttctat gagatcttac atttacctaa cttaaacgaa gaacaacgaa acgccttcat
     3721 ccaaagttta aaagatgacc caagccaaag cgctaacctt ttagcagaag ctaaaaagct
     3781 aaatgatgct caggcgccga aaggatccat gtctgggggc aaatacgtag actcggaggg
     3841 acatctctac acggtaccca tccgggaaca gggcaacatc tacaagccca acaacaaggc
     3901 catggcagac gagctgagcg agaagcaagt gtacgacgcg cacaccaagg agatcgacct
     3961 ggttaaccgc gaccctaaac acctcaacga tgacgtggtc aagattgact ttgaagatgt
     4021 gattgcagaa ccagaaggga cacacagttt tgacggcatt tggaaggcca gcttcaccac
     4081 cttcactgtg acgaaatact ggttttaccg cttgctgtct gccctctttg gcatcccgat
     4141 ggcactcatc tggggcattt acttcgccat tctctctttc ctgcacatct gggcagttgt
     4201 accatgcatt aagagcttcc tgattgagat tcagtgcatc agccgtgtct attccatcta
     4261 cgtccacacc gtctgtgacc cactctttga agctgttggg aaaatattca gcaatgtccg
     4321 catcaacatg cagaaagaaa caatcgaagg tcgtcatcat catcatcatc attaagaatt
     4381 cgccgctctt taacaattta tcagatccaa taggaggaac aatatgaaaa aaaccgcgat
     4441 tgcgattgcg gtggcgctgg cgggctttgc gaccgtggcg caggcgatgg cggaagtgat
     4501 ggtatctaaa ggagaggctg tcatcaaaga gtttatgaga tttaaagtcc acatggaggg
     4561 aagcatgaac ggtcatgagt ttgaaataga aggagaaggt gagggtcgac cttacgaagg
     4621 tacccagacc gcgaagttga aagttaccaa agggggcccc ttgccgttca gttgggacat
     4681 cctgtcccct caatttatgt acggaagcag ggcttttacg aaacatccgg ctgacattcc
     4741 cgattactat aagcaaagtt tccccgaggg ctttaagtgg gagcgcgtaa tgaattttga
     4801 ggatggcggt gctgttacag tcacacagga tacatccctc gaagacggaa cgttgatcta
     4861 caaagttaaa cttcgcggga ctaacttccc gcccgatggc cctgtgatgc aaaagaagac
     4921 tatgggatgg gaggccagca cggaaagact ctaccctgaa gatggcgtac ttaaaggtga
     4981 tataaaaatg gctctgcgcc tgaaggatgg aggtcgttac ctcgccgatt tcaagaccac
     5041 ctacaaagcc aaaaaacccg tgcagatgcc tggcgcatat aatgtagata ggaagctgga
     5101 catcactagc cataatgaag attacaccgt tgtagagcaa tacgaacgat cagagggccg
     5161 tcactcaacc ggcggtatgg atgaactcta taagtaatag ccgtcgacaa gcttgcggcc
     5221 gcactcgagt ctagattggt accacgcgtg cgcgctgatc cggctgctaa caaagcccga
     5281 aaggaagctg agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc
     5341 tctaaacggg tcttgagggg ttttttgctg aaaggaggaa ctatagccgg ataattct
//
