LOCUS       sequence-69950-e        6613 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Shuttle vector E. coli/B.meg.; encoding the signal peptide for 
            protein secretion of the Bacillus megaterium protein Pac under 
            control of the xylose inducible promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6613)
  AUTHORS   Stammen S, Muller BK, Korneli C, Biedendieck R, Gamer M, Franco-Lara
            E, Jahn D
  TITLE     High-yield intra- and extracellular protein production using 
            Bacillus megaterium.
  JOURNAL   Appl Environ Microbiol. 2010 Jun;76(12):4037-46. doi: 
            10.1128/AEM.00431-10. Epub 2010 Apr 30.
  PUBMED    20435764
REFERENCE   2  (bases 1 to 6613)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1128/AEM.00431-10"; journalName: "Appl Environ Microbiol"; date:
            "2010-06"; volume: "76"; issue: "12"; pages: "4037-46"
COMMENT     Sequence Label: pSPPac-hp
FEATURES             Location/Qualifiers
     source          1..6613
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             716..733
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     rep_origin      complement(874..1462)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(954..973)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1633..2424,2425..2493))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2256..2275
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2494..2598)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2666..2684
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     CDS             complement(2762..4138)
                     /codon_start=1
                     /gene="Bacillus subtilis tetL"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MNTSYSQSNLRHNQILIWLCILSFFSVLNEMVLNVSLPDIANDFN
                     KPPASTNWVNTAFMLTFSIGTAVYGKLSDQLGIKRLLLFGIIINCFGSVIGFVGHSFFS
                     LLIMARFIQGAGAAAFPALVMVVVARYIPKENRGKAFGLIGSIVAMGEGVGPAIGGMIA
                     HYIHWSYLLLIPMITIITVPFLMKLLKKEVRIKGHFDIKGIILMSVGIVFFMLFTTSYS
                     ISFLIVSVLSFLIFVKHIRKVTDPFVDPGLGKNIPFMIGVLCGGIIFGTVAGFVSMVPY
                     MMKDVHQLSTAEIGSVIIFPGTMSVIIFGYIGGILVDRRGPLYVLNIGVTFLSVSFLTA
                     SFLLETTSWFMTIIIVFVLGGLSFTKTVISTIVSSSLKQQEAGAGMSLLNFTSFLSEGT
                     GIAIVGGLLSIPLLDQRLLPMEVDQSTYLYSNLLLLFSGIIVISWLVTLNVYKHSQRDF
                     "
     CDS             complement(4338..5342)
                     /codon_start=1
                     /gene="repB"
                     /product="RepB replication protein"
                     /label=repB
                     /note="from Enterococcus faecalis plasmid pAM-alpha-1"
                     /db_xref="GI:22652809"
                     /protein_id="AAN03827.1"
                     /translation="MGVSFNIMCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDY
                     FELLHILEFKKAERVKDCAEILEYKQNRETGERKLYRVWFCKSRLCPMCNWRRAMKHGI
                     QSQKVVAEVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLV
                     GFMRATEVTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPN
                     VKVQMIRPKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLIS
                     YGGLLKEIHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
ORIGIN
        1 aactagacaa gtcagaagtc ttctcgagaa taatatttcc ttctaagtcg gttagaattc
       61 cgttaagata gtcgactcct atatcaatac caatcgagta gcctgcattc ttattaaaaa
      121 caagcattac aggtcttctg ccgcctctag attgccctgc cccaatttca aaaataaaat
      181 ctttttcaag cagtgtattt acttgagagg agacagtaga cttgtttaat cctgtaatct
      241 cagagagagt tgccctggag acaggggagt tcttcaaaat ttcatctaat attaattttt
      301 gattcatttt ttttactaaa gcttgatctg caatttgaat aataaccact cctttgttta
      361 tccaccgaac taagttggtg ttttttgaag cttgaattag atatttaaaa gtatcatatc
      421 taatattata actaaatttt ctaaaaaaaa cattgacata aacatttaat taatatatga
      481 tgagataaag ttagtttatt ggataaacaa actaactcaa ttaagatagt tgatggataa
      541 acttgttcac ttaaatcaag gaggtgaatg tacaatgaag acgaagtggc taatatcagt
      601 cataatccta tttgttttca tttttcctca aaatctagtt tttgctggga ctagttcgaa
      661 gatctccgga gctcccggga tccggtaccg gccgcatgcc ggcctgagag gatcgcatca
      721 ccatcaccat cactaaccgg tccaagaatt ggagccaatc aattcttgcg gagaactgtg
      781 aatgcgcaaa ccaacccttg gcagaacata tccatcgcgt ccgccatctc cagcagccgc
      841 acgcggcgca tctcgggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
      901 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
      961 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     1021 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     1081 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     1141 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     1201 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     1261 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     1321 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     1381 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     1441 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     1501 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     1561 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     1621 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     1681 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     1741 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     1801 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     1861 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     1921 tagtttgcgc aacgttgttg ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg
     1981 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     2041 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     2101 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     2161 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     2221 gcgaccgagt tgctcttgcc cggcgtcaac acgggataat accgcgccac atagcagaac
     2281 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     2341 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     2401 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     2461 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     2521 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     2581 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     2641 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtcttcaaga
     2701 attcctgtta taaaaaaagg atcaattttg aactctctcc caaagttgat cccttaacga
     2761 tttagaaatc cctttgagaa tgtttatata cattcaaggt aaccagccaa ctaatgacaa
     2821 tgattcctga aaaaagtaat aacaaattac tatacagata agttgactga tcaacttcca
     2881 taggtaacaa cctttgatca agtaagggta tggataataa accacctaca attgcaatac
     2941 ctgttccctc tgataaaaag ctggtaaagt taagcaaact cattccagca ccagcttcct
     3001 gctgtttcaa gctacttgaa acaattgttg atataactgt tttggtgaac gaaagcccac
     3061 ctaaaacaaa tacgattata attgtcatga accatgatgt tgtttctaaa agaaaggaag
     3121 cagttaaaaa gctaacagaa agaaatgtaa ctccgatgtt taacacgtat aaaggacctc
     3181 ttctatcaac aagtatccca ccaatgtagc cgaaaataat gacactcatt gttccaggga
     3241 aaataattac acttccgatt tcggcagtac ttagctggtg aacatctttc atcatataag
     3301 gaaccataga gacaaaccct gctactgttc caaatataat tcccccacaa agaactccaa
     3361 tcataaaagg tatatttttc cctaatccgg gatcaacaaa aggatctgtt actttcctga
     3421 tatgttttac aaatatcagg aatgacagca cgctaacgat aagaaaagaa atgctatatg
     3481 atgttgtaaa caacataaaa aatacaatgc ctacagacat tagtataatt cctttgatat
     3541 caaaatgacc ttttatcctt acttctttct ttaataattt cataagaaac ggaacagtga
     3601 taattgttat cataggaatg agtagaagat aggaccaatg aatataatgg gctatcattc
     3661 caccaatcgc tggaccgact ccttctccca tggctactat cgatccaata agaccaaatg
     3721 ctttacccct attttccttt ggaatatagc gcgcaactac aaccattacg agtgctggaa
     3781 atgcagctgc accagcccct tgaataaaac gagccataat aagtaaggaa aagaaagaat
     3841 ggccaacaaa cccaattacc gacccgaaac aatttattat aattccaaat aggagtaacc
     3901 ttttgatgcc taattgatca gatagctttc catatacagc tgttccaatg gaaaaggtta
     3961 acataaaggc tgtgttcacc cagtttgtac tcgcaggtgg tttattaaaa tcatttgcaa
     4021 tatcaggtaa tgagacgttc aaaaccattt catttaatac gctaaaaaaa gataaaatgc
     4081 aaagccaaat taaaatttgg ttgtgtcgta aattcgattg tgaataggat gtattcacat
     4141 ttcaccctcc aataatgagg gcagacgtag tttatagggt taatgatacg cttccctctt
     4201 ttaattgaac cctgttacat tcattacact tcataattaa ttcctcctaa acttgattaa
     4261 aacattttac cacatataaa ctaagtttta aattcagtat ttcatcactt atacaacaat
     4321 atggcccgtt tgttgaacta ctctttaata aaataatttt tccgttccca attccacatt
     4381 gcaataatag aaaatccatc ttcatcggct ttttcgtcat catctgtatg aatcaaatcg
     4441 ccttcttctg tgtcatcaag gtttaatttt ttatgtattt cttttaacaa accaccatag
     4501 gagattaacc ttttacggtg taaaccttcc tccaaatcag acaaacgttt caaattcttt
     4561 tcttcatcat cggtcataaa atccgtatcc tttacaggat attttgcagt ttcgtcaatt
     4621 gccgattgta tatccgattt atatttattt ttcggtcgaa tcatttgaac ttttacattt
     4681 ggatcatagt ctaatttcat tgcctttttc caaaattgaa tccattgttt ttgattcacg
     4741 tagttttctg tattcttaaa ataagttggt tccacacata ccaatacatg catgtgctga
     4801 ttataagaat tatctttatt atttattgtc acttccgttg cacgcataaa accaacaaga
     4861 tttttattaa tttttttata ttgcatcatt cggcgaaatc cttgagccat atctgacaaa
     4921 ctcttattta attcttcgcc atcataaaca tttttaactg ttaatgtgag aaacaaccaa
     4981 cgaactgttg gcttttgttt aataacttca gcaacaacct tttgtgactg aatgccatgt
     5041 ttcattgctc tcctccagtt gcacattgga caaagcctgg atttacaaaa ccacactcga
     5101 tacaactttc tttcgcctgt ttcacgattt tgtttatact ctaatatttc agcacaatct
     5161 tttactcttt cagccttttt aaattcaaga atatgcagaa gttcaaagta atcaacatta
     5221 gcgattttct tttctctcca tggtctcact tttccacttt ttgtcttgtc cactaaaacc
     5281 cttgattttt catctgaata aatgctacta ttaggacaca taatattaaa agaaaccccc
     5341 atctatttag ttatttgttt ggtcacttat aactttaaca gatggggttt ttctgtgcaa
     5401 ccaattttaa gggttttcaa tactttaaaa cacatacata ccaacacttc aacgcacctt
     5461 tcagcaacta aaataaaaat gacgttattt ctatatgtat caagataaga aagaacaagt
     5521 tcaaaaccat caaaaaaaga caccttttca ggtgcttttt ttattttata aactcattcc
     5581 ctgatctcga cttcgttctt tttttacctc tcggttatga gttagttcaa attcgttctt
     5641 tttaggttct aaatcgtgtt tttcttggaa ttgtgctgtt ttatccttta ccttgtctac
     5701 aaacccctta aaaacgtttt taaaggcttt taagccgtct gtacgttcct taagatcaac
     5761 gtgatatagg tttgctaacc tttgcgttca cttaactaac ttataggggt aacacttaaa
     5821 aaagaatcaa taacgataga aaccgctcct aaagcaggtg cattttttcc taacgaagaa
     5881 ggcaatagtt cacatttatt gtctaaatga gaatggactc tagaagaaac ttcgttttta
     5941 atcgtattta aaacaatggg atgagattca attatatgat ttctcaagat aacagcttct
     6001 atatcaaatg tattaaggat attggttaat ccaattccga tataaaagcc aaagttttga
     6061 agtgcattta acatttctac atcattttta tttgcgcgtt ccacaatctc ttttcgagaa
     6121 atattctttt cttctttaga gagcgaagcc agtaacgctt tttcagaagc atataattcc
     6181 caacagcctc gatttccaca gctgcatttg ggtccattaa aatctatcgt catatgaccc
     6241 atttccccag aaaaaccctg aacaccttta tacaattcgt tgttaataac aagtccagtt
     6301 ccaattccga tattaatact gatgtaaacg atgttttcat agttttttgt cataccaaat
     6361 actttttcac cgtatgctcc tgcattagct tcattttcaa caaaaaccgg aacattaaac
     6421 tcactctcaa ttaaaaactg caaatctttg atattccaat ttaagttagg catgaaaata
     6481 atttgctgat gacgatctac aaggcctgga acacaaattc ctattccgac tagaccataa
     6541 ggggactcag gcatatgggt tacaaaacca tgaataagtg caaataaaat ctcttttact
     6601 tcactagcgg aag
//
