LOCUS       sequence-206087-        5771 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express periplasmic, outer membrane, or secreted proteins in E. coli
            - SufI signal (TAT) and His6 tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5771)
  AUTHORS   Casasanta MA, Slade DJ
  TITLE     A Vector Suite for the Overexpression and Purification of Tagged 
            Outer Membrane, Periplasmic, and Secreted Proteins in E. coli.
  JOURNAL   Methods Mol Biol. 2019;1960:123-138. doi: 
            10.1007/978-1-4939-9167-9_11.
  PUBMED    30798527
REFERENCE   2  (bases 1 to 5771)
  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: "2019"; volume: "1960:123-138. doi"; pages: " 
            10.1007/978-1-4939-9167-9_11"
COMMENT     Sequence Label: pDJSVT91
FEATURES             Location/Qualifiers
     source          1..5771
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..15,16..32,33..38)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     terminator      complement(213..260)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     256..274
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(345..362)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             462..484
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    499..523
                     /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)."
     primer_bind     complement(523..542)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(524..542)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(626..645)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        851..928
                     /gene="lacI"
                     /label=lacI promoter
     CDS             929..2011
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(948..967)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             3323..3514
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(3508..3530)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3616..3758
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3671..3690
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3773..3790)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3944..4532)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4024..4043)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4703..5494,5495..5563))
                     /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     5326..5345
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5564..5668)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5736..5754
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ttctcatgtt tgacagctta tcatcgataa gctttaatgc ggtagtttat cacagttaaa
       61 ttgctaacgc agtcaggcac cgtgtatgaa atctaacaat gcgctcatcg tcatcctcgg
      121 caccgtcacc ctggatgctg taggcatagg cttggttatg ccggtactgc cgggcctctt
      181 gcgggatatc cggatatagt tcctcctttc agcaaaaaac ccctcaagac ccgtttagag
      241 gccccaaggg gttatgctag ttattgctca gcggtggcag cagccaactc agcttccttt
      301 cgggctttgt tagcagccgg atcctcgagg gtacccgcgg ccgcgtggtg gtggtggtgg
      361 tgttgcccgg ctgcgctggc cttcagggga acagcgcctg cacaaagtgc aatccccgat
      421 gcctgaatga actgacgccg agagagtgat cccatggtat atctccttct taaagttaaa
      481 caaaattatt tctagagggg aattgttatc cgctcacaat tcccctatag tgagtcgtat
      541 taatttcgcg ggatcgagat ctcgatcctc tacgccggac gcatcgtggc cggcatcacc
      601 ggcgccacag gtgcggttgc tggcgcctat atcgccgaca tcaccgatgg ggaagatcgg
      661 gctcgccact tcgggctcat gagcgcttgt ttcggcgtgg gtatggtggc aggccccgtg
      721 gccgggggac tgttgggcgc catctccttg catgcaccat tccttgcggc ggcggtgctc
      781 aacggcctca acctactact gggctgcttc ctaatgcagg agtcgcataa gggagagcgt
      841 cgagatcccg gacaccatcg aatggcgcaa aacctttcgc ggtatggcat gatagcgccc
      901 ggaagagagt caattcaggg tggtgaatgt gaaaccagta acgttatacg atgtcgcaga
      961 gtatgccggt gtctcttatc agaccgtttc ccgcgtggtg aaccaggcca gccacgtttc
     1021 tgcgaaaacg cgggaaaaag tggaagcggc gatggcggag ctgaattaca ttcccaaccg
     1081 cgtggcacaa caactggcgg gcaaacagtc gttgctgatt ggcgttgcca cctccagtct
     1141 ggccctgcac gcgccgtcgc aaattgtcgc ggcgattaaa tctcgcgccg atcaactggg
     1201 tgccagcgtg gtggtgtcga tggtagaacg aagcggcgtc gaagcctgta aagcggcggt
     1261 gcacaatctt ctcgcgcaac gcgtcagtgg gctgatcatt aactatccgc tggatgacca
     1321 ggatgccatt gctgtggaag ctgcctgcac taatgttccg gcgttatttc ttgatgtctc
     1381 tgaccagaca cccatcaaca gtattatttt ctcccatgaa gacggtacgc gactgggcgt
     1441 ggagcatctg gtcgcattgg gtcaccagca aatcgcgctg ttagcgggcc cattaagttc
     1501 tgtctcggcg cgtctgcgtc tggctggctg gcataaatat ctcactcgca atcaaattca
     1561 gccgatagcg gaacgggaag gcgactggag tgccatgtcc ggttttcaac aaaccatgca
     1621 aatgctgaat gagggcatcg ttcccactgc gatgctggtt gccaacgatc agatggcgct
     1681 gggcgcaatg cgcgccatta ccgagtccgg gctgcgcgtt ggtgcggata tctcggtagt
     1741 gggatacgac gataccgaag acagctcatg ttatatcccg ccgttaacca ccatcaaaca
     1801 ggattttcgc ctgctggggc aaaccagcgt ggaccgcttg ctgcaactct ctcagggcca
     1861 ggcggtgaag ggcaatcagc tgttgcccgt ctcactggtg aaaagaaaaa ccaccctggc
     1921 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     1981 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgta agttagctca
     2041 ctcattaggc accgggatct cgaccgatgc ccttgagagc cttcaaccca gtcagctcct
     2101 tccggtgggc gcggggcatg actatcgtcg ccgcacttat gactgtcttc tttatcatgc
     2161 aactcgtagg acaggtgccg gcagcgctct gggtcatttt cggcgaggac cgctttcgct
     2221 ggagcgcgac gatgatcggc ctgtcgcttg cggtattcgg aatcttgcac gccctcgctc
     2281 aagccttcgt cactggtccc gccaccaaac gtttcggcga gaagcaggcc attatcgccg
     2341 gcatggcggc cgacgcgctg ggctacgtct tgctggcgtt cgcgacgcga ggctggatgg
     2401 ccttccccat tatgattctt ctcgcttccg gcggcatcgg gatgcccgcg ttgcaggcca
     2461 tgctgtccag gcaggtagat gacgaccatc agggacagct tcaaggatcg ctcgcggctc
     2521 ttaccagcct aacttcgatc actggaccgc tgatcgtcac ggcgatttat gccgcctcgg
     2581 cgagcacatg gaacgggttg gcatggattg taggcgccgc cctatacctt gtctgcctcc
     2641 ccgcgttgcg tcgcggtgca tggagccggg ccacctcgac ctgaatggaa gccggcggca
     2701 cctcgctaac ggattcacca ctccaagaat tggagccaat caattcttgc ggagaactgt
     2761 gaatgcgcaa accaaccctt ggcagaacat atccatcgcg tccgccatct ccagcagccg
     2821 cacgcggcgc atctcgggca gcgttgggtc ctggccacgg gtgcgcatga tcgtgctcct
     2881 gtcgttgagg acccggctag gctggcgggg ttgccttact ggttagcaga atgaatcacc
     2941 gatacgcgag cgaacgtgaa gcgactgctg ctgcaaaacg tctgcgacct gagcaacaac
     3001 atgaatggtc ttcggtttcc gtgtttcgta aagtctggaa acgcggaagt cagcgccctg
     3061 caccattatg ttccggatct gcatcgcagg atgctgctgg ctaccctgtg gaacacctac
     3121 atctgtatta acgaagcgct ggcattgacc ctgagtgatt tttctctggt cccgccgcat
     3181 ccataccgcc agttgtttac cctcacaacg ttccagtaac cgggcatgtt catcatcagt
     3241 aacccgtatc gtgagcatcc tctctcgttt catcggtatc attaccccca tgaacagaaa
     3301 tcccccttac acggaggcat cagtgaccaa acaggaaaaa accgccctta acatggcccg
     3361 ctttatcaga agccagacat taacgcttct ggagaaactc aacgagctgg acgcggatga
     3421 acaggcagac atctgtgaat cgcttcacga ccacgctgat gagctttacc gcagctgcct
     3481 cgcgcgtttc ggtgatgacg gtgaaaacct ctgacacatg cagctcccgg agacggtcac
     3541 agcttgtctg taagcggatg ccgggagcag acaagcccgt cagggcgcgt cagcgggtgt
     3601 tggcgggtgt cggggcgcag ccatgaccca gtcacgtagc gatagcggag tgtatactgg
     3661 cttaactatg cggcatcaga gcagattgta ctgagagtgc accatatatg cggtgtgaaa
     3721 taccgcacag atgcgtaagg agaaaatacc gcatcaggcg ctcttccgct tcctcgctca
     3781 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     3841 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     3901 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3961 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4021 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     4081 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     4141 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     4201 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     4261 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     4321 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     4381 gaaggacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     4441 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     4501 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     4561 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     4621 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     4681 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     4741 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     4801 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     4861 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     4921 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     4981 cgccagttaa tagtttgcgc aacgttgttg ccattgctgc aggcatcgtg gtgtcacgct
     5041 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     5101 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     5161 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     5221 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     5281 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaac acgggataat accgcgccac
     5341 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     5401 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     5461 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     5521 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     5581 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     5641 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct
     5701 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     5761 gtcttcaaga a
//
