LOCUS       sequence-335991-        7979 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express ColicinB-V5-TEV-T7-mature MBP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7979)
  TITLE     Postle Lab Plasmids
REFERENCE   2  (bases 1 to 7979)
  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"
COMMENT     Sequence Label: pKP2085
FEATURES             Location/Qualifiers
     source          1..7979
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(1..6,7..23,24..29)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             2118..2159
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2160..2180
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2214..3314
                     /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
                     WYAVRTAVINAASGRQTVDAALAAAQT"
     CDS             3321..3338
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3723..3734
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4163..4182)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             complement(6035..6694)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     6609..6628
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(6695..6797)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(7189..7206)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7323..7868)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(7403..7422)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttgacagctt atcatcgata agctttaatg cggtagttta tcacagttaa attgctaacg
       61 cagtcaggca ccgtgtatga aatctaacaa tgcgctcatc gtcatcctcg gcaccgtcac
      121 cctggatgct gtaggcatag gcttggttat gccggtactg ccgggcctct tgcgggatat
      181 cgtccattcc gacagcatcg ccagtcacta tggcgtgctg ctagcgctat atgcgttgat
      241 gcaatttcta tgcgcacccg ttctcggagc actgtccgac cgctttggcc gccgcccagt
      301 cctgctcgct tcgctacttg gagccactat cgactacgcg atcatggcga ccacacccgt
      361 cctgtggatc cagaacagtt atgcctgtgc cgttttttgt ggtatcaggc gagaatagca
      421 cgtggtgtga atgcctgttt tttgatcgtt ttcacaaaaa ttgaaatcca cagtcttgac
      481 agagaaaaca cagcggcgta atttttgatc tgtacataaa accagtggtt atatgtacag
      541 tatttaattt taattgattg tttttaaagt caaagaggtt ttctatgagt gataatgaag
      601 gtagtgtacc gacagaaggt attgattacg gggacacaat ggttgtgtgg ccgtcaacag
      661 gacgaattcc gggcggtgat gtgaaacccg gaggctcatc aggtctcgct ccatccatgc
      721 ctccgggatg gggggattac agcccacaag gtatcgcact tgtacaaagt gttctttttc
      781 ctggaattat tcgccggatt attcttgata aagaacttga agagggagac tggtcgggat
      841 ggtctgtcag tgtgcatagc ccctggggaa acgagaaagt ttccgctgca cgaacagttc
      901 ttgaaaatgg tttacgtggt ggtttgccag aaccgtctcg cccggctgct gtttcttttg
      961 cccgtctgga gcctgcttcc ggaaatgagc aaaaaattat tcgtcttatg gttacacagc
     1021 aactggagca ggtaacggat atccctgcca gccagttacc agcagcgggt aataatgtac
     1081 cggtaaaata tcgtctgacg gaccttatgc agaatggtac acaatatatg gctattatcg
     1141 gaggtattcc gatgacagtg ccagtagtgg atgccgttcc agttccggac cggagtcgtc
     1201 cgggaaccaa tattaaagat gtttacagtg cccctgtatc accaaatcta ccggacctgg
     1261 tattaagtgt gggtcagatg aatactccag ttcggtctaa tcccgaaatc caggaagatg
     1321 gcgttatttc tgagacaggg aattatgttg aggctggtta tacgatgtcc agtaataatc
     1381 atgatgtcat tgtccgtttt cctgaaggca gtggagtttc tccgctatat atttcagccg
     1441 tggagattct ggacagtaat agtttaagcc agcgccagga agccgaaaat aacgcaaagg
     1501 atgacttcag agtcaagaaa gaacaagaaa atgacgagaa gacggtcctg acaaaaacca
     1561 gcgaggtcat cattagtgtc ggtgacaaag tcggggaata tcttggagat aaatacaagg
     1621 cgctttcccg tgaaattgca gagaatataa ataattttca gggaaaaacg attcgtagtt
     1681 atgatgatgc aatgtcttcc attaataagt taatggctaa ccccagcctt aaaataaatg
     1741 caacggacaa agaagccatt gtgaatgcgt ggaaagcatt taatgctgag gatatgggga
     1801 ataaatttgc tgcgttgggt aaaacgttca aagcagcaga ttatgcaata aaggcaaaca
     1861 acatcaggga gaagagtatt gagggttacc agactggtaa ctgggggcca ttaatgctgg
     1921 aagtcgagtc ctgggttatc agtgggatgg catctgctgt agctcttagt ttgttttctt
     1981 tgacattagg ctcggccctt atagcctttg gtctttcggc cacagttgtt ggttttgttg
     2041 gcgtagttat tgcaggtgct attggtgcat ttatcgatga taaatttgtt gatgagttga
     2101 atcacaagat cataaaaggc aaaccgattc cgaacccgct gctgggcctg gatagcaccg
     2161 aaaacctgta cttccagggt atggcgagca tgaccggcgg ccagcagatg ggcatgaaaa
     2221 tcgaagaagg taaactggta atctggatta acggcgataa aggctataac ggtctcgctg
     2281 aagtcggtaa gaaattcgag aaagataccg gaattaaagt caccgttgag catccggata
     2341 aactggaaga gaaattccca caggttgcgg caactggcga tggccctgac attatcttct
     2401 gggcacacga ccgctttggt ggctacgctc aatctggcct gttggctgaa atcaccccgg
     2461 acaaagcgtt ccaggacaag ctgtatccgt ttacctggga tgccgtacgt tacaacggca
     2521 agctgattgc ttacccgatc gctgttgaag cgttatcgct gatttataac aaagatctgc
     2581 tgccgaaccc gccaaaaacc tgggaagaga tcccggcgct ggataaagaa ctgaaagcga
     2641 aaggtaagag cgcgctgatg ttcaacctgc aagaaccgta cttcacctgg ccgctgattg
     2701 ctgctgacgg gggttatgcg ttcaagtatg aaaacggcaa gtacgacatt aaagacgtgg
     2761 gcgtggataa cgctggcgcg aaagcgggtc tgaccttcct ggttgacctg attaaaaaca
     2821 aacacatgaa tgcagacacc gattactcca tcgcagaagc tgcctttaat aaaggcgaaa
     2881 cagcgatgac catcaacggc ccgtgggcat ggtccaacat cgacaccagc aaagtgaatt
     2941 atggtgtaac ggtactgccg accttcaagg gtcaaccatc caaaccgttc gttggcgtgc
     3001 tgagcgcagg tattaacgcc gccagtccga acaaagagct ggcaaaagag ttcctcgaaa
     3061 actatctgct gactgatgaa ggtctggaag cggttaataa agacaaaccg ctgggtgccg
     3121 tagcgctgaa gtcttacgag gaagagttgg cgaaagatcc acgtattgcc gccactatgg
     3181 aaaacgccca gaaaggtgaa atcatgccga acatcccgca gatgtccgct ttctggtatg
     3241 ccgtgcgtac tgcggtgatc aacgccgcca gcggtcgtca gactgtcgat gcagccctgg
     3301 cagccgcgca gactaatgcg caccaccacc accaccacta attaaaagga aggcagtgtt
     3361 actgcatccc tttaaaatat ttaatgacaa gtaatggcat cagtaaaatc gcatatgtca
     3421 tggttaaaac agtaaagtaa atggagcaat ataaaataaa cagagtgatg acattatgag
     3481 ataataattt aaaaaaacca cctgctgttg ttagttccag attatagaac agtgtgatat
     3541 aaatcagaat ggcataaaaa acgggccccc agaaacagga ttgcacaaca agttttcttg
     3601 ttactggctt tatgatttct cttgtcttat atgtgcaaag aaaaacgaga atacccataa
     3661 ctggcgctgt cttgttgtag tttgacataa gtgttgtcat gacgggatca tgcaatgatg
     3721 taatagaagg gagaaatgcc accttattat ataagtagta cagcacttga gaatatggat
     3781 catttattcg aagtattaat atagccatta atggaataat cccgattatg gaaaatgcat
     3841 ataatatttc gtttgctttc ttgttcttat ctttattgct ggtcatatat ataccttttt
     3901 gtattttata tatagattat atatcagagt tctcaatata tttccagtat ttaatggagt
     3961 gtacagatat atatcttata tatggtagtt caggagctgc cctaccattt ttaacggcat
     4021 atcaggcaaa caatgttatg gggagcctct ctcatggggt ttgttcgccg gcggaggcgt
     4081 tagccagcgg cgaactctgg gatcctctac gccggacgca tcgtggccgg catcaccggc
     4141 gccacaggtg cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct
     4201 cgccacttcg ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc
     4261 gggggactgt tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac
     4321 ggcctcaacc tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga
     4381 ccgatgccct tgagagcctt caacccagtc agctccttcc ggtgggcgcg gggcatgact
     4441 atcgtcgccg cacttatgac tgtcttcttt atcatgcaac tcgtaggaca ggtgccggca
     4501 gcgctctggg tcattttcgg cgaggaccgc tttcgctgga gcgcgacgat gatcggcctg
     4561 tcgcttgcgg tattcggaat cttgcacgcc ctcgctcaag ccttcgtcac tggtcccgcc
     4621 accaaacgtt tcggcgagaa gcaggccatt atcgccggca tggcggccga cgcgctgggc
     4681 tacgtcttgc tggcgttcgc gacgcgaggc tggatggcct tccccattat gattcttctc
     4741 gcttccggcg gcatcgggat gcccgcgttg caggccatgc tgtccaggca ggtagatgac
     4801 gaccatcagg gacagcttca aggatcgctc gcggctctta ccagcctaac ttcgatcatt
     4861 ggaccgctga tcgtcacggc gatttatgcc gcctcggcga gcacatggaa cgggttggca
     4921 tggattgtag gcgccgccct ataccttgtc tgcctccccg cgttgcgtcg cggtgcatgg
     4981 agccgggcca cctcgacctg aatggaagcc ggcggcacct cgctaacgga ttcaccactc
     5041 caagaattgg agccaatcaa ttcttgcgga gaactgtgaa tgcgcaaacc aacccttggc
     5101 agaacatatc catcgcgtcc gccatctcca gcagccgcac gcggcgcatc tcgggcagcg
     5161 ttgggtcctg gccacgggtg cgcatgatcg tgctcctgtc gttgaggacc cggctaggct
     5221 ggcggggttg ccttactggt tagcagaatg aatcaccgat acgcgagcga acgtgaagcg
     5281 actgctgctg caaaacgtct gcgacctgag caacaacatg aatggtcttc ggtttccgtg
     5341 tttcgtaaag tctggaaacg cggaagtccc ctacgtgctg ctgaagttgc ccgcaacaga
     5401 gagtggaacc aaccggtgat accacgatac tatgactgag agtcaacgcc atgagcggcc
     5461 tcatttctta ttctgagtta caacagtccg caccgctgcc ggtagctcct tccggtgggc
     5521 gcggggcatg actatcgtcg ccgcacttat gactgtcttc tttatcatgc aactcgtagg
     5581 acaggtgccg gcagcgccca acagtccccc ggccacgggg cctgccacca tacccacgcc
     5641 gaaacaagcg ccctgcacca ttatgttccg gatctgcatc gcaggatgct gctggctacc
     5701 ctgtggaaca cctacatctg tattaacgaa gcgctaaccg tttttatcag gctctgggag
     5761 gcagaataaa tgatcatatc gtcaattatt acctccacgg ggagagcctg agcaaactgg
     5821 cctcaggcat ttgagaagca cacggtcaca ctgcttccgg tagtcaataa accggtaaac
     5881 cagcaataga cataagcggc tatttaacga ccctgccctg aaccgacgac cgggtcgaat
     5941 ttgctttcga atttctgcca ttcatccgct tattatcact tattcaggcg tagcaaccag
     6001 gcgtttaagg gcaccaataa ctgccttaaa aaaattacgc cccgccctgc cactcatcgc
     6061 agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa acggcatgat
     6121 gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat ttgcccatgg
     6181 tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac
     6241 tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta gggaaatagg
     6301 ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac tgccggaaat
     6361 cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt
     6421 aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc atacggaatt
     6481 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
     6541 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
     6601 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
     6661 caacggtggt atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc
     6721 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac
     6781 ctcttacgtg ccgatcaacg tctcattttc gccaaaagtt ggcccagggc ttcccggtat
     6841 caacagggac accaggattt atttattctg cgaagtgatc ttccgtcaca ggtatttatt
     6901 cggcgcaaag tgcgtcgggt gatgctgcca acttactgat ttagtgtatg atggtgtttt
     6961 tgaggtgctc cagtggcttc tgtttctatc agctgtccct cctgttcagc tactgacggg
     7021 gtggtgcgta acggcaaaag caccgccgga catcagcgct agcggagtgt atactggctt
     7081 actatgttgg cactgatgag ggtgtcagtg aagtgcttca tgtggcagga gaaaaaaggc
     7141 tgcaccggtg cgtcagcaga atatgtgata caggatatat tccgcttcct cgctcactga
     7201 ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg gcttacgaac ggggcggaga
     7261 tttcctggaa gatgccagga agatacttaa cagggaagtg agagggccgc ggcaaagccg
     7321 tttttccata ggctccgccc ccctgacaag catcacgaaa tctgacgctc aaatcagtgg
     7381 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggcgg ctccctcgtg
     7441 cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc cgctgttatg gccgcgtttg
     7501 tctcattcca cgcctgacac tcagttccgg gtaggcagtt cgctccaagc tggactgtat
     7561 gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     7621 caacccggaa agacatgcaa aagcaccact ggcagcagcc actggtaatt gatttagagg
     7681 agttagtctt gaagtcatgc gccggttaag gctaaactga aaggacaagt tttggtgact
     7741 gcgctcctcc aagccagtta cctcggttca aagagttggt agctcagaga accttcgaaa
     7801 aaccgccctg caaggcggtt ttttcgtttt cagagcaaga gattacgcgc agaccaaaac
     7861 gatctcaaga agatcatctt attaatcaga taaaatattt ctagatttca gtgcaattta
     7921 tctcttcaaa tgtagcacct gaagtcagcc ccatacgata taagttgtaa ttctcatgt
//
