LOCUS       sequence-441881-        5969 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for N-terminally His-tagged maltose binding 
            protein (10xHis-MBP) fused to a protein of interest.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5969)
  AUTHORS   Barthel S, Brenker L, Diehl C, Bohra N, Giaveri S, Paczia N, Erb TJ
  TITLE     In vitro transcription-based biosensing of glycolate for prototyping
            of a complex enzyme cascade.
  JOURNAL   Synth Biol (Oxf). 2024 Sep 20;9(1):ysae013. doi: 
            10.1093/synbio/ysae013. eCollection 2024.
  PUBMED    39399720
REFERENCE   2  (bases 1 to 5969)
  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: "Synth Biol 
            (Oxf)."; date: "2024-09-20"; volume: "9(1):ysae013. doi"; pages: " 
            10.1093/synbio/ysae013. eCollection 2024"
COMMENT     Sequence Label: pTE5400_pMBP_CDS dropout vector
FEATURES             Location/Qualifiers
     source          1..5969
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             132..143
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             150..863
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      875..946
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      962..989
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      1040..1097
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(1121..1138)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1292..1880)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1372..1391)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(2063..2157)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2178..2837)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2752..2771
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     2999..3017
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(3021..3064)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    3067..3088
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     primer_bind     4220..4239
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(4259..4336)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     4542..4561
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     4645..4664
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4645..4663
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4664..4688
                     /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)."
     RBS             4703..4725
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             4738..4767
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             4777..5874
                     /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="KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEE
                     KFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLI
                     AYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAA
                     DGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGET
                     AMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLE
                     NYLLTDEGLEAVNKDKPLGAVALKSYEEELVKDPRIAATMENAQKGEIMPNIPQMSAFW
                     YAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     5848..5871
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             5923..5934
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
ORIGIN
        1 tcatgcgtca ttttgaacat tttgtaaatc ttatttaata atgtgtgcgg caattcacat
       61 ttaatttatg aatgttttct taacatcgcg gcaactcaag aaacggcagg ttcggatctt
      121 agctactaga gaaagaggag aaatactaga tgcgtaaagg cgaagagctg ttcactggtg
      181 tcgtccctat tctggtggaa ctggatggtg atgtcaacgg tcataagttt tccgtgcgtg
      241 gcgagggtga aggtgacgca actaatggta aactgacgct gaagttcatc tgtactactg
      301 gtaaactgcc ggttccttgg ccgactctgg taacgacgct gacttatggt gttcagtgct
      361 ttgctcgtta tccggaccat atgaagcagc atgacttctt caagtccgcc atgccggaag
      421 gctatgtgca ggaacgcacg atttccttta aggatgacgg cacgtacaaa acgcgtgcgg
      481 aagtgaaatt tgaaggcgat accctggtaa accgcattga gctgaaaggc attgacttta
      541 aagaggacgg caatatcctg ggccataagc tggaatacaa ttttaacagc cacaatgttt
      601 acatcaccgc cgataaacaa aaaaatggca ttaaagcgaa ttttaaaatt cgccacaacg
      661 tggaggatgg cagcgtgcag ctggctgatc actaccagca aaacactcca atcggtgatg
      721 gtcctgttct gctgccagac aatcactatc tgagcacgca aagcgttctg tctaaagatc
      781 cgaacgagaa acgcgatcat atggttctgc tggagttcgt aaccgcagcg ggcatcacgc
      841 atggtatgga tgaactgtac aaatgaccag gcatcaaata aaacgaaagg ctcagtcgaa
      901 agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctctact agagtcacac
      961 tggctcacct tcgggtgggc ctttctgcgt ttatacgtct catgagacca ctcatcgcga
     1021 ctagtagcgg ccgctgcagt ccggcaaaaa agggcaaggt gtcaccaccc tgcccttttt
     1081 ctttaaaacc gaaaagatta cttcgcgtta tgcaggcttc ctcgctcact gactcgctgc
     1141 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
     1201 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     1261 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccacag gctccgcccc cctgacgagc
     1321 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     1381 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     1441 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     1501 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     1561 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     1621 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     1681 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     1741 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     1801 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     1861 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     1921 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
     1981 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
     2041 ggtctgacag ctcgaggctt ggattctcac caataaaaaa cgcccggcgg caaccgagcg
     2101 ttctgaacaa atccagatgg agttctgagg tcattactgg atctatcaac aggagtccaa
     2161 gcgagctcga tatcaaatta cgccccgccc tgccactcgt cgcagtactg ttgtaattca
     2221 ttaagcattc tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc
     2281 ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag
     2341 aagttgtcca tattggccac gtttaaatca aaactggtga aactcaccca gggattggct
     2401 gatacgaaaa acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa
     2461 cacgccacat cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc
     2521 cagagtgatg aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta
     2581 tcccatatca ccagctcacc gtctttcatt gccatacgaa attccggatg agcattcatc
     2641 aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc
     2701 tttaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac
     2761 tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca
     2821 gtgatttttt tctccatttt agcttcctta gctcctgaaa atctcgataa ctcaaaaaat
     2881 acgcccggta gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgcccgatc
     2941 aactcgagtg ccacctgacg tctaagaaac cattattatc atgacattaa cctataaaaa
     3001 taggcgtatc acgaggcaga atttcagata aaaaaaatcc ttagctttcg ctaaggatga
     3061 tttctggaat tcgcggccgc ttctagagcg atgagtctgg gttgaaggct ctcaagggca
     3121 tcggtcgaga tcccggtgcc taatgagtga gctaacttac attaattgcg ttgcgctcac
     3181 tgcccgcttt ccagtcggga aactgtcgtg ccagctgcat taatgaatcg gccaacgcgc
     3241 ggggagaggc ggtttgcgta ttgggcgcca gggtggtttt tcttttcacc agtgagactg
     3301 gcaacagctg attgcccttc accgcctggc cctgagagag ttgcagcaag cggtccacgc
     3361 tggtttgccc cagcaggcga aaatcctgtt tgatggtggt taacggcggg atataacatg
     3421 agctgtcttc ggtatcgtcg tatcccacta ccgagatatc cgcaccaacg cgcagcccgg
     3481 actcggtaat ggcgcgcatt gcgcccagcg ccatctgatc gttggcaacc agcatcgcag
     3541 tgggaacgat gccctcattc agcatttgca tggtttgttg aaaaccggac atggcactcc
     3601 agtcgccttc ccgttccgct atcggctgaa tttgattgcg agtgagatat ttatgccagc
     3661 cagccagacg cagacgcgcc gagacagaac ttaatgggcc cgctaacagc gcgatttgct
     3721 ggtgacccaa tgcgaccaga tgctccacgc ccagtcgcgt accgtcttca tgggagaaaa
     3781 taatactgtt gatgggtgtc tggtcagaga catcaagaaa taacgccgga acattagtgc
     3841 aggcagcttc cacagcaatg gcatcctggt catccagcgg atagttaatg atcagcccac
     3901 tgacgcgttg cgcgagaaga ttgtgcaccg ccgctttaca ggcttcgacg ccgcttcgtt
     3961 ctaccatcga caccaccacg ctggcaccca gttgatcggc gcgagattta atcgccgcga
     4021 caatttgcga cggcgcgtgc agggccagac tggaggtggc aacgccaatc agcaacgact
     4081 gtttgcccgc cagttgttgt gccacgcggt tgggaatgta attcagctcc gccatcgccg
     4141 cttccacttt ttcccgcgtt ttcgcagaaa cgtggctggc ctggttcacc acgcgggaaa
     4201 cggtctgata agagacaccg gcatactctg cgacatcgta taacgttact ggtttcacat
     4261 tcaccaccct gaattgactc tcttccgggc gctatcatgc cataccgcga aaggttttgc
     4321 gccattcgat ggtgtccggg atctcgacgc tctcccttat gcgactcctg cattaggaag
     4381 cagcccagta gtaggttgag gccgttgagc accgccgccg caaggaatgg tgcatgcaag
     4441 gagatggcgc ccaacagtcc cccggccacg gggcctgcca ccatacccac gccgaaacaa
     4501 gcgctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc ggcgatatag
     4561 gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc ggcgtagagg
     4621 atcgagatct cgatcccgcg aaattaatac gactcactat aggggaattg tgagcggata
     4681 acaattcccc tctagaaata attttgttta actttaagaa ggagatatac catgggccat
     4741 catcatcatc atcatcatca tcatcacagc agcggcaaaa tcgaagaagg taaactggta
     4801 atctggatta acggcgataa aggctataac ggcctcgctg aagtcggtaa gaaattcgag
     4861 aaagataccg gaattaaagt caccgttgag catccggata aactggaaga gaaattccca
     4921 caggttgcgg caactggcga tggccctgac attatcttct gggcacacga ccgctttggt
     4981 ggctacgctc aatctggcct gttggctgaa atcaccccgg acaaagcgtt ccaggacaag
     5041 ctgtatccgt ttacctggga tgccgtacgt tacaacggca agctgattgc ttacccgatc
     5101 gctgttgaag cgttatcgct gatttataac aaagatctgc tgccgaaccc gccaaaaacc
     5161 tgggaagaga tcccggcgct ggataaagaa ctgaaagcga aaggtaagag cgcgctgatg
     5221 ttcaacctgc aagaaccgta cttcacctgg ccgctgattg ctgctgacgg gggttatgcg
     5281 ttcaagtatg aaaacggcaa gtacgacatt aaagacgtgg gcgtggataa cgctggcgcg
     5341 aaagcgggtc tgaccttcct ggttgacctg attaaaaaca aacacatgaa tgcagacacc
     5401 gattactcca tcgcagaagc tgcctttaat aaaggcgaaa cagcgatgac catcaacggc
     5461 ccgtgggcat ggtccaacat cgacaccagc aaagtgaatt atggtgtaac ggtactgccg
     5521 accttcaagg gtcaaccatc caaaccgttc gttggcgtgc tgagcgcagg tattaacgcc
     5581 gccagtccga acaaagagct ggcaaaagag ttcctcgaaa actatctgct gactgatgaa
     5641 ggtctggaag cggttaataa agacaaaccg ctgggtgccg tagcgctgaa gtcttacgag
     5701 gaagagttgg tgaaagatcc gcgtattgcc gccactatgg aaaacgccca gaaaggtgaa
     5761 atcatgccga acatcccgca gatgtccgct ttctggtatg ccgtgcgtac tgcggtgatc
     5821 aacgccgcca gcggtcgtca gactgtcgat gaagccctga aagacgcgca gactaattcg
     5881 agctcgaaca acaacaacaa taacaataac aacaacctcg ggatcgaagg tcgtcatggg
     5941 tctcgtgaga cggaaagtga aacgtgatt
//
