LOCUS       sequence-352195-        7605 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses His6 tagged MBP-BirA fusion protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7605)
  TITLE     Bacterial Expression Plasmid for H6-MBP-BirA
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7605)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pH6-MBP-TEV-BirA
FEATURES             Location/Qualifiers
     source          1..7605
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..80
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             81..1163
                     /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(100..119)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        join(1406..1411,1412..1427,1428..1434)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    1442..1458
                     /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     1447..1469
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             1540..1557
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1567..1584
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             1588..2688
                     /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
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVVVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     2662..2685
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2701..2721
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2722..3684
                     /codon_start=1
                     /gene="birA"
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     CDS             2722..3681
                     /codon_start=1
                     /product="E. coli biotin protein ligase"
                     /label=BirA
                     /translation="KDNTVPLKLIALLANGEFHSGEQLGETLGMSRAAINKHIQTLRDW
                     GVDVFTVPGKGYSLPEPIQLLNAKQILGQLDGGSVAVLPVIDSTNQYLLDRIGELKSGD
                     ACIAEYQQAGRGRRGRKWFSPFGANLYLSMFWRLEQGPAAAIGLSLVIGIVMAEVLRKL
                     GADKVRVKWPNDLYLQDRKLAGILVELTGKTGDAAQIVIGAGINMAMRRVEESVVNQGW
                     ITLQEAGINLDRNTLAAMLIRELRAALELFEQEGLAPYLSRWEKLDNFINRPVKLIIGD
                     KEIFGISRGIDKQGALLLEQDGIIKPWMGGEISLRSAEK"
     primer_bind     complement(3696..3713)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3696..3712)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3705..3727)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(3891..3908)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      4061..4147
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      4239..4266
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        4285..4376
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4377..4445,4446..5237)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMEDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4595..4614)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      complement(5279..5792)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5474..5495)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5686..5705
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     rep_origin      5903..6491
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6392..6411
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6645..6662
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6677..6819
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6745..6764)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6905..6927
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(6921..7112)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ccgacaccat cgaatggtgc aaaacctttc gcggtatggc atgatagcgc ccggaagaga
       61 gtcaattcag ggtggtgaat gtgaaaccag taacgttata cgatgtcgca gagtatgccg
      121 gtgtctctta tcagaccgtt tcccgcgtgg tgaaccaggc cagccacgtt tctgcgaaaa
      181 cgcgggaaaa agtggaagcg gcgatggcgg agctgaatta cattcccaac cgcgtggcac
      241 aacaactggc gggcaaacag tcgttgctga ttggcgttgc cacctccagt ctggccctgc
      301 acgcgccgtc gcaaattgtc gcggcgatta aatctcgcgc cgatcaactg ggtgccagcg
      361 tggtggtgtc gatggtagaa cgaagcggcg tcgaagcctg taaagcggcg gtgcacaatc
      421 ttctcgcgca acgcgtcagt gggctgatca ttaactatcc gctggatgac caggatgcca
      481 ttgctgtgga agctgcctgc actaatgttc cggcgttatt tcttgatgtc tctgaccaga
      541 cacccatcaa cagtattatt ttctcccatg aagacggtac gcgactgggc gtggagcatc
      601 tggtcgcatt gggtcaccag caaatcgcgc tgttagcggg cccattaagt tctgtctcgg
      661 cgcgtctgcg tctggctggc tggcataaat atctcactcg caatcaaatt cagccgatag
      721 cggaacggga aggcgactgg agtgccatgt ccggttttca acaaaccatg caaatgctga
      781 atgagggcat cgttcccact gcgatgctgg ttgccaacga tcagatggcg ctgggcgcaa
      841 tgcgcgccat taccgagtcc gggctgcgcg ttggtgcgga tatctcagta gtgggatacg
      901 acgataccga agacagctca tgttatatcc cgccgttaac caccatcaaa caggattttc
      961 gcctgctggg gcaaaccagc gtggaccgct tgctgcaact ctctcagggc caggcggtga
     1021 agggcaatca gctgttgccc gtctcactgg tgaaaagaaa aaccaccctg gcgcccaata
     1081 cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt
     1141 cccgactgga aagcgggcag tgagcgcaac gcaattaatg taagttagct cactcattag
     1201 gcacaattct catgtttgac agcttatcat cgactgcacg gtgcaccaat gcttctggcg
     1261 tcaggcagcc atcggaagct gtggtatggc tgtgcaggtc gtaaatcact gcataattcg
     1321 tgtcgctcaa ggcgcactcc cgttctggat aatgtttttt gcgccgacat cataacggtt
     1381 ctggcaaata ttctgaaatg agctgttgac aattaatcat cggctcgtat aatgtgtgga
     1441 attgtgagcg gataacaatt tcacacagga aacagccagt ccgtttaggt gttttcacga
     1501 gcacttcacc aacaaggacc atagcatatg ggcagcagcc atcatcatca tcatcacagc
     1561 agcggcctgg tgccgcgcgg cagccatatg aaaatcgaag aaggtaaact ggtaatctgg
     1621 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     1681 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     1741 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     1801 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     1861 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     1921 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
     1981 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
     2041 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     2101 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     2161 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     2221 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     2281 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     2341 aagggtcaac catccaaacc gttcgttgtc gtgctgagcg caggtattaa cgccgccagt
     2401 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     2461 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     2521 ttggcgaaag atccacgtat tgccgccact atggaaaacg cccagaaagg tgaaatcatg
     2581 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     2641 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctca
     2701 gagaacctgt atttccaggg aaaggataac accgtgccac tgaaattgat tgccctgtta
     2761 gcgaacggtg aatttcactc tggcgagcag ttgggtgaaa cgctgggaat gagccgggcg
     2821 gctattaata aacacattca gacactgcgt gactggggcg ttgatgtctt taccgttccg
     2881 ggtaaaggat acagcctgcc tgagcctatc cagttactta atgctaaaca gatattgggt
     2941 cagctggatg gcggtagtgt agccgtgctg ccagtgattg actccacgaa tcagtacctt
     3001 cttgatcgta tcggagagct taaatcgggc gatgcttgca ttgcagaata ccagcaggct
     3061 ggccgtggtc gccggggtcg gaaatggttt tcgccttttg gcgcaaactt atatttgtcg
     3121 atgttctggc gtctggaaca aggcccggcg gcggcgattg gtttaagtct ggttatcggt
     3181 atcgtgatgg cggaagtatt acgcaagctg ggtgcagata aagttcgtgt taaatggcct
     3241 aatgacctct atctgcagga tcgcaagctg gcaggcattc tggtggagct gactggcaaa
     3301 actggcgatg cggcgcaaat agtcattgga gccgggatca acatggcaat gcgccgtgtt
     3361 gaagagagtg tcgttaatca ggggtggatc acgctgcagg aagcggggat caatctcgat
     3421 cgtaatacgt tggcggccat gctaatacgt gaattacgtg ctgcgttgga actcttcgaa
     3481 caagaaggat tggcacctta tctgtcgcgc tgggaaaagc tggataattt tattaatcgc
     3541 ccagtgaaac ttatcattgg tgataaagaa atatttggca tttcacgcgg aatagacaaa
     3601 cagggggctt tattacttga gcaggatgga ataataaaac cctggatggg cggtgaaata
     3661 tccctgcgta gtgcagaaaa ataatgaagc ttggcactgg ccgtcgtttt acaacgtcgt
     3721 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
     3781 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
     3841 aatggcgaat ggcagcttgg ctgttttggc ggatgagata agattttcag cctgatacag
     3901 attaaatcag aacgcagaag cggtctgata aaacagaatt tgcctggcgg cagtagcgcg
     3961 gtggtcccac ctgaccccat gccgaactca gaagtgaaac gccgtagcgc cgatggtagt
     4021 gtggggtctc cccatgcgag agtagggaac tgccaggcat caaataaaac gaaaggctca
     4081 gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc tcctgagtag
     4141 gacaaatccg ccgggagcgg atttgaacgt tgcgaagcaa cggcccggag ggtggcgggc
     4201 aggacgcccg ccataaactg ccaggcatca aattaagcag aaggccatcc tgacggatgg
     4261 cctttttgcg tttctacaaa ctcttttgtt tatttttcta aatacattca aatatgtatc
     4321 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     4381 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     4441 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     4501 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     4561 aacgtttccc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgtg
     4621 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     4681 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     4741 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     4801 gaccgaagga gctaaccgct tttttgcaca acatggagga tcatgtaact cgccttgatc
     4861 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     4921 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     4981 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     5041 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     5101 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     5161 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     5221 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttac
     5281 cccggttgat aatcagaaaa gccccaaaaa caggaagatt gtataagcaa atatttaaat
     5341 tgtaaacgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt
     5401 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     5461 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     5521 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat cacccaaatc
     5581 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     5641 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     5701 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     5761 cgccgcgctt aatgcgccgc tacagggcgc gtaaaaggat ctaggtgaag atcctttttg
     5821 ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg
     5881 tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc
     5941 aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc
     6001 tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtc cttctagtgt
     6061 agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
     6121 taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact
     6181 caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac
     6241 agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag
     6301 aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg
     6361 gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg
     6421 tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga
     6481 gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt
     6541 ttgctcacat gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct
     6601 ttgagtgagc tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg
     6661 aggaagcgga agagcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     6721 accgcatata tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagta
     6781 tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc gccaacaccc
     6841 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     6901 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgaggcag
     6961 ctgcggtaaa gctcatcagc gtggtcgtgc agcgattcac agatgtctgc ctgttcatcc
     7021 gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat aaagcgggcc
     7081 atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg gggatttctg
     7141 ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg ggttactgat
     7201 gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt atggatgcgg
     7261 cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac agatgtaggt
     7321 gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat ggtgcagggc
     7381 gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat tcatgttgtt
     7441 gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg tatcggtgat
     7501 tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa cgacaggagc
     7561 acgatcatgc gcacccgtgg ccaggaccca acgctgcccg aaatt
//
