LOCUS       sequence-447905-        6049 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  BccP protein purification.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6049)
  AUTHORS   Haidar-Ahmad N, Tomaro K, Lavallee-Adam M, Campbell-Valois F-X
  TITLE     The promiscuous biotin ligase TurboID reveals the proxisome of the 
            T3SS chaperone IpgC in Shigella flexneri.
  JOURNAL   mSphere. 2024 Nov 21;9(11):e0055324. doi: 10.1128/msphere.00553-24. 
            Epub 2024 Oct 31.
  PUBMED    39480076
REFERENCE   2  (bases 1 to 6049)
  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: "mSphere."; 
            date: "2024-11-21"; volume: "9(11):e0055324. doi"; pages: " 
            10.1128/msphere.00553-24. Epub 2024 Oct 31"
COMMENT     Sequence Label: pNHA8
FEATURES             Location/Qualifiers
     source          1..6049
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(10..24,25..30,31..47,48..54)
                     /label=T5 promoter
                     /note="bacteriophage T5 promoter for E. coli RNA 
                     polymerase, with embedded lac operator"
     protein_bind    30..46
                     /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)."
     protein_bind    62..78
                     /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             97..108
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             127..1227
                     /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
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     regulatory      1084..1093
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     1201..1224
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     terminator      1506..1600
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             1644..2303
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1710..1729)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      2368..2454
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(2547..3629)
                     /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     3591..3610
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(3630..3707)
                     /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."
     primer_bind     complement(3791..3813)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3899..4039
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     3954..3973
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4054..4071)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4225..4813)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4305..4324)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4984..5775,5776..5844))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5607..5626
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5845..5949)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6017..6035
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 ctcgagaaat cataaaaaat ttatttgctt tgtgagcgga taacaattat aatagattca
       61 attgtgagcg gataacaatt tcacacagaa ttcattaaag aggagaaatt aactatgaga
      121 ggatcgatga aaatcgaaga aggtaaactg gtaatctgga ttaacggcga taaaggctat
      181 aacggtctcg ctgaagtcgg taagaaattc gagaaagata ccggaattaa agtcaccgtt
      241 gagcatccgg ataaactgga agagaaattc ccacaggttg cggcaactgg cgatggccct
      301 gacattatct tctgggcaca cgaccgcttt ggtggctacg ctcaatctgg cctgttggct
      361 gaaatcaccc cggacaaagc gttccaggac aagctgtatc cgtttacctg ggatgccgta
      421 cgttacaacg gcaagctgat tgcttacccg atcgctgttg aagcgttatc gctgatttat
      481 aacaaagatc tgctgccgaa cccgccaaaa acctgggaag agatcccggc gctggataaa
      541 gaactgaaag cgaaaggtaa gagcgcgctg atgttcaacc tgcaagaacc gtacttcacc
      601 tggccgctga ttgctgctga cgggggttat gcgttcaagt atgaaaacgg caagtacgac
      661 attaaagacg tgggcgtgga taacgctggc gcgaaagcgg gtctgacctt cctggttgac
      721 ctgattaaaa acaaacacat gaatgcagac accgattact ccatcgcaga agctgccttt
      781 aataaaggcg aaacagcgat gaccatcaac ggcccgtggg catggtccaa catcgacacc
      841 agcaaagtga attatggtgt aacggtactg ccgaccttca agggtcaacc atccaaaccg
      901 ttcgttggcg tgctgagcgc aggtattaac gccgccagtc cgaacaaaga gctggcgaaa
      961 gagttcctcg aaaactatct gctgactgat gaaggtctgg aagcggttaa taaagacaaa
     1021 ccgctgggtg ccgtagcgct gaagtcttac gaggaagagt tggcgaaaga tccacgtatt
     1081 gccgccacca tggaaaacgc ccagaaaggt gaaatcatgc cgaacatccc gcagatgtcc
     1141 gctttctggt atgccgtgcg tactgcggtg atcaacgccg ccagcggtcg tcagactgtc
     1201 gatgaagccc tgaaagacgc gcagactgga tccgaaatca gtggtcacat cgtacgttcc
     1261 ccgatggttg gtactttcta ccgcacccca agcccggacg caaaagcgtt catcgaagtg
     1321 ggtcagaaag tcaacgtggg cgataccctg tgcatcgttg aagccatgaa aatgatgaac
     1381 cagatcgaag cggacaaatc cggtaccgtg aaagcaattc tggtcgaaag tggacaaccg
     1441 gtagaatttg acgagccgct ggtcgtcatc gagtaactgc agccaagctt aattagctga
     1501 gcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg atttgttcag
     1561 aacgctcggt tgccgccggg cgttttttat tggtgagaat ccaagctagc ttggcgagat
     1621 tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc accgttgata
     1681 tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct caatgtacct
     1741 ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag aaaaataagc
     1801 acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct catccggaat
     1861 ttcgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac ccttgttaca
     1921 ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac cacgacgatt
     1981 tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa aacctggcct
     2041 atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc tgggtgagtt
     2101 tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc gttttcacca
     2161 tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt caggttcatc
     2221 atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa cagtactgcg
     2281 atgagtggca gggcggggcg taattttttt aaggcagtta ttggtgccct taaacgcctg
     2341 gggtaatgac tctctagctt gaggcatcaa ataaaacgaa aggctcagtc gaaagactgg
     2401 gcctttcgtt ttatctgttg tttgtcggtg aacgctctcc tgagtaggac aaatccgccc
     2461 tctagattac gtgcagtcga tgataagctg tcaaacatga gaattgtgcc taatgagtga
     2521 gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     2581 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgcc
     2641 agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt caccgcctgg
     2701 ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg aaaatcctgt
     2761 ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc gtatcccact
     2821 accgagatat ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat tgcgcccagc
     2881 gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt cagcatttgc
     2941 atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc tatcggctga
     3001 atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc cgagacagaa
     3061 cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag atgctccacg
     3121 cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt ctggtcagag
     3181 acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat ggcatcctgg
     3241 tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag attgtgcacc
     3301 gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac gctggcaccc
     3361 agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg cagggccaga
     3421 ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg tgccacgcgg
     3481 ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt tttcgcagaa
     3541 acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc ggcatactct
     3601 gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact ctcttccggg
     3661 cgctatcatg ccataccgcg aaaggttttg caccattcga tggtgtcgga atttcgggca
     3721 gcgttgggtc ctggccacgg gtgcgcatga tctagagctg cctcgcgcgt ttcggtgatg
     3781 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
     3841 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggcg
     3901 cagccatgac ccagtcacgt agcgatagcg gagtgtatac tggcttaact atgcggcatc
     3961 agagcagatt gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag
     4021 gagaaaatac cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4081 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4141 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4201 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4261 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4321 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4381 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4441 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4501 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     4561 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     4621 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     4681 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     4741 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     4801 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     4861 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     4921 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     4981 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5041 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5101 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5161 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5221 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5281 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5341 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5401 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5461 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     5521 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     5581 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     5641 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     5701 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     5761 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     5821 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     5881 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     5941 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat
     6001 gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtcttcac
//
