LOCUS       40924_5739        5975 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector pBAD-HisB-PhoCl-MBP, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5975)
  AUTHORS   Zhang W, Lohman AW, Zhuravlova Y, Lu X, Wiens MD, Hoi H, Yaganoglu 
            S, Mohr MA, Kitova EN, Klassen JS, Pantazis P, Thompson RJ, Campbell
            RE.
  TITLE     Optogenetic control with a photocleavable protein, PhoCl
  JOURNAL   Nat. Methods (2017) In press
  PUBMED    28288123
REFERENCE   2  (bases 1 to 5975)
  AUTHORS   Zhang W.
  TITLE     Direct Submission
  JOURNAL   Submitted (01-FEB-2017) Chemistry, University of Alberta, 11227 
            Saskatchewan Drive, Department of Chemistry, CCIS 4-140, Edmonton, 
            Alberta T6G2C5, Canada
REFERENCE   3  (bases 1 to 5975)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5975)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Methods (2017) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (01-FEB-2017) Chemistry, University of Alberta, 11227 Saskatchewan 
            Drive, Department of Chemistry, CCIS 4-140, Edmonton, Alberta 
            T6G2C5, Canada"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: GeneStudio v. 2.2.0.0
            Sequencing Technology :: Illumina
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5975
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     CDS             331..348
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     CDS             352..384
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             388..411
                     /codon_start=1
                     /label=Xpress(TM) tag
                     /note="Xpress(TM) epitope tag, including an enterokinase 
                     recognition and cleavage site"
                     /translation="DLYDDDDK"
     CDS             427..1152
                     /codon_start=1
                     /label=PhoCl
                     /note="photocleavable protein that separates into two
                     fragments upon exposure to violet light (Zhang et al., 
                     2017)"
                     /translation="VIPDYFKQSFPEGYSWERSMTYEDGGICIATNDITMEGDSFINKI
                     HFKGTNFPPNGPVMQKRTVGWEASTEKMYERDGVLKGDVKMKLLLKGGGHYRCDYRTTY
                     KVKQKPVKLPDYHFVDHRIEILSHDKDYNKVKLYEHAVARNSTDSMDELYKGGSGGMVS
                     KGEETITSVIKPDMKNKLRMEGNVNGHAFVIEGEGSGKPFEGIQTIDLEVKEGAPLPFA
                     YDILTTAFHYGNRVFTKYPR"
     CDS             1168..2268
                     /codon_start=1
                     /label=MBP
                     /note="maltose binding protein from E. coli"
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     CDS             2317..2328
                     /codon_start=1
                     /label=Factor Xa site
                     /note="Factor Xa recognition and cleavage site"
                     /translation="IEGR"
     terminator      2546..2632
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      2724..2751
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     promoter        2770..2861
                     /label=AmpR promoter
     CDS             2862..3719
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      3893..4481
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(4667..4807)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(5074..5949)
                     /codon_start=1
                     /label=araC
                     /note="L-arabinose regulatory protein"
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
ORIGIN
        1 aagaaaccaa ttgtccatat tgcatcagac attgccgtca ctgcgtcttt tactggctct
       61 tctcgctaac caaaccggta accccgctta ttaaaagcat tctgtaacaa agcgggacca
      121 aagccatgac aaaaacgcgt aacaaaagtg tctataatca cggcagaaaa gtccacattg
      181 attatttgca cggcgtcaca ctttgctatg ccatagcatt tttatccata agattagcgg
      241 atcctacctg acgcttttta tcgcaactct ctactgtttc tccatacccg ttttttgggc
      301 taacaggagg aattaaccat ggggggttct catcatcatc atcatcatgg tatggctagc
      361 atgactggtg gacagcaaat gggtcgggat ctgtacgacg atgacgataa ggatccgagc
      421 tcgagtgtga tccctgacta cttcaagcag agcttccccg agggctacag ctgggagcgc
      481 agcatgacct acgaggacgg cggcatctgc atcgccacca acgacatcac aatggagggg
      541 gacagcttca tcaacaagat ccacttcaag ggcacgaact tcccccccaa cggccccgtg
      601 atgcagaaga ggaccgtggg ctgggaggcc agcaccgaga agatgtacga gcgcgacggc
      661 gtgctgaagg gcgacgtgaa gatgaagctg ctgctgaagg gcggcggcca ctatcgctgc
      721 gactaccgca ccacctacaa ggtcaagcag aagcccgtaa agctgcccga ctaccacttc
      781 gtggaccacc gcatcgagat cctgagccac gacaaggact acaacaaggt gaagctgtac
      841 gagcacgccg tggcccgcaa ctccaccgac agcatggacg agctgtacaa gggtggcagc
      901 ggtggcatgg tgagcaaggg cgaggagacc attacaagcg tgatcaagcc tgacatgaag
      961 aacaagctgc gcatggaggg caacgtgaac ggccacgcct tcgtgatcga gggcgagggc
     1021 agcggcaagc ccttcgaggg catccagacg attgatttgg aggtgaagga gggcgccccg
     1081 ctgcccttcg cctacgacat cctgaccacc gccttccact acggcaaccg cgtgttcacc
     1141 aagtacccac ggggaggtgg aggtaccatg aaaatcgaag aaggtaaact ggtaatctgg
     1201 attaacggcg ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat
     1261 accggaatta aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt
     1321 gcggcaactg gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac
     1381 gctcaatctg gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat
     1441 ccgtttacct gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt
     1501 gaagcgttat cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa
     1561 gagatcccgg cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac
     1621 ctgcaagaac cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag
     1681 tatgaaaacg gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcgaaagcg
     1741 ggtctgacct tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac
     1801 tccatcgcag aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg
     1861 gcatggtcca acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc
     1921 aagggtcaac catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt
     1981 ccgaacaaag agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg
     2041 gaagcggtta ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag
     2101 ttggcgaaag atccacgtat tgccgccacc atggaaaacg cccagaaagg tgaaatcatg
     2161 ccgaacatcc cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc
     2221 gccagcggtc gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg
     2281 aacaacaaca acaataacaa taacaacaac ctcgggatcg agggaaggat ttcataaaag
     2341 cttggctgtt ttggcggatg agagaagatt ttcagcctga tacagattaa atcagaacgc
     2401 agaagcggtc tgataaaaca gaatttgcct ggcggcagta gcgcggtggt cccacctgac
     2461 cccatgccga actcagaagt gaaacgccgt agcgccgatg gtagtgtggg gtctccccat
     2521 gcgagagtag ggaactgcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc
     2581 ctttcgtttt atctgttgtt tgtcggtgaa cgctctcctg agtaggacaa atccgccggg
     2641 agcggatttg aacgttgcga agcaacggcc cggagggtgg cgggcaggac gcccgccata
     2701 aactgccagg catcaaatta agcagaaggc catcctgacg gatggccttt ttgcgtttct
     2761 acaaactctt ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat
     2821 aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc
     2881 gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa
     2941 cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac
     3001 tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga
     3061 tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtgttgac gccgggcaag
     3121 agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca
     3181 cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca
     3241 tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa
     3301 ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc
     3361 tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa
     3421 cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     3481 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     3541 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     3601 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     3661 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     3721 aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat
     3781 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     3841 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     3901 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     3961 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     4021 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     4081 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg
     4141 gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc
     4201 ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg
     4261 aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg
     4321 cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag
     4381 ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc
     4441 gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct
     4501 ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc
     4561 ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc
     4621 gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgcctg atgcggtatt
     4681 ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc agtacaatct
     4741 gctctgatgc cgcatagtta agccagtata cactccgcta tcgctacgtg actgggtcat
     4801 ggctgcgccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     4861 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     4921 accgtcatca ccgaaacgcg cgaggcagca gatcaattcg cgcgcgaagg cgaagcggca
     4981 tgcataatgt gcctgtcaaa tggacgaagc agggattctg caaaccctat gctactccgt
     5041 caagccgtca attgtctgat tcgttaccaa ttatgacaac ttgacggcta catcattcac
     5101 tttttcttca caaccggcac ggaactcgct cgggctggcc ccggtgcatt ttttaaatac
     5161 ccgcgagaaa tagagttgat cgtcaaaacc aacattgcga ccgacggtgg cgataggcat
     5221 ccgggtggtg ctcaaaagca gcttcgcctg gctgatacgt tggtcctcgc gccagcttaa
     5281 gacgctaatc cctaactgct ggcggaaaag atgtgacaga cgcgacggcg acaagcaaac
     5341 atgctgtgcg acgctggcga tatcaaaatt gctgtctgcc aggtgatcgc tgatgtactg
     5401 acaagcctcg cgtacccgat tatccatcgg tggatggagc gactcgttaa tcgcttccat
     5461 gcgccgcagt aacaattgct caagcagatt tatcgccagc agctccgaat agcgcccttc
     5521 cccttgcccg gcgttaatga tttgcccaaa caggtcgctg aaatgcggct ggtgcgcttc
     5581 atccgggcga aagaaccccg tattggcaaa tattgacggc cagttaagcc attcatgcca
     5641 gtaggcgcgc ggacgaaagt aaacccactg gtgataccat tcgcgagcct ccggatgacg
     5701 accgtagtga tgaatctctc ctggcgggaa cagcaaaata tcacccggtc ggcaaacaaa
     5761 ttctcgtccc tgatttttca ccaccccctg accgcgaatg gtgagattga gaatataacc
     5821 tttcattccc agcggtcggt cgataaaaaa atcgagataa ccgttggcct caatcggcgt
     5881 taaacccgcc accagatggg cattaaacga gtatcccggc agcaggggat cattttgcgc
     5941 ttcagccata cttttcatac tcccgccatt cagag
//
