LOCUS       Exported                9485 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector pEBM113, complete sequence.
ACCESSION   KC779545
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9485)
  AUTHORS   Mearls EB, Olson DG, Herring CD, Lynd LR.
  TITLE     Development of a regulatable plasmid-based gene expression system 
            for Clostridium thermocellum
  JOURNAL   Appl. Microbiol. Biotechnol. 99 (18), 7589-7599 (2015)
  PUBMED    25994254
REFERENCE   2  (bases 1 to 9485)
  AUTHORS   Argyros A, Mearls EB.
  TITLE     Direct Submission
  JOURNAL   Submitted (13-MAR-2013) Engineering, Dartmouth College, 14 
            Engineering Dr, Hanover, NH 03755, USA
REFERENCE   3  (bases 1 to 9485)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..9485
                     /organism="Expression vector pEBM113"
                     /mol_type="other DNA"
                     /country="USA"
                     /label=derived from Clostridium thermocellum strain BL21
                     /note="derived from Clostridium thermocellum strain BL21"
                     /db_xref="taxon:1381394"
     regulatory      56..309
                     /regulatory_class="terminator"
     terminator      57..304
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      493..1166
     rep_origin      complement(552..1140)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1311..2171)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCDTLLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKRSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     CDS             complement(2267..3070)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(3071..3286)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      3309..3827
     misc_feature    3319..3822
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     rep_origin      3976..5510
     CDS             complement(4051..5055)
                     /codon_start=1
                     /gene="repB"
                     /product="RepB"
                     /label=repB
                     /note="replication protein B"
                     /protein_id="AGT95726.1"
                     /translation="MGVSFNIMCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDY
                     FELLHILEFKKAERVKDCAEILEYKQNRETGERKLYRVWFCKSRLCPMCNWRRAMKHGI
                     QSQKVVAEVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLV
                     GFMRATEVTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPN
                     VKVQMIRPKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLIS
                     YGGLLKEIHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
     gene            complement(4051..5055)
                     /gene="repB"
                     /label=repB
     regulatory      5541..6188
                     /regulatory_class="promoter"
                     /gene="gapD"
     gene            5541..6188
                     /gene="gapD"
                     /label=gapD
     CDS             6189..6839
                     /codon_start=1
                     /gene="cat"
                     /product="Cat"
                     /label=cat
                     /note="chloramphenicol acetyltransferase"
                     /protein_id="AGT95727.1"
                     /translation="MNFNKIDLDNWKRKEIFNHYLNQQTTFSITTEIDISVLYRNIKQE
                     GYKFYPAFIFLVTRVINSNTAFRTGYNSDGELGYWDKLEPLYTIFDGVSKTFSGIWTPV
                     KNDFKEFYDLYLSDVEKYNGSGKLFPKTPIPENAFSLSIIPWTSFTGFNLNINNNSNYL
                     LPIITAGKFINKGNSIYLPLSLQVHHSVCDGYHAGLFMNSIQELSDRPNDWLL"
     gene            6189..6839
                     /gene="cat"
                     /label=cat
     regulatory      7417..7456
                     /regulatory_class="terminator"
     regulatory      7464..7502
                     /regulatory_class="promoter"
                     /gene="celC"
     gene            7464..7502
                     /gene="celC"
                     /label=celC
     CDS             7626..8663
                     /codon_start=1
                     /gene="glyR3"
                     /product="GlyR3"
                     /label=glyR3
                     /note="repressor"
                     /protein_id="AGT95728.1"
                     /translation="MTSEEIAKLCGVSRATVSRVINNSPNVKEETRQKILAVIKEKNYV
                     PIAPARRLAGIDSNIIGLFVLDIDISESKSRVSESTYFSRLINLIIDQANNFGFQVLVS
                     IITSQKQLSEIRNLFMSRTIFSGIFIGAFNDEIQLDDDIIMQHPTIIIDRQSERMVKKP
                     NRLVVNLDNFEGAYNATQFLIKLGHTRIGHISGDLRKLSGIERYEGYKKALEDAGLGFD
                     KNLVREGNFLDDSGYRLAREILKENVTAIFCANDVMAISAIKAIKETGLSVPDDISVIG
                     FDNTAIGNYIMPALTTVNAPLEHIAEACIESLKYFCEHKHFKQKEIRVKTDLIIRDSTK
                     RALEF"
     gene            7626..8663
                     /gene="glyR3"
                     /label=glyR3
     CDS             8676..9485
                     /codon_start=1
                     /gene="spo0A"
                     /product="Spo0A"
                     /label=spo0A
                     /note="sporulation response regulator A"
                     /protein_id="AGT95729.1"
                     /translation="MTSNKIRVVIADDNREFGDILYEYLNNQEDIEVVGVARDGIEAYE
                     LIVEKLPDIAILDIIMPHLDGLGVLEKIGATAISKRPLFIILSAVGQDKITQRALALGA
                     EYYVVKPFDMEVLISRIRQLKNVNQPNVIRQDGLSGEVKSSYHPPQPKNLEAEVTNIMH
                     EIGVPAHIKGYQYLRDAIIMVVKDLDVINSITKQLYPTIAKEYNTTPSRVERAIRHAIE
                     VAWSRGQIDTIDSLFGYTINVGKGKPTNSEFIAMVADKLRLEMKMAT"
     gene            8676..9485
                     /gene="spo0A"
                     /label=spo0A