LOCUS       Exported               10329 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pT3C, complete sequence.
ACCESSION   EF127523
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10329)
  AUTHORS   Li XL, Skory CD, Ximenes EA, Jordan DB, Dien BS, Hughes SR, Cotta 
            MA.
  TITLE     Expression of an AT-rich xylanase gene from the anaerobic fungus 
            Orpinomyces sp. strain PC-2 in and secretion of the heterologous 
            enzyme by Hypocrea jecorina
  JOURNAL   Appl. Microbiol. Biotechnol. 74 (6), 1264-1275 (2007)
  PUBMED    17225100
REFERENCE   2  (bases 1 to 10329)
  AUTHORS   Li X-L.
  TITLE     Direct Submission
  JOURNAL   Submitted (20-NOV-2006) Fermentation Biotechnology Research Unit, 
            National Center for Agricultural Utilization Research, USDA-ARS, 
            1815 N. University Street, Peoria, IL 61604, USA
REFERENCE   3  (bases 1 to 10329)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10329
                     /organism="Expression vector pT3C"
                     /lab_host="Hypocrea jecorina"
                     /mol_type="other DNA"
                     /label=constructed from pBluescript KS(-)
                     /note="constructed from pBluescript KS(-)"
                     /db_xref="taxon:428319"
     promoter        21..125
                     /gene="bla"
                     /label=AmpR promoter
     CDS             126..986
                     /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"
     rep_origin      1157..1745
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    2033..2054
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2069..2099
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2107..2123
                     /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     2131..2147
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2168..2186
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     regulatory      2206..4420
                     /regulatory_class="promoter"
                     /note="from Hypocrea jecorina cel7A"
     RBS             3933..3941
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3 end of the 16S rRNA (Chen et al., 1994)"
     CDS             4421..4684
                     /codon_start=1
                     /gene="egII/cel5A"
                     /product="EGII"
                     /label=egII/cel5A
                     /note="includes signal peptide, carbohydrate binding 
                     module, and hinge sequences; fuses to Orpinomyces xynA for 
                     expression"
                     /protein_id="ABO21312.1"
                     /translation="MNKSVAPLLLAASILYGGAVAQQTVWGQCGGIGWSGPTNCAPGSA
                     CSTLNPYYAQCIPGATTITTSTRPPSGPTTTTRATSTSSSTPP"
     gene            4421..4684
                     /gene="egII/cel5A"
                     /label=egII/cel5A
     CDS             complement(4559..4570)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      4702..5410
                     /regulatory_class="terminator"
                     /note="from Hypocrea jecorina cel7A"
     CDS             5785..6810
                     /codon_start=1
                     /gene="hygR"
                     /product="hygromycinB phosphotransferase"
                     /label=hygR
                     /note="APH; hygromycin resistance protein"
                     /protein_id="ABO21313.1"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGQFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     gene            5785..6810
                     /gene="hygR"
                     /label=hygR
     misc_feature    6826..9648
                     /label=Hypocrea jecorina cel7A 3 region
                     /note="Hypocrea jecorina cel7A 3 region"
     promoter        complement(9686..9704)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9711..9727)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      9869..10324
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"