LOCUS       Exported                8593 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pNGWA, complete sequence.
ACCESSION   EU680845
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8593)
  AUTHORS   Busso D, Delagoutte-Busso B, Moras D.
  TITLE     Construction of a set Gateway-based destination vectors for 
            high-throughput cloning and expression screening in Escherichia coli
  JOURNAL   Anal. Biochem. 343 (2), 313-321 (2005)
  PUBMED    15993367
REFERENCE   2  (bases 1 to 8593)
  AUTHORS   Busso D, Delagoutte-Busso B, Salim L, Moras D.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-APR-2008) Structural Biology and Genomics Platform, 
            IGBMC, 1, rue Laurent Fries, Illkirch 67404, France
REFERENCE   3  (bases 1 to 8593)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8593
                     /organism="Expression vector pNGWA"
                     /mol_type="other DNA"
                     /note="Gateway-adapted expression vector for E. coli: N-ter
                     NusA tag - C-ter His tag"
                     /db_xref="taxon:526725"
     promoter        27..45
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      27..44
                     /regulatory_class="promoter"
                     /note="T7"
     misc_feature    46..70
                     /label=lac operator
                     /note="lac operator"
     protein_bind    46..70
                     /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             85..107
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     regulatory      100..106
                     /regulatory_class="ribosome_binding_site"
     misc_feature    123..1604
                     /label=NusA solubility enhancer
                     /note="NusA solubility enhancer"
     CDS             123..1604
                     /codon_start=1
                     /gene="nusA"
                     /product="transcription elongation protein (N utilization 
                     substance protein A)"
                     /label=NusA
                     /note="highly soluble in E. coli"
                     /translation="NKEILAVVEAVSNEKALPREKIFEALESALATATKKKYEQEIDVR
                     VQIDRKSGDFDTFRRWLVVDEVTQPTKEITLEAARYEDESLNLGDYVEDQIESVTFDRI
                     TTQTAKQVIVQKVREAERAMVVDQFREHEGEIITGVVKKVNRDNISLDLGNNAEAVILR
                     EDMLPRENFRPGDRVRGVLYSVRPEARGAQLFVTRSKPEMLIELFRIEVPEIGEEVIEI
                     KAAARDPGSRAKIAVKTNDKRIDPVGACVGMRGARVQAVSTELGGERIDIVLWDDNPAQ
                     FVINAMAPADVASIVVDEDKHTMDIAVEAGNLAQAIGRNGQNVRLASQLSGWELNVMTV
                     DDLQAKHQAEAHAAIDTFTKYLDIDEDFATVLVEEGFSTLEELAYVPMKELLEIEGLDE
                     PTVEALRERAKNALATIAQAQEESLGDNKPADDLLNLEGVDRDLAFKLAARGVCTLEDL
                     AEQGIDDLADIEGLTDEKAGALIMAARNICWFGDEA"
     misc_recomb     1623..1747
                     /label=AttR1
                     /note="AttR1"
     protein_bind    1623..1747
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1772..1802
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1856..2515
                     /codon_start=1
                     /product="chloramphenicol resistance protein"
                     /label=chloramphenicol resistance protein
                     /protein_id="ACD44871.1"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     CDS             2857..3162
                     /codon_start=1
                     /product="CcdB"
                     /label=CcdB
                     /protein_id="ACD44872.1"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     misc_recomb     3203..3327
                     /label=AttR2
                     /note="AttR2"
     protein_bind    complement(3203..3327)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    3341..3358
                     /label=6 His affinity tag
                     /note="6 His affinity tag"
     CDS             3341..3358
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      3425..3472
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     regulatory      3426..3472
                     /regulatory_class="terminator"
                     /note="T7"
     rep_origin      3509..3964
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        3991..4095
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4096..4956
                     /codon_start=1
                     /product="ampicillin resistance protein"
                     /label=ampicillin resistance protein
                     /protein_id="ACD44873.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5101..5774
                     /label=pBR322
                     /note="pBR322"
     rep_origin      5127..5715
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    5901..6043
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(6145..6336)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(7145..8227)
                     /codon_start=1
                     /product="LacI"
                     /label=LacI
                     /protein_id="ACD44874.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(8228..8305)
                     /gene="lacI"
                     /label=lacI promoter