LOCUS       Exported                6690 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pD-GEX1, complete sequence.
ACCESSION   EF431916
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6690)
  AUTHORS   Ergin A, Bussow K, Sieper J, Thiel A, Duchmann R, Adam T.
  TITLE     Homologous High-Throughput Expression and Purification of Highly 
            Conserved E. coli Proteins
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6690)
  AUTHORS   Ergin A, Adam T, Bussow K.
  TITLE     Direct Submission
  JOURNAL   Submitted (09-FEB-2007) Gastroenterology, Charite Berlin (CBF), 
            Hindenburgdamm 30, Berlin 12200, Germany
REFERENCE   3  (bases 1 to 6690)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6690
                     /organism="Cloning vector pD-GEX1"
                     /mol_type="other DNA"
                     /db_xref="taxon:429337"
     promoter        183..211
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    219..235
                     /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)."
     CDS             258..1004
                     /codon_start=1
                     /gene="gst"
                     /product="glutathione S-transferase"
                     /label=gst
                     /note="contains His-tag"
                     /protein_id="ABO28844.1"
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPKSDLI
                     EGRGIHHHHHHHSHQTSLYKKAERET"
     gene            258..1004
                     /gene="gst"
                     /label=gst
     CDS             258..911
                     /codon_start=1
                     /product="glutathione S-transferase from Schistosoma 
                     japonicum"
                     /label=GST
                     /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK
                     FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY
                     GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL
                     YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPK"
     CDS             921..932
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     misc_feature    939..959
                     /label=seven-histidine tag
                     /note="seven-histidine tag"
     CDS             939..959
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     misc_feature    964..1088
                     /label=attR1 recombination site
                     /note="attR1 recombination site"
     protein_bind    969..1093
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        1118..1148
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1202..1882
                     /codon_start=1
                     /gene="cmR"
                     /product="chloramphenicol resistance protein"
                     /label=cmR
                     /protein_id="ABO28845.1"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     gene            1202..1882
                     /gene="cmR"
                     /label=cmR
     CDS             2202..2507
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB"
                     /label=ccdB
                     /protein_id="ABO28846.1"
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     gene            2202..2507
                     /gene="ccdB"
                     /label=ccdB
     protein_bind    complement(2548..2672)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     misc_feature    2553..2677
                     /label=attR2 recombination site
                     /note="attR2 recombination site"
     promoter        2993..3097
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3098..3958
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /protein_id="ABO28847.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            3098..3958
                     /gene="bla"
                     /label=bla
     rep_origin      4129..4717
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        4961..5038
                     /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."
     CDS             5039..6121
                     /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"
     CDS             5162..6121
                     /codon_start=1
                     /gene="lacI"
                     /allele="Q"
                     /product="LacI"
                     /label=lacI
                     /note="Lac repressor"
                     /protein_id="ABO28848.1"
                     /translation="MAELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSR
                     ADQLGASVVVSMVERSGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPA
                     LFLDVSDQTPINSIIFSHEDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKY
                     LTRNQIQPIAEREGDWSAMSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLR
                     VGADISVVGYDDTEDSSCYIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVS
                     LVKRKTTLAPNTQTASPRALADSLMQLARQVSRLESGQ"
     gene            5162..6121
                     /gene="lacI"
                     /allele="Q"
                     /label=lacI
     protein_bind    6134..6155
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6170..6200
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6208..6224
                     /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     6232..6248
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             6244..81
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVGISLSTARCTNASGVR
                     QPSEAVVWLCRS"
     primer_bind     complement(6264..6280)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"