LOCUS       Exported                4680 bp ds-DNA    circular SYN 12-3-2014
DEFINITION  Compact bacterial vector encoding tRNA genes for rare Arg and Ile 
            codons, for ligation-independent cloning (LIC) and addition of a 
            C-terminal 6xHis tag.
ACCESSION   .
VERSION     .
KEYWORDS    pMCSG81
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4680)
  AUTHORS   Midwest Center for Structural Genomics
  TITLE     Direct Submission
COMMENT     For LIC, amplify the vector with the F and R primers and treat with 
            T4 DNA polymerase plus dCTP.
FEATURES             Location/Qualifiers
     source          1..4680
                     /organism="synthetic DNA construct"
                     /lab_host="Escherichia coli"
                     /mol_type="other DNA"
     terminator      26..73
                     /note="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(140..157)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /note="6xHis"
                     /translation="HHHHHH"
     CDS             complement(158..160)
                     /codon_start=1
                     /product="start codon"
                     /note="ATG"
                     /translation="M"
     RBS             169..174
                     /note="ribosome binding site"
     protein_bind    205..229
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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)."
     promoter        complement(230..248)
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     tRNA            complement(523..599)
                     /gene="argU"
                     /locus_tag="
                     "
                     /product="E. coli tRNA-Arg recognizing AGA and AGG"
                     /note="argU"
     tRNA            complement(869..944)
                     /gene="ileX"
                     /locus_tag="
                     "
                     /product="E. coli tRNA-Ile recognizing AUA"
                     /note="ileX"
     promoter        1084..1161
                     /gene="lacI"
                     /note="lacI promoter"
                     /note="
                     "
     CDS             1162..2244
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /note="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"
     rep_origin      complement(2673..3261)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3432..4292)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(4293..4396)
                     /gene="bla"
                     /note="AmpR promoter"