LOCUS       Exported                6148 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pTRC-LIC, complete sequence.
ACCESSION   EF460847
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6148)
  AUTHORS   Loppnau P, Kozieradzki I, Arrowsmith C.
  TITLE     Expression vectors for high-throughput In-Fusion cloning
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6148)
  AUTHORS   Loppnau P, Kozieradzki I, Arrowsmith C.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-FEB-2007) Structural Genomics Consortium, University 
            of Toronto, 100 College Street, Toronto, ON M5G1L5, Canada
REFERENCE   3  (bases 1 to 6148)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6148
                     /organism="Expression vector pTRC-LIC"
                     /mol_type="other DNA"
                     /db_xref="taxon:435753"
     promoter        1..92
                     /gene="bla"
                     /label=AmpR promoter
     CDS             93..953
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /note="ampicillin resistance protein"
                     /protein_id="ABP57192.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPTAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            93..953
                     /gene="bla"
                     /label=bla
     rep_origin      1124..1712
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     rep_origin      1711
                     /label=colE1 from pBR322
                     /note="colE1 from pBR322"
     misc_feature    1898..2038
                     /label=bom
                     /note="basis of mobility region from pBR322"
     promoter        2224..2301
                     /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             2302..3384
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /protein_id="ABP57193.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     gene            2302..3384
                     /gene="lacI"
                     /label=lacI
     regulatory      3441..3686
                     /regulatory_class="promoter"
                     /note="trc from pKK233-2"
     promoter        3616..3645
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     regulatory      3616..3621
                     /regulatory_class="minus_35_signal"
                     /note="derived from trp promoter"
     regulatory      3639..3645
                     /regulatory_class="minus_10_signal"
                     /note="derived from lacUV5 promoter"
     protein_bind    3651..3671
                     /label=lac repressor protein binding site
                     /bound_moiety="lac repressor protein"
     protein_bind    3653..3669
                     /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)."
     misc_feature    3678..3692
                     /note="5-LIC sequence; add to upstream PCR primer"
     protein_bind    3720..3753
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        3768..4213
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             4214..5635
                     /codon_start=1
                     /gene="sacB"
                     /product="levansucrase"
                     /label=sacB
                     /note="Bacillus subtilis negative selection gene; converts 
                     sucrose to a toxic product; allows selection of recombinant
                     plasmids on media containing 5% sucrose"
                     /protein_id="ABP57194.1"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     gene            4214..5635
                     /gene="sacB"
                     /label=sacB
     misc_feature    5708..5722
                     /note="3-LIC sequence; add complement to downstream PCR 
                     primer"
     misc_feature    5723..6148
                     /note="derived from the rrnB operon; the entire operon is 
                     not contained in this vector"
     gene            5804..6129
                     /gene="rrnB"
                     /label=rrnB
     rRNA            5804..5923
                     /gene="rrnB"
                     /product="5S ribosomal RNA"
                     /label=rrnB
     terminator      5924..6010
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     regulatory      5927..5970
                     /regulatory_class="terminator"
                     /gene="rrnB"
                     /note="rrnB T1"
     regulatory      6102..6129
                     /regulatory_class="terminator"
                     /gene="rrnB"
                     /note="rrnB T2"
     terminator      6102..6129
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"