LOCUS       Exported                6646 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pMal-C2T, complete sequence.
ACCESSION   JF795283
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6646)
  AUTHORS   Arce-Rodriguez A, Durante-Rodriguez G, Platero R, Krell T, Calles B,
            de Lorenzo V.
  TITLE     The Crp regulator of Pseudomonas putida: evidence of an unusually 
            high affinity for its physiological effector, cAMP
  JOURNAL   Environ. Microbiol. 14 (3), 702-713 (2012)
  PUBMED    22040086
REFERENCE   2  (bases 1 to 6646)
  AUTHORS   Calles B.
  TITLE     Direct Submission
  JOURNAL   Submitted (08-APR-2011) Systems Biology Program, Centro Nacional de 
            Biotecnologia, Darwin, 3, Cantoblanco, Madrid 28049, Spain
REFERENCE   3  (bases 1 to 6646)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6646
                     /organism="Cloning vector pMal-C2T"
                     /mol_type="other DNA"
                     /db_xref="taxon:1163665"
     promoter        3..80
                     /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."
     regulatory      15..46
                     /regulatory_class="promoter"
                     /gene="lacI"
                     /note="lacIq promoter; overexpressing mutant"
     gene            15..46
                     /gene="lacI"
                     /label=lacI
     CDS             81..1163
                     /codon_start=1
                     /product="LacI"
                     /label=LacI
                     /protein_id="AFE02924.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     misc_feature    1406..3307
                     /label=malE expression cassette
                     /note="malE expression cassette"
     promoter        1406..1434
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     regulatory      1406..1433
                     /regulatory_class="promoter"
                     /note="Ptac promoter"
     protein_bind    1442..1458
                     /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             1528..2916
                     /codon_start=1
                     /product="MalE-lacZalpha fusion protein"
                     /label=MalE-lacZalpha fusion protein
                     /note="maltose binding protein (MBP) fusion"
                     /protein_id="AFE02922.1"
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQTNSSSNNNNNNNNNNLGLVPRGSEFGSSRVDLQ
                     ASLALAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWQLG
                     CFGG"
     CDS             1528..2628
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     misc_feature    2677..2694
                     /label=Region: thrombin recognition sequence
                     /note="Region: thrombin recognition sequence"
     CDS             2677..2694
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     misc_feature    2695..2734
                     /label=multiple cloning site
                     /note="multiple cloning site"
     misc_feature    2734..2913
                     /label=Region: lacZalpha
                     /note="Region: lacZalpha"
     primer_bind     complement(2736..2752)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     terminator      3101..3187
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     regulatory      3104..3307
                     /regulatory_class="terminator"
                     /note="rrnB T1T2 transcription terminators"
     terminator      3279..3306
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        3326..3417
                     /gene="bla"
                     /label=AmpR promoter
     CDS             3418..4278
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /note="confers resistance to ampicillin"
                     /protein_id="AFE02921.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERSPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            3418..4278
                     /gene="bla"
                     /label=bla
     rep_origin      complement(4319..4832)
                     /direction=LEFT
                     /label=M13 origin of replication
                     /note="M13 origin of replication"
     rep_origin      complement(4320..4833)
                     /direction=LEFT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     rep_origin      4943..5531
                     /note="pMB1 origin of replication (RNAII -35 to RNA/DNA 
                     switch point)"
     rep_origin      4944..5532
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    5718..5860
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(5962..6153)
                     /codon_start=1
                     /gene="rop"
                     /product="ROP protein"
                     /label=rop
                     /protein_id="AFE02923.1"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     gene            complement(5962..6153)
                     /gene="rop"
                     /label=rop