LOCUS       Exported               10495 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pLNMCL, complete sequence.
ACCESSION   KC999858
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10495)
  AUTHORS   Shahana S, Childers DS, Ballou ER, Bohovych I, Odds FC, Gow NA, 
            Brown AJ.
  TITLE     New Clox Systems for Rapid and Efficient Gene Disruption in Candida 
            albicans
  JOURNAL   PLoS ONE 9 (6), E100390 (2014)
  PUBMED    24940603
REFERENCE   2  (bases 1 to 10495)
  AUTHORS   Shahana S, Childers DS, Brown AJP.
  TITLE     Direct Submission
  JOURNAL   Submitted (01-MAY-2013) Microbiology, School of Medical Sciences, 
            Institute of Medical Sciences - Foresterhill, Aberdeen, 
            Aberdeenshire AB25 2ZD, United Kingdom
REFERENCE   3  (bases 1 to 10495)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..10495
                     /organism="Cloning vector pLNMCL"
                     /mol_type="other DNA"
                     /db_xref="taxon:1415790"
     CDS             complement(663..1757)
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKVDAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVDPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     promoter        complement(1770..2174)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(2474..2929)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     3074..3090
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3097..3115
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3148..3164
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    join(3178..3211,7597..7630)
                     /label=loxP
                     /note="loxP"
     protein_bind    3178..3211
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     terminator      3622..3819
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(3834..4406)
                     /codon_start=1
                     /product="NAT1"
                     /label=NAT1
                     /note="confers nourseothrycin resistance; obtained from 
                     pJK805"
                     /protein_id="AGZ63434.1"
                     /translation="MSTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDAGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     promoter        complement(4420..4763)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     regulatory      4827..6158
                     /regulatory_class="promoter"
                     /gene="pMET3"
                     /note="MET3 promoter"
     gene            4827..6158
                     /gene="pMET3"
                     /label=pMET3
     CDS             6186..6593
                     /codon_start=1
                     /product="Cre"
                     /label=Cre
                     /note="Cre recombinase"
                     /protein_id="AGZ63435.1"
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQAL"
     protein_bind    7597..7630
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     complement(7675..7691)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(7721..7739)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7760..7776)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7784..7800
                     /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)."
     promoter        complement(7808..7838)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     rep_origin      complement(8162..8750)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(8921..9781)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFFHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(9782..9886)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    9923..10426
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"