LOCUS       Exported                7109 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Recombinase expression vector pSH66, complete sequence.
ACCESSION   HQ412576
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7109)
  AUTHORS   Gueldener U, Heinisch J, Koehler GJ, Voss D, Hegemann JH.
  TITLE     A second set of loxP marker cassettes for Cre-mediated multiple gene
            knockouts in budding yeast
  JOURNAL   Nucleic Acids Res. 30 (6), E23 (2002)
  PUBMED    11884642
REFERENCE   2  (bases 1 to 7109)
  AUTHORS   Hegemann JH, Heick SB.
  TITLE     Delete and Repeat: A Comprehensive Toolkit for Sequential Gene 
            Knockout in the Budding Yeast Saccharomyces cerevisiae
  JOURNAL   Methods Mol. Biol. 765, 189-206 (2011)
  PUBMED    21815094
REFERENCE   3  (bases 1 to 7109)
  AUTHORS   Hegemann JH, Heick SB.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-OCT-2010) Lehrstuhl fur Funktionelle Genomforschung 
            der Mikroorganismen, Heinrich-Heine-Universitaet, 
            Universitatsstrasse 1, Duesseldorf, NRW 40225, Germany
REFERENCE   4  (bases 1 to 7109)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7109
                     /organism="Recombinase expression vector pSH66"
                     /mol_type="other DNA"
                     /db_xref="taxon:1007539"
     regulatory      187..584
                     /regulatory_class="promoter"
                     /note="TEF"
     gene            241..1360
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     promoter        241..584
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             585..1157
                     /codon_start=1
                     /gene="nat1"
                     /product="nourseothricin N-acetyltransferase"
                     /label=nat1
                     /note="nourseothricin resistance"
                     /protein_id="AEB90910.1"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     gene            585..1157
                     /gene="nat1"
                     /label=nat1
     regulatory      1158..1422
                     /regulatory_class="terminator"
                     /note="CYC1"
     terminator      1163..1360
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     rep_origin      complement(1481..1936)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     2081..2097
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2107..2125
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      complement(2134..2392)
                     /regulatory_class="terminator"
                     /note="CYC1"
     terminator      2144..2391
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     CDS             complement(2710..3741)
                     /codon_start=1
                     /gene="cre"
                     /product="recombinase"
                     /label=cre
                     /protein_id="AEB90911.1"
                     /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRTLDSETGAMVRLLE
                     DGD"
     gene            complement(2710..3741)
                     /gene="cre"
                     /label=cre
     primer_bind     complement(3839..3855)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      complement(3850..4317)
                     /regulatory_class="promoter"
                     /note="GAL1"
     promoter        complement(3870..4311)
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    4194..4311
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     promoter        complement(4335..4353)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4374..4390)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4398..4414
                     /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(4422..4452)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     rep_origin      4760..4832
     rep_origin      complement(4776..5364)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5535..6395)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /note="ampicillin resistance protein"
                     /protein_id="AEB90912.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(5535..6395)
                     /gene="bla"
                     /label=bla
     promoter        complement(6396..6500)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6534..7048
                     /label=CEN6/ARSH4
                     /note="CEN6/ARSH4"
     misc_feature    6537..7040
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"