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ID   PCGS990    preliminary; circular DNA; SYN; 14000 BP.
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AC   ATCC77417;
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DT   01-DEC-1994 (Rel. 10, Created)
DT   01-JUL-1995 (Rel. 12, Last updated, Version 1)
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DE   Saccharomyces/E.coli plasmid vector pCGS990 - incomplete.
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KW   cloning vector.
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OS   Cloning vector
OC   Artificial sequences; Cloning vehicles.
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RN   [1]
RC   pCGS990 from pBR322 & LYS2 gene
RA   Smith D.R., Smyth A.P., Strauss W.M., Moir D.T.;
RT   "Incorporation of copy-number control elements into yeast
RT   artificial chromosomes by targeted homologous recombination";
RL   Mamm. Genome 4:141-147(1993).
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CC   Conversion vector to insert copy number control elements into
CC   existing pYAC4-derived clones by targeted homologous recombination.
CC   pCGS990 should be linearized with SalI before transformation of the
CC   YAC-carrying strain and Lys+ clones selected. Homologous
CC   recombination between pCGS990 and the target YAC results in
CC   replacement of the centric pYAC4 arm with sequences from pCGS990.
CC   Recombinant clones will be Trp-.
CC   Approximately 0.5-2.5 percent of transformants will have the desired
CC   phenotype (Lys+, Ura+, Trp-).  These clones may be grown in liquid MST
CC   medium for 2-5 days to increase the YAC copy number.
CC   The cis-acting elements necessary for amplification are the thymidine
CC   kinase gene and a conditional CEN4 regulated by GAL1.
CC   The order of the major features in the plasmid is: BamHI -
CC   Tetrahymena telomere - ampR - thymidine kinase -
CC   EcoRI - LYS2 - ARS1 - GAL1 promoter - CEN4 - ClaI - SalI. [1]
CC   Medium is LB plus ampicillin (ATCC medium number 1227) 37C
CC   Deposited by: D.T.Moir
CC   NM (pCGS990)
CC   CM (no)
CC   NA (ds-DNA)
CC   TP (circular)
CC   ST ()
CC   TY (plasmid)
CC   SP (ATCC)
CC   HO (E.coli HB101)(Saccharomyces cerevisiae)
CC   CP ()
CC   FN (cloning)(shuttle)
CC   SE ()
CC   PA ()
CC   BR ()
CC   OF ()
CC   OR ()
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FH   Key             Location/Qualifiers
FH
FT   misc_feature    0..0
FT                   /note="1. pBR322 4361bp, ori/amp gene
FT                   2. yeast, LYS2 gene
FT                   -> pCGS990 14000bp"
FT   rep_origin      0..0
FT                   /note="ORI E. coli pMB1 (ColE1 and pBR322)"
FT   rep_origin      0..0
FT                   /note="ORI yeast ARS1"
FT   misc_feature    0..0
FT                   /note="yeast centromere CEN4"
FT   CDS             0..0
FT                   /note="ANT E. coli beta-lactamase gene (bla)
FT                   ampicillin resistance gene (apr/amp)"
FT   CDS             0..0
FT                   /note="ANT yeast LYS2 gene"
FT   promoter        0..0
FT                   /note="PRO yeast GAL1 gene"
FT   promoter        0..0
FT                   /note="PRO HSV tk gene"
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SQ   Sequence 1 BP; 0 A; 0 C; 0 G; 0 T; 1 other;
     n
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