Yeast artificial chromosomes

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Yeast artificial chromosomes (YACs) are synthetic double stranded linear constructs containing the elements necessary for replication as independent chromosomes in yeast. These elements are:

  • an autonomous replication sequence (ARS): ARS1, chromosome III ARS, ARSH4
  • a centromere: CEN4, CEN6: centromeres consist of three centromere determining elements, CDE I, CDE II, and CDE III.
  • a telomeric sequence at each end

Typically the chromosome also contains a selection marker such as TRP1, Lys2 or Ura3.

Minimal size for a YAC is between 50kb and 100kb, while maximum sizes are 1Mb to 3Mb.

A common tool for constructing YACs is a shuttle plasmid such as pYAC4 which replicates in E. coli, has a multiple cloning site, and a pair of telomeres which can be cleaved to form a linear fragment. Available as an E.coli plasmid ATCC 67379, sequence at U01086.

There are two common centromere sequences, CEN4 and CEN6. CEN4 is found in most yeast centromere-containiing vectors, such as pYAC4. These vectors typically use ARS1 sequences.

The pRS313- pRS316 plasmids use the CEN6 + ARSH4 cassette (Sikorski89).

Yeast strains

  • AB1380 (MATa ura3-52 trp1 lys2-1 ade2-1 can1-100 his5) (Burke87) ATCC 204682
  • J57D (MATa, ura3-52, trp1 ade2-101 can1-100 leu2-3, 112 his3-6) (Haldi96)
    • claimed to be 2-3x more efficient at large insert cloning than AB1380
  • W303-1a (MATa leu2-3,112 trp1-1 can1-100 ura3-1 ade2-1 his3-11,15 ybp1-1) Rothstein notes (see Veal03) Open Biosystems: YSC1058
  • YPH500 (MATα ura3-52 lys2-801_amber ade2-101_ochre trp1-Δ63 his1-Δ200 leu2-Δ1) ATCC:204680 (Sikorski89)
  • S288C (MATα SUC2 gal2 mal mel flo1 flo8-1 hap1) ATCC: 204508 (Mortimer86)


YAC Plasmids

  • pYAC4 (Burke87) has Sup4 for ade2 selection, interrupted by the cloning site (EcoRI), ura3, a pair of facing telomeres surrounding a his3 gene (linearized and cut out with BamHI), a pBR322 based E. coli replication and ampR region, a barely functional TRP1 gene, ARS1 and CEN4. Sequence at Genbank U01086
  • pJS97 / pJS98 plasmids (Shero91) are available at ATCC 77191 as a two-plasmid kit. Each plasmid supplies one chromosome end.
    • pJS97 has CEN4 and ARSH4, URA3 gene, Sup11 tRNA for ade2 selection, a pUC19 derived E. coli replication region, including ampR, and a telomere.
    • pJS98 has ARSH4, a functional TRP1 gene, a pUC19 derived E. coli replication region including ampR, and a telomere.
  • pCGS966 (Smith90, Smith92, Moir93) has ARS1 on both arms, Gal inducible extra copy production, NeoR for mammalian expression and a functional promoter for the Trp gene, unlike pYAC4.

References

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  1. Error fetching PMID 2996783: [Hieter85]
  2. Error fetching PMID 3537685: [Mann86]
  3. Error fetching PMID 3519363: [Moritmer86]
  4. Error fetching PMID 3033825: [Burke87]
  5. Error fetching PMID 2552293: [Cottarel89]
  6. Error fetching PMID 2542900: [Anand89]
  7. Error fetching PMID 2659436: [Sikorski89]
  8. Error fetching PMID 2189412: [Sgaramella90]
  9. Error fetching PMID 2236036: [Smith90]
  10. Error fetching PMID 2276741: [Wells90]
  11. Error fetching PMID 2196439: [VanHouten90]
  12. Error fetching PMID 2091693: [McCormick90]
  13. Error fetching PMID 2091697: [Burke90]
  14. Error fetching PMID 2071158: [Shero92]
  15. Error fetching PMID 2005791: [Burke91]
  16. Error fetching PMID 2045094: [Connelly91]
  17. Error fetching PMID 1336106: [Smith92]
  18. Error fetching PMID 1620611: [Ragoussis92]
  19. Error fetching PMID 1427849: [deBruin92]
  20. Error fetching PMID 1406623: [Deshpande92]
  21. Error fetching PMID 1536007: [Marahrens92]
  22. Error fetching PMID 1544568: [Christianson92]
  23. Error fetching PMID 8462878: [Moir93]
  24. Spencer F, Ketner G, Connelly C, and Hieter P. Targeted recombination-based cloning and manipulation of large DNA segments in yeast. Methods: A companion to methods in enzymology 1993; 5, 161-175 (no pubmed entry)

    [Spencer93]

  25. Error fetching PMID 8163163: [Kuhn94]
  26. Error fetching PMID 7959756: [Hugerat94]
  27. Error fetching PMID 7959757: [Spencer94]
  28. Error fetching PMID 8524833: [Hamer95]
  29. Error fetching PMID 8841544: [Mahmood95]
  30. Error fetching PMID 8880145: [Mahmood96]
  31. Error fetching PMID 8854865: [Haldi96]
  32. US Patent 5776745 Ketner et al. 1998

    [Ketner98]

  33. Error fetching PMID 10087193: [Kooprina99]
  34. Error fetching PMID 10954614: [Cocchia00]
  35. Error fetching PMID 12743123: [Veal03]
All Medline abstracts: PubMed HubMed
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