pG23A
(Plasmid #102884)

• Purpose: Plasmid used for the C-terminal tagging of Cdc14 with mCherry and the auxin-inducible degron in yeast.
• Depositing Lab: Matthias Heinemann
• Publication: Papagiannakis et al Mol Cell. 2017 Jan 19;65(2):285-295. doi: 10.1016/j.molcel.2016.11.018. Epub 2016 Dec 15.

Backbone

• Vector backbone: HO-poly-KanMX4-HO
• Backbone manufacturer: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC55758/
• Backbone size w/o insert (bp): 2531
• Total vector size (bp): 5849
• Vector type: Yeast Expression
• Selectable markers: NatMX (select with Nourseotrichin)

Growth in Bacteria

• Bacterial Resistance(s): Ampicillin, 100 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: Cdc14/mCherry-AID(71-114)-NatMX
• Alt name: OAF3
• Species: S. cerevisiae (budding yeast)
• Insert Size (bp): 2328
• Entrez Gene: CDC14 (a.k.a. YFR028C, OAF3)
• Promoter: Native promoter
• Tags / Fusion Proteins:
  • mCherry (C terminal on insert)
  • AID(71-114) (C terminal on insert)

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: ATGACGAATCCATGCAGG
• 3′ sequencing primer: ggtcacagcttgtctgtaa

Resource Information

• Supplemental Documents:
  • G23A.txt
• A portion of this plasmid was derived from a plasmid made by: The AID(71-114) sequence was retrieved from: Morawska, M., and Ulrich, H.D. (2013). An expanded tool kit for the auxin-inducible degron system in budding yeast. Yeast 30, 341–351. mCherry was retrieved from the pBS35 plasmid.

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Takara Bio Limited Use Label License (formerly Clontech)
• Industry Terms:
  • Not Available to Industry

Depositor Comments

This plasmid can be used to dynamically and conditionally deplete Cdc14 in yeast.

A characterization guide can be found here:
https://www.nature.com/articles/s41598-017-04791-6

Detailed description of plasmid construction can be found in the Supplemental Experimental Procedures of the original paper:
http://www.cell.com/molecular-cell/pdfExtended/S1097-2765(16)30726-2