pMS81(rpl-28p::mKate2::unc-54 3'UTR::LoxP::rps-0p::HYGR∆)
(Plasmid #154840)

• Purpose: Insertion of rpl-28p::mKate2::unc-54 3'UTR into a split hygromycin landing pad. Can be modified to insert a different gene(s) of interest.
• Depositing Lab: Patrick Phillips
• Publication: Stevenson et al G3 (Bethesda). 2020 Aug 19. pii: g3.120.401400. doi: 10.1534/g3.120.401400.

Backbone

• Vector backbone: pUC57
• Backbone size w/o insert (bp): 2470
• Total vector size (bp): 6176
• Vector type: Worm Expression, Cre/Lox, CRISPR

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: homology arm:rpl-28p::mKate2::unc-54 3'UTR::LoxP::rps-0p::HYGR∆
• Species: C. elegans (nematode), Synthetic
• Mutation: HYGR∆ encodes aa 1-226

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: caggaaacagctatgac (M13-rev)
• 3′ sequencing primer: Ctatgcggcatcagagcagatt

Resource Information

• Supplemental Documents:
  • pMS81.gb
• A portion of this plasmid was derived from a plasmid made by: rpl-28p is from pBCN39-R4R3. mKate2 coding sequence and unc-54 3’UTR is from pDD285. The hygromycin resistance gene fragment was modified from pCFJ1663. All three plasmids were obtained from Addgene. All remaining insert sequences were from C. elegans genomic DNA or were synthesized for our lab.

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • Evrogen Limited Use Label License for FPs
• Industry Terms:
  • Not Available to Industry

Depositor Comments

The LoxP site has a point mutation that in our hands does not noticeably impact function.

The rpl-28p::mKate2::unc-54 3'UTR transgene is flanked by restriction sites to facilitate removal and replacement of this gene.

Additional protocol information can be found at: github.com/phillips-lab/SLP