GFP-MYO5C
(Plasmid #135404)

• Purpose: Expresses GFP-tagged human MYO5C in mammalian cells.
• Depositing Lab: Richard Cheney
• Publication: Jacobs et al Mol Biol Cell. 2009 Nov;20(21):4471-88. doi: 10.1091/mbc.e08-08-0865. Epub 2009 Sep 9.

Backbone

• Vector backbone: pEGFP-C2
• Backbone manufacturer: Clontech
• Backbone size w/o insert (bp): 4735
• Vector type: Mammalian Expression
• Selectable markers: Neomycin (select with G418)

Growth in Bacteria

• Bacterial Resistance(s): Kanamycin, 50 μg/mL
• Growth Temperature: 37°C
• Growth Strain(s): DH5alpha
• Copy number: High Copy

Gene/Insert

• Gene/Insert name: MYO5C
• Species: H. sapiens (human)
• Mutation: Relative to the GenBank MYO5C cDNA sequence reported in AF272390 and based on its numbering, the MYO5C sequence in GFP-MYO5C differs at 2 sites (AF272390>GFP-MYO5C): t907c (silent) c1584t (P552L) The human genome sequence for MYO5C also contains an L at aa 552, so the L in the GFP-MYO5C plasmid appears to be correct. Note: downstream of the natural MYO5C stop codon (between the SalI and BamHI sites), the GFP-MYO5C plasmid contains 42 nucleotides from the TOPO-TA vector used to assemble the plasmid.
• Entrez Gene: MYO5C
• Promoter: CMV
• Tag / Fusion Protein:
  • EGFP (N terminal on insert)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: XhoI (unknown if destroyed)
• 3′ cloning site: BamHI (unknown if destroyed)
• 5′ sequencing primer: EGFP-C

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
• Industry Terms:
  • Not Available to Industry

Depositor Comments

The official HUGO gene symbol for the human myosin-Vc heavy chain gene is MYO5C, so we recommend that MYO5C be used to denote the MYO5C gene, plasmids, and protein.

The GFP-MYO5C mammalian expression plasmid (9972 bp) was made by Damon T. Jacobs in the Richard E. Cheney lab and consists of nucleotides 20-5252 of the human myosin-Vc heavy chain GenBank sequence AF272390 inserted into the XhoI and BamH1 sites of the multiple cloning site in the pEGFP-C2 plasmid. The GFP is fused to the N-terminus of the full-length MYO5C heavy chain (aa 1-1742). The GFP-MYO5C plasmid and its localization to secretory granules was first described in Jacobs, DT, Weigert, R, Grode, KD, Donaldson, JG, and Cheney, RE, Myosin-Vc is a molecular motor that functions in secretory granule trafficking, Mol. Biol. Cell 20:4471-4488, 2009. Myosin-Vc is expressed in exocrine secretory tissues such as breast, prostate, pancreas, salivary, and lacrimal glands (Rodriguez, OC, and Cheney, RE, Human myosin-Vc is a novel class V myosin expressed in epithelial cells J Cell Sci 115:991-1004, 2001). Prolonged high-level expression should be avoided because under these conditions full-length GFP-MYO5c sometimes forms large puncta similar to those induced by overexpressing the MYO5C tail as a dominant negative. The pEGFP-C2 backbone contains a neomycin resistance cassette, so use Kan plates for bacterial selection and G418 for eukaryotic selection.