pCvGAPDH-NanoLuc^opt
(Plasmid #255781)

• Purpose: Expresses NanoLuc luciferase (codon optimized for Chromera) in Chromera velia and Toxoplasma gondii
• Depositing Lab: Ke Hu
• Publication: Tengganu et al J Cell Sci. 2026 Jan 1;139(1):jcs.264400. doi: 10.1242/jcs.264400. Epub 2026 Jan 13.

Backbone

• Vector backbone: pBluescript
• Backbone size w/o insert (bp): 2931
• Total vector size (bp): 5295
• Vector type: Bacterial Expression ; Chromera velia, Toxoplasma gondii

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: Cv β-tubulin intron 1 + GAPDH Kozak + NanoLuc
• Alt name: NanoLuc luciferase
• Insert Size (bp): 628
• Mutation: codon optimized for Chromera velia
• Promoter: putative C. velia GAPDH promoter (1090 bp)

Cloning Information

• Cloning method: Gibson Cloning
• 5′ sequencing primer: pBluescript KS
• 3′ sequencing primer: pBluescript SK

Terms and Licenses

• Academic/Nonprofit Terms:
  • UBMTA
  • NanoLuc Luciferase Limited Use Label License
• Industry Terms:
  • Not Available to Industry

Depositor Comments

Derived from pCvGAPDH-NanoLuc, a pBluescript-based vector containing the Chromera velia GAPDH promoter (1099 bp; VEuPathDB: Cvel_5775), NanoLuc CDS, and GAPDH 3′ UTR (632 bp). The pCvGAPDH-NanoLuc vector was digested with NsiI (∼200 bp upstream of the start codon within the promoter) and AflII (immediately downstream of the NanoLuc stop codon) to generate the backbone.

A synthetic gBlock (IDT) was assembled into this backbone by Gibson assembly to replace the excised region. The insert contains ~200 bp of the CvGAPDH promoter, the first intron of C. velia β-tubulin (VEuPathDB: Cvel_33153), and a NanoLuc CDS codon-optimized for C. velia, with ~20 bp overlaps matching the NsiI and AflII ends.

The β-tubulin intron lies within the 5′ UTR, with its 3′ end 9 bp upstream of the start codon, immediately preceding the Kozak sequence (gctcagaagATG) without disrupting the ATG.