PE-SUMO CRYGS Q16C
(Plasmid #217674)

• Purpose: For expression of SUMO tagged human gamma S crystallin with Q16C mutation to act as a biological control for ligation experiment
• Depositing Lab: Larry David
• Publication: Completion of previous SUMO tagged human crystallin submission plus plasmids for native chemical ligation to produced gamma S crystallin (unpublished)

Backbone

• Vector backbone: PE-SUMO
• Backbone manufacturer: LifeSensors
• Backbone size w/o insert (bp): 5761
• Total vector size (bp): 6269
• Modifications to backbone: None
• Vector type: Bacterial Expression

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: CRYGS
• Species: H. sapiens (human)
• Mutation: Glutamine 16 changed to Cysteine
• GenBank ID: CRYGS
• Entrez Gene: CRYGS (a.k.a. CRYG8, CTRCT20)
• Promoter: T7
• Tags / Fusion Proteins:
  • 6XHis (N terminal on backbone)
  • SUMO (N terminal on backbone)

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: Bsal (unknown if destroyed)
• 3′ cloning site: Xbal (unknown if destroyed)
• 5′ sequencing primer: T7
• 3′ sequencing primer: T7 term

Resource Information

• A portion of this plasmid was derived from a plasmid made by: DNASU HsCD00303433

Terms and Licenses

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

Depositor Comments

Expresses well in a soluble state and tags are removed efficiently using ULP-1 treatment. N-terminal methionine not included so it mirrors the mature sequence in the lens that has the methionine removed. This Q16C mutant of gS was used as a control to compare to the same protein created using native chemical ligation. The cysteine 16 was added to act as a ligation site to join synthetic peptide 1-15 to the N-terminus in order to place isoaspartate residues into the protein at positions 12 and 14 to investigate the impact on protein structure.