lentiCRISPRv2-sgRNAs-del-CDK5RAP2-SVA-BFP
(Plasmid #202824)

• Purpose: Lentiviral vector modified to express two sgRNAs that delete the intronic SVA within the CDK5RAP2 gene with EF-1apha for Cas9 and BFP expression
• Depositing Lab: Matthew Anderson
• Publication: Nadler et al Commun Biol. 2023 Mar 30;6(1):347. doi: 10.1038/s42003-023-04683-8.

Backbone

• Vector backbone: modified version of 98291
• Modifications to backbone: This vector was modified from Addgene 98291. The hygromycin resistance gene was excised by digestion with PshAI and MluI. T2A and blue fluorescent protein (BFP) were inserted in frame after the MluI site. Two sgRNAs were then cloned in tandem, each with their U6 promoter into digested lentiCRISPRv2-BFP vector (KpnI and NheI) using NEBuilder HiFi DNA Assembly(NEB).
• Vector type: Mammalian Expression, Lentiviral

Growth in Bacteria

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

Gene/Insert

• Gene/Insert name: two sgRNAs (each with a U6 promoter) to delete SVA within CDK5RAP2 gene
• gRNA/shRNA sequence: see map
• Species: H. sapiens (human)
• Promoter: U6 - two

Cloning Information

• Cloning method: Restriction Enzyme
• 5′ cloning site: KpnI (not destroyed)
• 3′ cloning site: NheI (not destroyed)
• 5′ sequencing primer: whole plasmid sequencing
• 3′ sequencing primer: whole plasmid sequencing

Terms and Licenses

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