Patent ID: 11866703
Assignee: LANZHOU VETERINARY RESEARCH INSTITUTE, CHINESE ACADEMY OF AGRICULTURAL SCIENCES
Field: Biotechnology (Chemistry)
Classification: CPC C | IPC C

Claim 2:
3. The knockout method according to claim 2, further comprising constructing the pEtNMT::DHFR plasmid before step a, comprising:
i. with a pSAG1::Cas9-U6::sgUPRT plasmid as a template, conducting amplification using a CRISPR-EtNMT-F primer and a CRISPR-R primer to obtain an amplification product;
ii. subjecting the amplification product to KLD reaction to obtain a reaction product;
iii. transforming the reaction product into competent cells; and
iv. extracting a resulting plasmid to obtain the pCRISPR::EtNMT plasmid; wherein
the CRISPR-EtNMT-F primer has the nucleotide sequence shown in SEQ ID NO: 1; and
the CRISPR-R primer has the nucleotide sequence shown in SEQ ID NO: 2.