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US 11,339,370 B2Granted
Cell line with METTL3 gene knocked out, its construction method and interference vector
AssigneeZhejiang University
InventorsXin Zong, Yizhen Wang, Jing Zhao
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Abstract
A pig intestinal tract epithelium cell line with METTL3 gene knocked out and a construction method therefor. A gene interference vector for METTL3 form a pig is also provided.
Claims3
A method for constructing METTL3 gene knockout cell line, comprising:
- processing gene knockout by an m6A methyltransferase gene Mettl3 to obtain a pig intestinal epithelial cell line which is capable of tolerating LPS (lipopolysaccharide) stimulation led fatty acid absorption disorder;
- comprising steps of:
- (1) designing shRNA according to the m6A methyltransferase gene mettl3,
- wherein sequence of the mettl3 is shown in SEQ ID No.1, and sequence of the shRNA is shown in SEQ ID No.2 and SEQ ID No.3;
- (2) phosphorylating m6A methyltransferase gene mettl3 shRNA;
- (3) connecting the phosphorylated mettl3 shRNA to Lentivirus Expression Vector to obtain a mettl3 shRNA lentivirus expression vector;
- (4) enveloping the mettl3 shRNA lentivirus expression vector obtained in step (3) with 293A cells;
- (5) infecting the mettl3 shRNA lentivirus obtained in step (4) with intestinal epithelial cells, and then by puromycin, screening and obtaining the intestinal epithelial cells which are genetically modified to knock out the m6A methyltransferase gene mettl3.
5 more elements
Dependencies are inferred from the claim text.