CRISPR-Cas9 targeted knockout human colorectal carcinoma cell DEAF1 gene and specific sgRNA thereof

Abstract

The invention discloses a CRISPR/Cas9 targeted knockout human colorectal carcinoma cell DEAF1 gene and specific sgRNA thereof. The CRISPR/Cas9 targeted knockout human colorectal carcinoma cell DEAF1 gene and the specific sgRNA thereof are characterized in that: firstly, sgRNA of a second exon of the specific targeted DEAF1 gene is obtained and the base sequence of the sgRNA is shown as SEQ ID NO.1; secondly, the sgRNA of the DEAF1 gene is constructed into a lentiviral vector system, which contains Cas9 protein; finally, the CRISPR/Cas9 lentivirus containing the sgRNA is infected with human colorectal carcinoma cell HT-29 cell, so that a cell strain of which DEAF1 protein expression level is obviously reduced is obtained. The CRISPR/Cas9 targeted knockout human colorectal carcinoma cell DEAF1 gene disclosed by the invention has the advantages of simple operation steps, good sgRNA target ability and high cutting efficiency for the DEAF1 gene; in addition, the constructed CRISPR/Cas9 lentiviral vector system has the advantage of high knockout efficiency and can specifically knock out the DEAF1 gene to obtain the human colorectal carcinoma cells knocking out the DEAF1 gene, and therebya powerful tool is provided for further studying an action mechanism of DEAF1 in the colorectal carcinoma cells.

Description

Technical field [0001] The invention belongs to the field of genetic engineering, and more specifically relates to CRISPR / Cas9 targeted knockout of human intestinal cancer cell DEAF1 gene and its specific sgRNA. Background technique [0002] RBP-J interaction and microtubule-associated protein (DEAF1; C12orf52) is a highly conserved protein with a protein size of 36kD, which has no significant homology with any other protein. DEAF1 can interfere with Notch and RBP-J-mediated transcription. DEAF1 quickly shuttles between the nucleus and the cytoplasm, and most importantly, it mediates the nuclear export of RBP-J tubulin fibers. DEAF1, as a new RBP-J interaction protein, down-regulates Notch by interfering with the transcription factor RBP-J. Notch genes encode a class of highly conserved cell surface receptors that regulate the development of cells from sea urchins to humans. Notch signal affects many processes of normal cell morphogenesis, including the specialization of pluri...

AI Technical Summary

Problems solved by technology

At present, researchers have done a lot of research on the pathogenesis of colorectal cancer, and have discovered many pathways and mechanisms involved in the progression of colorectal cancer, such as activation of proto-oncogenes, inactivation of

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