Construction of Rho GDIa-sgRNAs plasmids by clustered regularly interspaced short palindromic repeats/associated protein 9 and the effect on migration of Hepa 1-6 cell line in mouse
10.16098/j.issn.0529-1356.2021.01.008
- Author:
Ming-Juan QU
1
;
Yan-Min LI
1
;
Jing XIE
1
;
Miao-Miao QIN
1
;
Jing WANG
1
;
Ju-Hua ZHOU
1
Author Information
1. School of Life Sciences, Ludong University
- Publication Type:Journal Article
- Keywords:
Clustered regularly interspaced short palindromic repeats/associated protein 9;
Hepatoma Hepal-6 cell line;
Mouse;
PX458 plasmid;
Rho guanine dissociation inhibitor α;
RT-PCR;
Single guide RNA
- From:
Acta Anatomica Sinica
2021;52(1):55-59
- CountryChina
- Language:Chinese
-
Abstract:
Objective To construct the recombinant plasmids of knocking down Rho guanine dissociation inhibitor α (GDIα ) gene by using clustered regularly interspaced short palindromic repeats/associated protein 9 (CRISPR/Cas9) technique, and investigate the effect of Rho GDIα interference on the migration of Hepa 1-6 cells of mouse in order to provide the method of prevention and treatment of liver cancer. Methods To construct and identify the PX458-Rho GDIα-single guide (sg) RNAs by using CRISPR/Cas9 technique. And the Hepa 1-6 cells were transfected by liposomes with PX458-Rho GDIα-sgRNAs for 48 hours respectively, and cells treated with PX458 plasmids were used as control. The migration ability of Hepa 1-6 was checked by wound healing assay and Transwell assay, respectively. Results The expression of Rho GDIα was depressed in group of PX458-Rho GDIα-sgRNAl transfection which was detected by using RT-PCR. The migration distance of Hepa 1-6 in PX458-Rho GDIα-sgRNAl transfection group was significantly promoted comparing with the control group which was transfected with PX458 only, and the cell number of PX458-Rho GDIα-sgRNAl group was more than that in control group by using transwell assay, indicating concluded that knocking down of Rho GDIα promoted the migration ability of Hepal-6 cells. Conclusion The result is explicit that in vivo, Rho GDIα may inhibit the migration of Hepal-6 partially. Overexpression of Rho GDIα might be used as an important method to prevent the metastasize of carcinoma.
