Effect Of HMGA2 Gene Knockout On Human Papillary Thyroid Carcinoma Cell Line TPC-1

Objective To study the effect of HMGA2 gene knockout on the invasion and metastasis of human papillary thyroid carcinoma cell line(Thyroid papillary carcinoma-1,TPC-1).Method 1.Construction of lentiviral vector p LV [2g RNA]-EGFP:T2A:Puro-U6 >{h HMGA2 [g RNA#A1] *}-U6 > {h HMGA2 [g RNA#A2] *}: the Dual-g RNA sequence targeting HMGA2 gene was designed,and the synthesized fragment was cloned into p LV [2g RNA]-EGFP:T2A:Puro-U6 vector.Construction of TPC-1 cell line with stable knockout of HMGA2 gene: the constructed recombinant plasmid and vector were packaged with lentivirus and infected with TPC-1 cells.HMGA2 knockout cell lines were screened by puromycin,and the knockout efficiency of HMGA2 m RNA in cells was detected by real-time fluorescence quantitative q PCR 3.The ability of cell migration was detected by scratch test and the ability of cell proliferation was observed by CCK-8 test in wild type(WT)group and HMGA2 gene knockout(KO-HMGA2)group.Result 1.After sequencing verification,successfully constructed p LV [2g RNA]-EGFP targeting HMGA2 gene: T2A:Puro-U6>{ h HMGA2 [g RNA#A1] *}-U6>{h HMGA2 [g RNA#A2]*} plasmid.2.After monoclonal screening,a TPC-1 cell line completely knocked out of HMGA2 was successfully obtained.No expression of HMGA2 m RNA was found in TPC-1 cells knocked out by HMGA2 by real-time fluorescence quantitative q PCR.3.The migration ability of the KO-HMGA2 group was significantly lower than that of the WT group,and the proliferation ability was significantly weaker than that of the WT group.Conclusion 1.The CRISPR/Cas9 gene editing system can quickly and effectively edit the HMGA2 gene of the human papillary thyroid carcinoma cell line TPC-1 2.The CRISPR/Cas9 gene editing system can be used to construct a stable cell line of the human papillary thyroid carcinoma cell line TPC-1knocked out of the HMGA2 gene.3.Knockout of HMGA2 gene may reduce the invasion and metastasis ability of human papillary thyroid carcinoma cell line TPC-1cells.