Construction of trophoblastic cell line HPT-8/pSilencer4.1-HtrA1 with silenced expression of HtrA1

VernacularTitle:HtrA1表达被沉默的滋养细胞株HPT-8/pSilencer4.1-HtrA1的构建
Author: Lu ZONG ; Wenli GOU ; Yulin GUO
Publication Type:Journal Article
Keywords: trophoblastic cell; HPT-8; HtrA1; plasmid
From: Journal of Xi'an Jiaotong University(Medical Sciences) 2015;(6):749-752,757
CountryChina
Language:Chinese
Abstract: Objective To construct the plasmid pSilencer4.1/HtrA1 and stably transfect human trophoblastic cell line HPT-8.Methods We detected the expression of HtrA1 in cell line HPT-8 with immunohistochemical SABC,constructed the plasmid pSilencer4.1/HtrA1,transfected human trophoblastic cell line HPT-8 with plasmid pSilencer4.1/HtrA1 and negative plasmid pSilencer4.1, and observed the cell expression after transfection. Results About 90% of cellular cytoplasm of HPT-8 was dyed brown while the nucleus was negatively stained. Selective concentration of G418 was 200 μg/mL for HPT-8 to be transfected pSilencer4.1/HtrA1.With RT-PCR and Western blot methods,the expression of HPT-8 transfected plasmid Psilence4.1/HtrA1 was significantly downregulated compared with that of cells with negative plasmid and normal cells (P < 0.05 ).There was no significant difference in absorbance value between blank plasmid group and HPT-8 group (P >0.05).Conclusion We successfully constructed stable trophoblastic cell line HPT-8/ pSilencer4.1-HtrA1 with silenced expression of HtrA1,which lays foundation for further research.